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  • 251. Gan, Haiyun
    et al.
    Yu, Chuanhe
    Devbhandari, Sujan
    Sharma, Sushma
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Han, Junhong
    Chabes, Andrei
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Remus, Dirk
    Zhang, Zhiguo
    Checkpoint Kinase Rad53 Couples Leading- and Lagging-Strand DNA Synthesis under Replication Stress2017Inngår i: Molecular Cell, ISSN 1097-2765, E-ISSN 1097-4164, Vol. 68, nr 2, s. 446-455Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The checkpoint kinase Rad53 is activated during replication stress to prevent fork collapse, an essential but poorly understood process. Here we show that Rad53 couples leading- and lagging-strand synthesis under replication stress. In rad53-1 cells stressed by dNTP depletion, the replicative DNA helicase, MCM, and the leading-strand DNA polymerase, Pol ε, move beyond the site of DNA synthesis, likely unwinding template DNA. Remarkably, DNA synthesis progresses further along the lagging strand than the leading strand, resulting in the exposure of long stretches of single-stranded leading-strand template. The asymmetric DNA synthesis in rad53-1 cells is suppressed by elevated levels of dNTPs in vivo, and the activity of Pol ε is compromised more than lagging-strand polymerase Pol δ at low dNTP concentrations in vitro. Therefore, we propose that Rad53 prevents the generation of excessive ssDNA under replication stress by coordinating DNA unwinding with synthesis of both strands.

  • 252.
    Ganai, Rais Ahmad
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Structural and biochemical basis for the high fidelity and processivity of DNA polymerase ε2015Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    DNA polymerase epsilon (Pol ε) is a multi-subunit B-family DNA polymerase that is involved in leading strand DNA replication in eukaryotes. DNA Pol ε in yeast consists of four subunits, Pol2, Dpb2, Dpb3, and Dpb4. Pol2 is the catalytic subunit and Dpb2, Dpb3, and Dpb4 are the accessory subunits. Pol2 can be further divided into an N-terminal catalytic core (Pol2core) containing both the polymerase and exonuclease active sites and a C-terminus domain. We determined the X-ray crystal structure of Pol2core at 2.2 Å bound to DNA and with an incoming dATP. Pol ε has typical fingers, palm, thumb, exonuclease, and N-terminal domains in common with all other B-family DNA polymerases. However, we also identified a seemingly novel domain we named the P-domain that only appears to be present in Pol ε. This domain partially encircles the nascent duplex DNA as it leaves the active site and contributes to the high intrinsic processivity of Pol ε.

    To ask if the crystal structure of Pol2core can serve as a model for catalysis by Pol ε, we investigated how the C-terminus of Pol2 and the accessory subunits of Pol ε influence the enzymatic mechanism by which Pol ε builds new DNA efficiently and with high fidelity. Pre-steady state kinetics revealed that the exonuclease and polymerization rates were comparable between Pol2core and Pol ε. However, a global fit of the data over five nucleotide-incorporation events revealed that Pol ε is slightly more processive than Pol2 core. The largest differences were observed when measuring the time for loading the polymerase onto a 3' primer-terminus and the subsequent incorporation of one nucleotide. We found that Pol ε needed less than a second to incorporate the first nucleotide, but it took several seconds for Pol2core to incorporate similar amounts of the first nucleotide.

    B-family polymerases have evolved an extended β-hairpin loop that is important for switching the primer terminus between the polymerase and exonuclease active sites. The high-resolution structure of Pol2core revealed that Pol ε does not possess an extended β-hairpin loop. Here, we show that Pol ε can processively transfer a mismatched 3' primer-terminus between the polymerase and exonuclease active sites despite the absence of a β-hairpin loop. Additionally we have characterized a series of amino acid substitutions in Pol ε that lead to altered partitioning of the 3'primer-terminus between the two active sites.

    In a final set of experiments, we investigated the ability of Pol ε to displace the downstream double-stranded DNA while carrying out DNA synthesis. Pol ε displaced only one base pair when encountering double-stranded DNA after filling a gap or a nick. However, exonuclease deficient Pol ε carries out robust strand displacement synthesis and can reach the end of the templates tested here. Similarly, an abasic site or a ribonucleotide on the 5'-end of the downstream primer was efficiently displaced but still only by one nucleotide. However, a flap on the 5'-end of the blocking primer resembling a D-loop inhibited Pol ε before it could reach the double-stranded junction. Our results are in agreement with the possible involvement of Pol ε in short-patch base excision repair and ribonucleotide excision repair but not in D-loop extension or long-patch base excision repair.

  • 253. Garbacz, Marta A.
    et al.
    Cox, Phillip B.
    Sharma, Sushma
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Lujan, Scott A.
    Chabes, Andrei
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Kunkel, Thomas A.
    The absence of the catalytic domains of Saccharomyces cerevisiae DNA polymerase ϵ strongly reduces DNA replication fidelity2019Inngår i: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 47, nr 8, s. 3986-3995Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The four B-family DNA polymerases α, δ, ϵ and ζ cooperate to accurately replicate the eukaryotic nuclear genome. Here, we report that a Saccharomyces cerevisiae strain encoding the pol2-16 mutation that lacks Pol ϵ's polymerase and exonuclease activities has increased dNTP concentrations and an increased mutation rate at the CAN1 locus compared to wild type yeast. About half of this mutagenesis disappears upon deleting the REV3 gene encoding the catalytic subunit of Pol ζ. The remaining, still strong, mutator phenotype is synergistically elevated in an msh6Δ strain and has a mutation spectrum characteristic of mistakes made by Pol δ. The results support a model wherein slow-moving replication forks caused by the lack of Pol ϵ's catalytic domains result in greater involvement of mutagenic DNA synthesis by Pol ζ as well as diminished proofreading by Pol δ during replication.

  • 254. Garcia-Aljaro, Cristina
    et al.
    Melado-Rovira, Silvia
    Milton, Debra L.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Blanch, Anicet R.
    Quorum-sensing regulates biofilm formation in Vibrio scophthalmi2012Inngår i: BMC Microbiology, ISSN 1471-2180, E-ISSN 1471-2180, Vol. 12, s. 287-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background: In a previous study, we demonstrated that Vibrio scophthalmi, the most abundant Vibrio species among the marine aerobic or facultatively anaerobic bacteria inhabiting the intestinal tract of healthy cultured turbot (Scophthalmus maximus), contains at least two quorum-sensing circuits involving two types of signal molecules (a 3-hydroxy-dodecanoyl-homoserine lactone and the universal autoinducer 2 encoded by luxS). The purpose of this study was to investigate the functions regulated by these quorum sensing circuits in this vibrio by constructing mutants for the genes involved in these circuits.

    Results: The presence of a homologue to the Vibrio harveyi luxR gene encoding a main transcriptional regulator, whose expression is modulated by quorum-sensing signal molecules in other vibrios, was detected and sequenced. The V. scophthalmi LuxR protein displayed a maximum amino acid identity of 82% with SmcR, the LuxR homologue found in Vibrio vulnificus. luxR and luxS null mutants were constructed and their phenotype analysed. Both mutants displayed reduced biofilm formation in vitro as well as differences in membrane protein expression by mass-spectrometry analysis. Additionally, a recombinant strain of V. scophthalmi carrying the lactonase AiiA from Bacillus cereus, which causes hydrolysis of acyl homoserine lactones, was included in the study.

    Conclusions: V. scophthalmi shares two quorum sensing circuits, including the main transcriptional regulator luxR, with some pathogenic vibrios such as V. harveyi and V. anguillarum. However, contrary to these pathogenic vibrios no virulence factors (such as protease production) were found to be quorum sensing regulated in this bacterium. Noteworthy, biofilm formation was altered in luxS and luxR mutants. In these mutants a different expression profile of membrane proteins were observed with respect to the wild type strain suggesting that quorum sensing could play a role in the regulation of the adhesion mechanisms of this bacterium.

  • 255. Garzón, J.
    et al.
    Rodríguez, R.
    Kong, Ziqing
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Chabes, Andrei
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Rodríguez-Acebes, S.
    Méndez, J.
    Moreno, S.
    García-Higuera, I.
    Shortage of dNTPs underlies altered replication dynamics and DNA breakage in the absence of the APC/C cofactor Cdh12017Inngår i: Oncogene, ISSN 0950-9232, E-ISSN 1476-5594, Vol. 36, nr 42, s. 5808-5818Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The APC/C-Cdh1 ubiquitin-ligase complex targets cell cycle regulators for proteosomal degradation and helps prevent tumor development and accumulation of chromosomal aberrations. Replication stress has been proposed to be the main driver of genomic instability in the absence of Cdh1, but the real contribution of APC/C-Cdh1 to efficient replication, especially in normal cells, remains unclear. Here we show that, in primary MEFs, acute depletion or permanent ablation of Cdh1 slowed down replication fork movement and increased origin activity. Partial inhibition of origin firing does not accelerate replication forks, suggesting that fork progression is intrinsically limited in the absence of Cdh1. Moreover, exogenous supply of nucleotide precursors, or ectopic overexpression of RRM2, the regulatory subunit of Ribonucleotide Reductase, restore replication efficiency, indicating that dNTP availability could be impaired upon Cdh1 loss. Indeed, we found reduced dNTP levels in Cdh1-deficient MEFs. Importantly, DNA breakage is also significantly alleviated by increasing intracellular dNTP pools, strongly suggesting that genomic instability is the result of aberrant replication. These observations highlight the relevance of APC/C-Cdh1 activity during G1 to ensure an adequate supply of dNTPs to the replisome, prevent replication stress and the resulting chromosomal breaks and, ultimately, suppress tumorigenesis.

  • 256. Gause, Maria
    et al.
    Hovhannisyan, Hayk
    Kan, Tatiana
    Institute of Gene Biology, Russian Academy of Sciences, Moscow, Russia.
    Kuhfittig, Steffi
    Mogila, Vladic
    Georgiev, Pavel
    hobo Induced rearrangements in the yellow locus influence the insulation effect of the gypsy su(Hw)-binding region in Drosophila melanogaster1998Inngår i: Genetics, ISSN 0016-6731, E-ISSN 1943-2631, Vol. 149, nr 3, s. 1393-1405Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The su(Hw) protein is responsible for the insulation mediated by the su(Hw)-binding region present in the gypsy retrotransposon. In the y2 mutant, su(Hw) protein partially inhibits yellow transcription by repressing the function of transcriptional enhancers located distally from the yellow promoter with respect to gypsy. y2 mutation derivatives have been induced by the insertion of two hobo copies on the both sides of gypsy: into the yellow intron and into the 5' regulatory region upstream of the wing and body enhancers. The hobo elements have the same structure and orientation, opposite to the direction of yellow transcription. In the sequence context, where two copies of hobo are separated by the su(Hw)-binding region, hobo-dependent rearrangements are frequently associated with duplications of the region between the hobo elements. Duplication of the su(Hw)-binding region strongly inhibits the insulation of the yellow promoter separated from the body and wing enhancers by gypsy. These results provide a better insight into mechanisms by which the su(Hw)-binding region affects the enhancer function.

  • 257.
    Gerpe, M.
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Kling, P.
    Berg, A. H.
    Olsson, P.-E.
    Arctic char (Salvelinus alpinus) metallothionein: cDNA sequence, expression, and tissue-specific inhibition of cadmium-mediated metallothionein induction by 17ß-estradiol, 4-OH-PCB 30, and PCB 1042000Inngår i: Environmental Toxicology and Chemistry, ISSN 0730-7268, E-ISSN 1552-8618, Vol. 19, nr 3, s. 638-645Artikkel i tidsskrift (Fagfellevurdert)
  • 258.
    Gharibyan, Anna
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Amyloids here, amyloids there…What’s wrong with them?2012Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Amyloid formation is inherent property of proteins which under certain circumstances can become a pathologic feature of a group of diseases called amyloidosis. There are about 30 known human amyloidosis and more than 27 identified proteins involved in these pathologies.  Besides these proteins, there are a growing number of proteins non-related to diseases shown to form amyloid-like structures in vitro, which make them excellent tools for studying amyloid formation mechanisms, physicochemical properties of different amyloid species and the nature of their influence on tissues and cells.  It is important to understand the mechanisms by which amyloids interact with different types of cells, as the leading hypothesis in amyloid field suggests that amyloids and especially their intermediate states are the main harmful, toxic species causing tissue and cell degeneration.

    Using de-novo synthesized protein albebetin as a model of amyloidogenic protein, we demonstrated that it forms amyloid-like structures under physiological conditions (pH 7 and 37°C). During aggregation it forms 2 different types of intermediate oligomers — cross-b sheet containing and lacking β-sheet oligomers. Only the former induces cellular toxicity in a dose dependent manner. Further aggregation leads to the formation of fully mature amyloid-like fibrils, which are not toxic to the cells during studied period of incubation.

    Another model protein in our studies was hen egg white lysozyme, which readily forms amyloid under denaturing conditions (pH 2,2 and 57°C). In contrast to albebetin and many other proteins reported in the literature, we showed that both oligomers and mature fibrils from hen lysozyme affect cell viability. Targeting different mechanisms involved in cellular death, we revealed that oligomers induce slow and apoptotic-like cell death, while mature fibrils cause rapid and mainly necrotic-like cellular death.   

    One of the important aspects of amyloid studies is to develop measures for inhibiting or re-directing the process of amyloid formation to abolish or neutralize toxic amyloid species. Among the agents having inhibitory or modulatory properties small, phenol containing molecules are widely studied. We investigated the effect of the novel nootropic drug noopept on amyloid formation process of α-synuclein, as this drug is a small dipeptide containing a phenol ring. We showed that noopept is able to modulate amyloid formation process by accelerating it to rapid conversion of α-synuclein into fully mature fibrils, thus eliminating the stage of population of toxic oligomeric species.  Using wide range of cytotoxicity assays we showed that amyloid-like fibrils formed in the presence of noopept have no cytotoxic properties.  As this medicine is becoming popular and freely available in some countries as a cognitive enhancer, neuroprotective and nootropic agent, further detailed investigations and clinical trials are needed to assess the safety and benefit of noopept in particular for the patients with amyloid related neurodegenerative diseases (such as Parkinson’s or Alzheimer’s diseases).    

    While in vitro models are useful to study some specific aspects of protein aggregation, their properties and effects on cell viability, it is very difficult or practically impossible to create an absolutely accurate model of in vivo situation. Therefore, it is important to turn to in vivo/ex vivo studies to relate the knowledge accumulated from in vitro studies to the real situation in the body.

    Using human brain hippocampus tissues from individuals with Alzheimer’s disease, we found that besides well-known and widely accepted main pathological hallmark — Ab peptide deposition, S100A9 and S100A8 pro-inflammatory calcium-binding proteins are also localized in the plaques and in surrounding tissues and very explicitly co-localized with Ab. Moreover, we found the presence of S100A9 within the neuronal cells, which has not been reported before and can be an important clue for understanding the mechanisms of neurodegeneration. In vitro cytotoxicity studies showed that S100A9 protein can efficiently induce cytotoxicity when added exogenously to the neuronal cell culture. These findings suggest that S100A8 and S100A9 proteins play an important role in Alzheimer’s pathology, and potentially can be candidates for the amyloid plaque formation and neurodegeneration. Whether they are associated with inflammatory processes underlying the early onset of disease or produced and accumulated as a consequence of A-beta induced pathology remain to be clarified.

    We found that Alzheimer’s disease is not the only pathology associated with A-beta and S100A9 deposition in a form of plaques. Immunohistochemical studies of an aortic valve surgically removed from a patient with aortic stenosis revealed plaque-like structures positively stained with A-beta and S100A9 proteins. These areas are also positively stained with fibril-specific antibodies as well as with Congo red, which also shows very distinct apple-green birefringence under the polarized light. Besides, there is intracellular localization and co-localization of both proteins in interstitial cells throughout the whole fibrous tissue of the valve. The presented case report is the first finding suggesting inflammatory protein S100A9 as well as A-beta peptide as potential candidates for amyloid formation in aortic stenosis valves.  We suggest that there is a specific interaction between A-beta and S100A9 during amyloid formation, which can be involved in amyloid-associated pathology in various tissues and organs in the body, which can potentially be caused by inflammatory processes, particularly by its chronic, long lasting forms.

  • 259.
    Gharibyan, Anna
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Narayana, Vinod
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Ankarcrona, Maria
    Karolinska Institute.
    Brännström, Thomas
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap.
    Olofsson, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Morozova-Roche, Ludmilla
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Emerging role of inflammatory S100A9 in Alzheimer’s disease amyloid growth and neurodegenerationManuskript (preprint) (Annet vitenskapelig)
  • 260.
    Gharibyan, Anna
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Narayana, Vinod
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Habib, Ahsan
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Sulniute, Rima
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Olofsson, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Henein, Michael
    Umeå universitet, Medicinska fakulteten, Institutionen för folkhälsa och klinisk medicin, Medicin.
    Morozova-Roche, Ludmilla
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Inflammatory S100A9 and Aβ amyloids in heart valve of patient with aortic stenosisManuskript (preprint) (Annet vitenskapelig)
  • 261. Gisterå, Anton
    et al.
    Robertson, Anna-Karin L
    Andersson, John
    Ketelhuth, Daniel FJ
    Ovchinnikova, Olga
    Nilsson, Stefan K
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Fysiologisk kemi.
    Lundberg, Anna M
    Li, Ming O
    Flavell, Richard A
    Hansson, Göran K
    Transforming growth factor-beta signaling in T cells promotes stabilization of atherosclerotic plaques through an interleukin-17-dependent pathway2013Inngår i: Science Translational Medicine, ISSN 1946-6234, E-ISSN 1946-6242, Vol. 5, nr 196, s. 196ra100-Artikkel i tidsskrift (Fagfellevurdert)
  • 262. Gnanasundram, Sivakumar Vadivel
    et al.
    Fåhraeus, Robin
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Translation Stress Regulates Ribosome Synthesis and Cell Proliferation2018Inngår i: International Journal of Molecular Sciences, ISSN 1422-0067, E-ISSN 1422-0067, Vol. 19, nr 12, artikkel-id 3757Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Ribosome and protein synthesis are major metabolic events that control cellular growth and proliferation. Impairment in ribosome biogenesis pathways and mRNA translation is associated with pathologies such as cancer and developmental disorders. Processes that control global protein synthesis are tightly regulated at different levels by numerous factors and linked with multiple cellular signaling pathways. Several of these merge on the growth promoting factor c-Myc, which induces ribosome biogenesis by stimulating Pol I, Pol II, and Pol III transcription. However, how cells sense and respond to mRNA translation stress is not well understood. It was more recently shown that mRNA translation stress activates c-Myc, through a specific induction of E2F1 synthesis via a PI3K delta-dependent pathway. This review focuses on how this novel feedback pathway stimulates cellular growth and proliferation pathways to synchronize protein synthesis with ribosome biogenesis. It also describes for the first time the oncogenic activity of the mRNA, and not the encoded protein.

  • 263. Goldberg, Emily L.
    et al.
    Asher, Jennifer L.
    Molony, Ryan D.
    Shaw, Albert C.
    Zeiss, Caroline J.
    Wang, Chao
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Morozova-Roche, Ludmilla A.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Herzog, Raimund I.
    Iwasaki, Akiko
    Dixit, Vishwa Deep
    beta-Hydroxybutyrate deactivates Neutrophil NLRP3 inflammasome to relieve gout flares2017Inngår i: Cell reports, ISSN 2211-1247, E-ISSN 2211-1247, Vol. 18, nr 9, s. 2077-2087Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Aging and lipotoxicity are two major risk factors for gout that are linked by the activation of the NLRP3 inflammasome. Neutrophil-mediated production of interleukin-1 beta (IL-1 beta) drives gouty flares that cause joint destruction, intense pain, and fever. However, metabolites that impact neutrophil inflammasome remain unknown. Here, we identified that ketogenic diet (KD) increases beta-hydroxybutyrate (BHB) and alleviates urate crystal-induced gout without impairing immune defense against bacterial infection. BHB inhibited NLRP3 inflammasome in S100A9 fibril-primed and urate crystal-activated macrophages, which serve to recruit inflammatory neutrophils in joints. Consistent with reduced gouty flares in rats fed a ketogenic diet, BHB blocked IL-1 beta in neutrophils in a NLRP3-dependent manner in mice and humans irrespective of age. Mechanistically, BHB inhibited the NLRP3 inflammasome in neutrophils by reducing priming and assembly steps. Collectively, our studies show that BHB, a known alternate metabolic fuel, is also an anti-inflammatory molecule that may serve as a treatment for gout.

  • 264.
    Goldsteins, Gundars
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Persson, Håkan
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Andersson, Karin
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Olofsson, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Dacklin, Ingrid
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Edvinsson, Åsa
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Saraiva, Maria João
    Lundgren, Erik
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Exposure of cryptic epitopes on transthyretin only in amyloid and in amyloidogenic mutants1999Inngår i: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 96, nr 6, s. 3108-3113Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The structural requirements for generation of amyloid from the plasma protein transthyretin (TTR) are not known, although it is assumed that TTR is partly misfolded in amyloid. In a search for structural determinants important for amyloid formation, we generated a TTR mutant with high potential to form amyloid. We demonstrated that the mutant represents an intermediate in a series of conformational changes leading to amyloid. Two monoclonal antibodies were generated against this mutant; each displayed affinity to ex vivo TTR and TTR mutants with amyloidogenic folding but not to wild-type TTR or mutants exhibiting the wild-type fold. Two cryptic epitopes were mapped to a domain of TTR, where most mutations associated with amyloidosis occur and which we propose is displaced at the initial phase of amyloid formation, opening up new surfaces necessary for autoaggregation of TTR monomers. The results provide direct biochemical evidence for structural changes in an amyloidogenic intermediate of TTR.

  • 265. Gomes, Ana Rita
    et al.
    Bushell, Ellen
    Schwach, Frank
    Girling, Gareth
    Anar, Burcu
    Quail, Michael A.
    Herd, Colin
    Pfander, Claudia
    Modrzynska, Katarzyna
    Rayner, Julian C.
    Billker, Oliver
    Wellcome Trust Sanger Institute, Hinxton Cambridge CB10 1SA, UK.
    A genome-scale vector resource enables high-throughput reverse genetic screening in a malaria parasite2015Inngår i: Cell Host and Microbe, ISSN 1931-3128, E-ISSN 1934-6069, Vol. 17, nr 3, s. 404-413Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The genome-wide identification of gene functions in malaria parasites is hampered by a lack of reverse genetic screening methods. We present a large-scale resource of barcoded vectors with long homology arms for effective modification of the Plasmodium berghei genome. Cotransfecting dozens of vectors into the haploid blood stages creates complex pools of barcoded mutants, whose competitive fitness can be measured during infection of a single mouse using barcode sequencing (barseq). To validate the utility of this resource, we rescreen the P. berghei kinome, using published kinome screens for comparison. We find that several protein kinases function redundantly in asexual blood stages and confirm the targetability of kinases cdpk1, gsk3, tkl3, and PBANKA_082960 by genotyping cloned mutants. Thus, parallel phenotyping of barcoded mutants unlocks the power of reverse genetic screening for a malaria parasite and will enable the systematic identification of genes essential for in vivo parasite growth and transmission.

  • 266. Gomzikova, Marina O.
    et al.
    Zhuravleva, Margarita N.
    Miftakhova, Regina R.
    Arkhipova, Svetlana S.
    Evtugin, Vladimir G.
    Khaiboullina, Svetlana F.
    Kiyasov, Andrey P.
    Persson, Jenny L.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Mongan, Nigel P.
    Pestell, Richard G.
    Rizvanov, Albert A.
    Cytochalasin B-induced membrane vesicles convey angiogenic activity of parental cells2017Inngår i: OncoTarget, ISSN 1949-2553, E-ISSN 1949-2553, Vol. 8, nr 41, s. 70496-70507Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Naturally occurring extracellular vesicles (EVs) play essential roles in intracellular communication and delivery of bioactive molecules. Therefore it has been suggested that EVs could be used for delivery of therapeutics. However, to date the therapeutic application of EVs has been limited by number of factors, including limited yield and full understanding of their biological activities. To address these issues, we analyzed the morphology, molecular composition, fusion capacity and biological activity of Cytochalasin B-induced membrane vesicles (CIMVs). The size of these vesicles was comparable to that of naturally occurring EVs. In addition, we have shown that CIMVs from human SH-SY5Y cells contain elevated levels of VEGF as compared to the parental cells, and stimulate angiogenesis in vitro and in vivo.

  • 267. Govindasamy, K
    et al.
    Jebiwott, S
    Jaijyan, D K
    Davidow, A
    Ojo, K K
    Van Voorhis, W C
    Brochet, M
    Billker, Oliver
    Wellcome Trust Sanger Institute, Malaria Programme, Hinxton, UK.
    Bhanot, P
    Invasion of hepatocytes by Plasmodium sporozoites requires cGMP‐dependent protein kinase and calcium dependent protein kinase 42016Inngår i: Molecular Microbiology, ISSN 0950-382X, E-ISSN 1365-2958, Vol. 102, nr 2, s. 349-363Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Invasion of hepatocytes by sporozoites is essential for Plasmodium to initiate infection of the mammalian host. The parasite's subsequent intracellular differentiation in the liver is the first developmental step of its mammalian cycle. Despite their biological significance, surprisingly little is known of the signalling pathways required for sporozoite invasion. We report that sporozoite invasion of hepatocytes requires signalling through two second‐messengers – cGMP mediated by the parasite's cGMP‐dependent protein kinase (PKG), and Ca2+, mediated by the parasite's calcium‐dependent protein kinase 4 (CDPK4). Sporozoites expressing a mutated form of Plasmodium berghei PKG or carrying a deletion of the CDPK4 gene are defective in invasion of hepatocytes. Using specific and potent inhibitors of Plasmodium PKG and CDPK4, we demonstrate that PKG and CDPK4 are required for sporozoite motility, and that PKG regulates the secretion of TRAP, an adhesin that is essential for motility. Chemical inhibition of PKG decreases parasite egress from hepatocytes by inhibiting either the formation or release of merosomes. In contrast, genetic inhibition of CDPK4 does not significantly decrease the number of merosomes. By revealing the requirement for PKG and CDPK4 in Plasmodium sporozoite invasion, our work enables a better understanding of kinase pathways that act in different Plasmodium stages.

  • 268.
    Grabbe, Caroline
    et al.
    Goethe University Frankfurt am Main, Germany.
    Dikic, Ivan
    Goethe University Frankfurt am Main, Germany.
    Functional roles of ubiquitin-like domain (ULD) and ubiquitin-binding domain (UBD) containing proteins2009Inngår i: Chemical Reviews, ISSN 0009-2665, E-ISSN 1520-6890, Vol. 109, nr 4, s. 1481-1494Artikkel i tidsskrift (Fagfellevurdert)
  • 269.
    Grabowski, Pawel
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Telomere length as prognostic parameter in chronic lymphocytic leukemia2011Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    B-cell chronic lymphocytic leukemia (B-CLL) is the most common leukemia among the adult population in western countries and accounts for 30-40% of all leukemias. With survival time ranging from months to decades, the clinical course of individual CLL patients is highly variable. This heterogeneity and in the end the need for means to identify the patients with less favorable disease has encouraged the search for biomarkers that can predict the prognosis.

    Telomeres are repetitive structures protecting the chromosomal endings and shorten at each cell division. Telomere length (TL) has been indicated as a prognostic factor both in hematological malignancies and solid tumors. In B-CLL, TL is associated with mutation status of the immunoglobulin heavy chain variable (IGHV) gene and with clinical course. In the present thesis the main aim was to evaluate TL as a biomarker in B-CLL using a quantitative PCR-based method for TL determination.

    In paper I, TL was shown to be a prognostic factor for stage A and stage B/C patients, whereas IGHV mutation status predicted outcome only in stage A patients. Moreover, IGHV mutated CLL cases were subdivided by TL into two groups with different prognosis, a subdivision not seen for unmutated cases. Interestingly, the IGHV-mutated group with short telomeres had en overall survival close to that of the unmutated cases. Thus, a combination of IGHV mutation status and telomere length gave an improved subclassification of CLL identifying previously unrecognized patient groups with different outcomes.

    TL correlates with cellular origin of B-cell malignancies in relation to the germinal center (GC). In paper II different B-cell lymphoma/leukemia subtypes were analyzed. Shortest telomeres were found in IGHV unmutated CLLs, differing significantly from IGHV mutated cases. Contrary to this, mantle cell lymphomas (MCL) demonstrated similar TL regardless of IGHV mutation status. TL differed significantly between GC-like and non-GC-like diffuse large B-cell lymphomas (DLBCL) and follicular lymphomas (FL) had shorter telomeres than GC-like DLBCL. Hairy cell leukemias, which display Ig gene intraclonal heterogeneity, had longer telomeres than FLs and non-GC-DLBCL, but shorter than GC-DLBCL. In conclusion, TL seemed not to simply correlate with GC origin.

    Paper III presents a B-CLL cohort assessed for TL, genomic aberrations, IGHV mutation status, CD38 and ZAP-70 expression. An inverse correlation existed between TL and IGHV homology, CD38 and ZAP-70 expression. The presence of genomic aberrations was similar among patients regardless of TL. In contrast, 13q deletion, a favorable biomarker, was more frequent in patients with long telomeres, while 11q and 17p deletions (markers of less favorable outcome) were more frequent in the subgroup with short telomeres.

    In paper IV a large group of mainly indolent CLL cases from a population based cohort was studied again showing an association between TL and prognosis, especially in “good” prognosis cases as defined by other biomarkers. Multivariate analysis indicated a strong connection between IGHV mutation status, lipoprotein lipase (LPL) expression and TL. A comparison of TL in diagnostic and follow up samples demonstrated a significant correlation, and also in the follow samples TL constituted a significant biomarker for survival.

  • 270.
    Grabowski, Pawel
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Hultdin, Magnus
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Karlsson, Karin
    Tobin, Gerard
    Aleskog, Anna
    Thunberg, Ulf
    Laurell, Anna
    Sundström, Christer
    Rosenquist, Richard
    Roos, Göran
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Telomere length as a prognostic parameter in chronic lymphocytic leukemia with special reference to VH gene mutation status2005Inngår i: Blood, ISSN 0006-4971, E-ISSN 1528-0020, Vol. 105, nr 12, s. 4807-4812Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    B-cell chronic lymphocytic leukemia (CLL) consists of 2 prognostic entities where cases with mutated immunoglobulin VH genes have better outcome than unmutated cases. VH-mutated CLLs display longer telomeres compared with unmutated cases and telomere length has been indicated to predict outcome, although the prognostic value of telomere length has not been fully established in CLL. We analyzed telomere length, VH gene mutation status, and clinical parameters in a large series of CLL. Telomere length was assessed by quantitative polymerase chain reaction (PCR), giving a very good correlation to telomere length estimated by Southern blotting (P < .001). The prognostic information given by mutation status (n = 282) and telomere length (n = 246) was significant (P < .001, respectively). Telomere length was a prognostic factor for stage A (P = .021) and stage B/C (P = .018) patients, whereas mutation status predicted outcome only in stage A patients (P < .001). Furthermore, mutated CLLs were subdivided by telomere length into 2 groups with different prognoses (P = .003), a subdivision not seen for unmutated cases (P = .232). Interestingly, the VH-mutated group with short telomeres had an overall survival close to that of the unmutated cases. Thus, by combining VH mutation status and telomere length, an improved subclassification of CLL was achieved identifying previously unrecognized patient groups with different outcomes.

  • 271.
    Graffmo, Karin Sixtensdotter
    Umeå universitet, Medicinsk fakultet, Medicinsk biovetenskap.
    Of mice and men: SOD1 associated human amyotrophic lateral sclerosis and transgenic mouse models2007Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Amyotrophic lateral sclerosis, ALS, is a progressive fatal neurodegenerative disorder affecting motor neurones in motor cortex, brain stem and spinal cord. This inevitably leads to paralysis, respiratory failure and death. In about 5% of patients with ALS there is an association with mutations in gene for the abundant intracellular scavenging enzyme superoxide dismutase1, SOD1. The noxious property of SOD1 is proposed to be due to gain of function. In familial cases the inheritance is most commonly dominant.

    This study focus on two disparate SOD1 mutations occurring in Scandinavia. The recessive D90A mutation which has properties similar to that of the normal wild-type human SOD1. The dominantly inherited G127insTGGG mutation, G127X, causes a C-terminal truncation of the last 21 amino acids and is a highly unstable protein.

    Transgenic mice were created expressing D90A and G127X mutated human SOD1. Results from studies of tissue from the central nervous system of patients carrying either of these mutations were compared with similar tissue collected from transgenic mice generated with the same mutations. Tissue from the mice were also compared to central nervous tissue from several other transgenic mouse strains expressing human wild type SOD1 as well as other ALS associated human SOD1 mutations.

    The transgenic mice expressing D90A respectively G127X mutated human SOD1 develop motor neurone disease. Microscopic studies of central nervous tissues from G127X transgenic mice reveals inclusions of aggregated misfolded SOD1 in motor neurones and adjacent supporting cells. These inclusions are composed of detergent resistant aggregates and preceded by accumulations of minute quantities of detergent-soluble aggregates. The inclusions mimic those found in G127X patients.

    In D90A transgenic mice the progression, as in the humans, was slower and the mice, as the patients, showed bladder disturbance. In the D90A patients, the SOD1 inclusions mimic those found in sporadic ALS patients.

    Aggregation of SOD1 in central nervous tissue appears to be related to severity of disease. Degenerative features as vacuolization and gliosis precedes phenotypic alterations. Changes are seen not only in motor areas but also in higher centres of the telencephalon.

  • 272.
    Granberg, I
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB), Anatomi.
    Lindell, Björn
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB), Anatomi.
    Eriksson, Per-Olof
    Umeå universitet, Medicinska fakulteten, Institutionen för odontologi, Klinisk oral fysiologi.
    Pedrosa-Domellöf, Fatima
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk vetenskap, Oftalmiatrik.
    Stål, Per
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB), Anatomi.
    Capillary supply in relation to myosin heavy chain fibre composition of human intrinsic tongue muscles2010Inngår i: Cells Tissues Organs, ISSN 1422-6405, E-ISSN 1422-6421, Vol. 192, nr 5, s. 303-313Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The capillary supply and myosin heavy chain (MyHC) composition of three different intrinsic tongue muscles was analysed in the anterior and posterior regions of the human tongue with biochemical and immunohistochemical techniques. Mean capillary density for the whole tongue was 796 ± 82 cap/mm², without regional differences. The overall number of capillaries around each fibre (CAF) was higher in the posterior than in the anterior region (2.5 vs. 2.1, p = 0.009). However, correcting for regional differences in fibre size, CAF per fibre area was higher in the anterior region (4.3 vs. 3.0, p < 0.001). Muscle fibres containing fast MyHCs predominated in the anterior region (78.7%), consisting of MyHCIIa (58.5%), MyHCIIx (1.0%), MyHCIIa+MyHCIIx (11.3%) and MyHCI+MyHCIIa (7.9%). Fibres containing slow MyHC predominated in the posterior region (65.2%), consisting of MyHCI (45.5%) and MyHCI+MyHCIIa (19.7%). A minor fibre population (<2%) contained unusual MyHC isoforms, namely MyHC foetal, MyHC slow-tonic, MyHC α-cardiac or MyHC embryonic. The microvascularization of the human tongue was twice as high as in human limb muscles. Regional similarities in capillary supply, but differences in fibre phenotype composition, suggest that human tongue muscle fibres are fatigue resistant independently of MyHC content. High frequency of hybrid fibres, that is fibres co-expressing two or more MyHC isoforms, indicates a wider spectrum of fibre contractile properties than in limb muscles. In conclusion, human intrinsic tongue muscles showed internal specialization in distribution of MyHC isoforms and capillary supply, but not in the expression of unusual MyHCs.

  • 273.
    Granholm, Susanne
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för odontologi, Molekylär paradontologi.
    Henning, Petra
    Göteborgs universitet.
    Lerner, Ulf H
    Umeå universitet, Medicinska fakulteten, Institutionen för odontologi, Molekylär paradontologi.
    Comparisons between the effects of calcitonin receptor-stimulating peptide and intermedin and other peptides in the calcitonin family on bone resorption and osteoclastogenesis2011Inngår i: Journal of Cellular Biochemistry, ISSN 0730-2312, E-ISSN 1097-4644, Vol. 112, nr 11, s. 3300-3312Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Calcitonin receptor-stimulating peptide (CRSP) and intermedin (IMD) are two recently discovered peptides in the calcitonin (CT) family of peptides. CRSP and IMD, similar to CT, calcitonin gene-related peptide (CGRP) and amylin (AMY), but in contrast to adrenomedullin (ADM), inhibited bone resorption in mouse calvarial bones. CRSP and IMD, similar to CT, CGRP, AMY, but in contrast to ADM, decreased formation of osteoclasts and number of pits in bone marrow macrophage cultures stimulated by M-CSF and RANKL, with no effect on the expression of a number of genes associated with osteoclast progenitor cell differentiation. CRSP and IMD inhibited osteoclastogenesis at a late stage but had no effect on DC-STAMP mRNA. IMD, similar to CGRP, AMY and ADM stimulated cyclic AMP formation in M-CSF expanded osteoclast progenitor cells lacking CT receptors. RANKL induced CT receptors and a cyclic AMP response also to CT and CRSP, and increased the cyclic AMP response to CGRP, AMY and IMD but decreased the response to ADM. Our data demonstrate that CRSP and IMD share several functional properties of peptides in the CT family of peptides, including inhibition of bone resorption and osteoclast formation. The data also show that the reason why ADM does not inhibit osteoclast activity or formation is related to the fact that RANKL decreases ADM receptor signalling through the adenylate cyclase-cyclic AMP pathway. Finally, the findings indicate that activation by CGRP, AMY and IMD may include activation of both CT and CT receptor-like receptors. J. Cell. Biochem. © 2011 Wiley-Liss, Inc.

  • 274. Grasso, Francesca
    et al.
    Frisan, Teresa
    Department Cell and Molecular Biology, Karolinska Institutet.
    Bacterial Genotoxins: Merging the DNA Damage Response into Infection Biology2015Inngår i: Biomolecules, E-ISSN 2218-273X, Vol. 5, nr 3, s. 1762-1782Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Bacterial genotoxins are unique among bacterial toxins as their molecular target is DNA. The consequence of intoxication or infection is induction of DNA breaks that, if not properly repaired, results in irreversible cell cycle arrest (senescence) or death of the target cells. At present, only three bacterial genotoxins have been identified. Two are protein toxins: the cytolethal distending toxin (CDT) family produced by a number of Gram-negative bacteria and the typhoid toxin produced by Salmonella enterica serovar Typhi. The third member, colibactin, is a peptide-polyketide genotoxin, produced by strains belonging to the phylogenetic group B2 of Escherichia coli. This review will present the cellular effects of acute and chronic intoxication or infection with the genotoxins-producing bacteria. The carcinogenic properties and the role of these effectors in the context of the host-microbe interaction will be discussed. We will further highlight the open questions that remain to be solved regarding the biology of this unusual family of bacterial toxins.

  • 275.
    Grimsholm, Ola
    Umeå universitet, Medicinsk fakultet, Integrativ medicinsk biologi.
    Neuropeptides and neurotrophins in arthritis: studies on the human and mouse knee joint2008Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Neuropeptides, such as substance P (SP) and bombesin/gastrin-releasing peptide (BN/GRP), and neurotrophins are involved in neuro-immunomodulatory processes and have marked trophic, growth-promoting and inflammation-modulating properties. The impact of these modulators in rheumatoid arthritis (RA) is, however, unclear. An involvement of the innervation, including the peptidergic innervation, is frequently proposed as an important factor for arthritic disease. Many patients with RA, but not all, benefit from treatment with anti-TNF medications.

    The studies presented here aimed to investigate the roles of neuropeptides, with an emphasis on BN/GRP and SP, and neurotrophins, especially with attention to brain-derived neurotrophic factor (BDNF), in human and murine knee joint tissue. The expression patterns of these substances and their receptors in synovial tissue from patients with either RA or osteoarthritis (OA) were studied in parallel with the levels of these factors in blood and synovial fluid from patients with RA and from healthy controls. Correlation studies were also performed comparing the levels of neuropeptides with those of pro-inflammatory cytokines [tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6)]. Furthermore, the impact of anti-TNF treatment on the levels of BDNF in blood was investigated. In a murine model of RA, the expression of these substances on articular chondrocytes along with their expression in synovial tissue was investigated.

    The expression of BN/GRP in human synovial tissue was confined to fibroblast-like and mononuclear-like cells whereas SP was detected in nerve-related structures. Receptors for these neuropeptides (GRP-R and NK-1R) were frequently present in blood vessel walls, and on fibroblast-like and mononuclear-like cells. The expression of BDNF and its receptors, p75 neurotrophin receptor and TrkB, was mainly confined to nerve structures. The levels of SP, and particularly those of BN/GRP, in synovial fluid and peripheral blood correlated with the levels of pro-inflammatory cytokines. There were clearly more correlations between SP-BN/GRP and inflammatory parameters than between BDNF and these factors. Plasma levels of BDNF were decreased following anti-TNF-treatment. In the joints of the murine model, there was a marked expression of neurotrophins, neurotrophin receptors and NK-1R/GRP-R in the articular chondrocytes. The expression was down-regulated in the arthritic animals. A neurotrophin system was found to develop in the inflammatory infiltrates of the synovium in the arthritic mice.

    The results presented suggest that there is a local, and not nerve-related, supply of BN/GRP in the human synovial tissue. Furthermore, BN/GRP and SP have marked effects in the synovial tissue of patients with RA, i.e., there were abundant receptor expressions, and these neuropeptides are, together with cytokines, likely to be involved in the neuro-immunomodulation that occurs in arthritis. The observations do on the whole suggest that the neuropeptides, rather than BDNF, are related to inflammatory processes in the human knee joint. A new effect of anti-TNF treatment; i.e., lowering plasma levels of BDNF, was observed. Severe arthritis, as in the murine model, lead to a decrease in the levels of neurotrophin, and neurotrophin and neuropeptide receptor expressions in the articular cartilage. This fact might be a drawback for the function of the chondrocytes. Certain differences between the expression patterns in the synovial tissue of the murine model and those of human arthritic synovial tissue were noted. It is obvious that local productions in the synovial tissue, nerve-related supply in this tissue and productions in chondrocytes to different extents occur for the investigated substances.

  • 276. Grong, Eivind
    et al.
    Kulseng, Bård
    Arbo, Ingerid Brænne
    Nord, Christoffer
    Umeå universitet, Medicinska fakulteten, Umeå centrum för molekylär medicin (UCMM).
    Eriksson, Maria
    Umeå universitet, Medicinska fakulteten, Umeå centrum för molekylär medicin (UCMM).
    Ahlgren, Ulf
    Umeå universitet, Medicinska fakulteten, Umeå centrum för molekylär medicin (UCMM).
    Mårvik, Ronald
    Sleeve gastrectomy, but not duodenojejunostomy, preserves total beta-cell mass in Goto-Kakizaki rats evaluated by three-dimensional optical projection tomography2016Inngår i: Surgical Endoscopy, ISSN 0930-2794, E-ISSN 1432-2218, Vol. 30, nr 2, s. 532-542Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Background In type 2 diabetes mellitus, there is a progressive loss of beta-cell mass. Bariatric surgery has in recent investigations showed promising results in terms of diabetes remission, but little is established regarding the effect of surgery on the survival or regeneration of pancreatic beta-cells. In this study, we aim to explore how bariatric surgery with its subsequent hormonal alterations affects the islets of Langerhans.

    Methods Twenty-four Goto-Kakizaki rats were operated with duodenojejunostomy (DJ), sleeve gastrectomy (SG) or sham operation. From the 38th week after surgery, body weight, fasting blood glucose, glycosylated hemoglobin, mixed meal tolerance with repeated measures of insulin, glucagon-like peptide 1, gastrin and total ghrelin were evaluated. Forty-six weeks after surgery, the animals were euthanized and the total beta-cell mass in all animals was examined by three-dimensional volume quantification by optical projection tomography based on the signal from insulin-specific antibody staining.

    Results Body weight did not differ between groups (Pg = 0.37). SG showed lower fasting blood glucose compared to DJ and sham (Pg = 0.037); HbA1c levels in SG were lower compared to DJ only (p\0.05). GLP-1 levels were elevated for DJ compared to SG and sham (Pg = 0.001), whereas gastrin levels were higher in SG compared to the two other groups (Pg = 0.002). Beta-cell mass was significantly greater in animals operated with SG compared to both DJ and sham (p = 0.036).

    Conclusion Sleeve gastrectomy is superior to duodenojejunostomy and sham operation when comparing the preservation of beta-cell mass 46 weeks after surgery in Goto-Kakizaki rats. This could be related to both the increased gastrin levels and the long-term improvement in glycemic parameters observed after this procedure.

  • 277.
    Grubbström, Olivia
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik. Biomedicinprogrammet.
    In vitro study of drugs against mammalian ribonucleotide reductase2014Independent thesis Basic level (degree of Bachelor), 10 poäng / 15 hpOppgave
  • 278.
    Grundström, Robert
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Is DNA polymerase ε a new driver in cancer development?: Functional studies of cancer associated mutations2015Independent thesis Basic level (degree of Bachelor), 10 poäng / 15 hpOppgave
  • 279.
    Grundström, Thomas
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Hauser, Jannek
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Grundström, Christine
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Kumar, Ramesh
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Ahmed, Tanzeel
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Mechanisms controlling diversification and affinity maturation of antibodies2015Inngår i: International Journal of Molecular Medicine, ISSN 1107-3756, E-ISSN 1791-244X, Vol. 36, s. S43-S43Artikkel i tidsskrift (Annet vitenskapelig)
  • 280.
    Gu, Weigang
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för farmakologi och klinisk neurovetenskap, Neurologi. Umeå universitet, Medicinska fakulteten, Institutionen för folkhälsa och klinisk medicin, Medicin.
    Brännström, Thomas
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Rosqvist, Roland
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Wester, Per
    Umeå universitet, Medicinska fakulteten, Institutionen för folkhälsa och klinisk medicin, Medicin.
    Cell division in the cerebral cortex of adult rats after photothrombotic ring stroke.2009Inngår i: Stem Cell Research, ISSN 1876-7753, Vol. 2, nr 1, s. 68-77Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Neurogenesis has been shown to occur in the cerebral cortex in adult rats after ischemic stroke. The origin of the newborn neurons is largely unknown. This study aimed to explore cell division in the poststroke penumbral cortex. Adult male Wistar rats were subjected to photothrombotic ring stroke. After repeated delivery of the DNA duplication marker BrdU, the animals were sacrificed at various times poststroke. BrdU was detected by immunohistochemistry/immunofluorescence labeling, as was the M-phase marker Phos H3 and the spindle components alpha-tubulin/gamma-tubulin. DNA damage was examined by TUNEL staining. Cell type was ascertained by double immunolabeling with the neuronal markers Map-2ab/beta-tubulin III and NeuN/Hu or the astrocyte marker GFAP. From 16h poststroke, BrdU-immunolabeled cells appeared in the penumbral cortex. From 24h, Phos H3 was colocalized with BrdU in the nuclei. Mitotic spindles immunolabeled by alpha-tubulin/gamma-tubulin appeared inside the cortical cells containing BrdU-immunopositive nuclei. Unexpectedly, the markers of neuronal differentiation, Map-2ab/beta-tubulin III/NeuN/Hu, were expressed in the Phos H3-immunolabeled cells, and NeuN was detected in some cells containing spindles. This study suggests that in response to a sublethal ischemic insult, endogenous cells with neuronal immunolabeling may duplicate their nuclear DNA and commit cell mitosis to generate daughter neurons in the penumbral cortex in adult rats.

  • 281.
    Gu, Xiaolian
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    p63 and epithelial homeostasis: studies of p63 under normal, hyper-proliferative and malignant conditions2010Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Background: The p63 gene is a member of the p53 transcription factor family and can produce six different proteins using two promoters and differential splicing. Expression of p63 is required for proper formation of epithelial tissues. Studies on the transcriptional control of specific genes involved in cell survival, proliferation, differentiation and adhesion have revealed the contributions of p63 to the continuously renewing stratified epithelium. In this thesis, the aim was to improve our understanding of the roles of p63 in epithelial homeostasis by investigating expression of p63 in normal and benign hyper-proliferative epithelia and exploring the influence of p63 deregulation on cancer progression.

    Materials and methods: Using quantitative real time RT-PCR and immunohistochemistry, we first examined the expression of different p63 isoforms in patients diagnosed with psoriasis - a benign hyper-proliferative and inflammatory skin disease. Afterwards, we investigated responses of p63 in psoriatic epidermis upon Narrowband-UVB (NB-UVB) phototherapy. At the same time, we studied the potential impact of p63 in carcinogenesis by searching for p63 transcriptional targets in a cell line derived from squamous cell carcinoma of the head and neck (SCCHN) - the sixth most common cancer worldwide with over-expression of the ∆Np63α protein as a common feature. p63 gene silencing and microarray were used to identify p63 regulated genes. Real time RT-PCR, western blot, immunohistochemistry, chromatin immunoprecipitation, transient transfection and reporter assays were performed to confirm specific genes as direct p63 targets.

    Results: Significant down-regulation of p63 mRNA levels was found in psoriatic lesions compared to patients’ own clinically normal skin. Moreover, a trend of decreased TAp63 mRNA levels was seen in patients’ normal skin compared to age- and sex-matched healthy controls. Following NB-UVB phototherapy, an effective first line therapy for psoriasis, expression of p63 was not significantly affected. However, significant changes in p53, FABP5, miR-21 and miR-125b were found. Surprisingly, location and expression levels of p63 proteins detected by immunohistochemistry were similar under all skin conditions. A direct transcriptional regulation of TRAF4 by p63 was seen in the SCCHN cell line and we further found that the localization of the TRAF4 protein was associated with histological differentiation of SCCHN cells. However, unlike its over-expression in SCCHN, similar TRAF4 mRNA expression levels were seen in psoriatic lesions as compared to healthy controls. Besides TRAF4, a total of 127 genes were identified as potentially p63 regulated in the SCCHN cell line and strikingly, about 20% of these genes are involved in cell adhesion or migration.

    Conclusions: Dysregulation of p63 isoforms in psoriatic epidermis, especially decreased TAp63 expression, and their resistance to NB-UVB phototherapy implicated a contribution of p63 to the psoriasis phenotype. Transcriptional regulation of genes involved in multiple biological pathways indicated that over-expression of p63 in SCCHN might account for altered cell differentiation, adhesion and migration, thus contributing to SCCHN. In conclusion, our studies have found additional mechanisms through which p63 guarded homeostasis of the established epithelium. Deregulation of p63 might play a role in distinct pathological conditions by participating in diverse cellular pathways under different microenvironments.

  • 282.
    Gu, Xiaolian
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Coates, Philip J
    Boldrup, Linda
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Nylander, Karin
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    p63 contributes to cell invasion and migration in squamous cell carcinoma of the head and neck2008Inngår i: Cancer Letters, ISSN 0304-3835, E-ISSN 1872-7980, Vol. 263, nr 1, s. 26-34Artikkel i tidsskrift (Fagfellevurdert)
  • 283.
    Gu, Xiaolian
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Coates, Philip
    MacCallum, Stephanie
    Boldrup, Linda
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Sjöström, Björn
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk vetenskap, Öron- näs- och halssjukdomar.
    Nylander, Karin
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    TRAF4 is potently induced by TAp63 isoforms and localised according to differentiation in SCCHN2007Inngår i: Cancer Biology & Therapy, ISSN 1538-4047, E-ISSN 1555-8576, Vol. 6, nr 12, s. 1979-1983Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    p63, a member of the p53 family, is overexpressed in squamous cell carcinoma of the head and neck (SCCHN) and some other tumors of epithelial origin. As a transcription factor, p63 can bind to p53-type response elements and there is some overlap between p53 family transcriptional targets. Tumor necrosis factor receptor associated factor 4 (TRAF4) is a p53 regulated gene which is overexpressed in many human carcinomas. We investigated the involvement of p63 in regulation of TRAF4 and the expression of the TRAF4 protein in SCCHN. Disrupting endogenous p63 expression resulted in downregulation of TRAF4 mRNA and protein in an SCCHN cell line. Endogenous p63 bound to the TRAF4 promoter in vivo and reporter assays showed that p63, p73 and p53 can all transactivate TRAF4, with TAp63 isoforms being the most potent activators. The level of TRAF4 activation by TAp63 was two-fold higher than by p53, and TRAF4 was ten-fold more responsive to TAp63 than another p63-target, IGFBP3. Nuclear expression of TRAF4 was seen in normal oral epithelium and highly/moderately differentiated SCCHN, whereas cytoplasmic expression of TRAF4 was seen in poorly differentiated SCCHN. These results indicate that TRAF4 is a common target of p53 family members and that localization of TRAF4 is associated with differentiation of SCCHN cells.

  • 284.
    Gu, Xiaolian
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Lundqvist, Elisabet N
    Umeå universitet, Medicinska fakulteten, Institutionen för folkhälsa och klinisk medicin, Dermatologi och venereologi.
    Coates, Philip J
    Thurfjell, Niklas
    Wettersand, Emma
    Nylander, Karin
    Dysregulation of TAp63 mRNA and protein levels in psoriasis2006Inngår i: Journal of Investigative Dermatology, ISSN 0022-202X, E-ISSN 1523-1747, Vol. 126, nr 1, s. 137-141Artikkel i tidsskrift (Fagfellevurdert)
  • 285.
    Gu, Xiaolian
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Nylander, E
    Coates, PJ
    Nylander, Karin
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Little effect on p63 but significant effect on miR-21 and miR-125b by NB-UVB phototherapy on psoriatic lesions2010Manuskript (preprint) (Annet vitenskapelig)
    Abstract [en]

    Psoriasis is an inflammatory skin disease in which dysregulation of p63, a member of the p53 family and crucial for skin development and maintenance, has been shown. Though currently incurable, many therapies are available including narrowband ultraviolet B (NB-UVB) phototherapy. To further elucidate the role of p63 in psoriasis and increase our understanding of the mechanisms of phototherapy, we studied the effects of NB-UVB treatment on p63 expression. Expression of p53 was also studied due to its functional role in the response of skin to UV. In addition, we investigated expression of miR-203, miR-125b and miR-21, as these microRNAs are p63 and/or p53 regulators and their involvement in psoriasis pathogenesis has previously been suggested. Skin biopsies from 12 psoriasis patients were collected before, during and at the final session of phototherapy. Real time RT-PCR and immunohistochemistry showed that epidermal p63 mRNA and protein levels were not significantly affected following phototherapy, whereas a significant increase in p53 mRNA expression and protein accumulation was found. NB-UVB treatment also significantly affected expression of miR-21 and miR-125b, whereas individual clinical improvement seemed related to p53 status only. Our results indicate that even though NB-UVB phototherapy causes diverse molecular changes, induction of p53 is pivotal for successful treatment of psoriasis, and unresolved p63 abnormality in the treated epidermis of psoriasis patients further indicate a role for p63 in psoriasis.

  • 286.
    Gu, Xiaolian
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Wang, Lixiao
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Boldrup, Linda
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Coates, Philip J.
    Fåhraeus, Robin
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi. RECAMO, Masaryk Memorial Cancer Institute, 656 53 Brno, Czech Republic; Équipe Labellisée Ligue Contre le Cancer, INSERM UMRS1162, Institut de Génétique Moléculaire, Université Paris 7, IUH Hôpital St. Louis, 75010 Paris, France.
    Sgaramella, Nicola
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    Wilms, Torben
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk vetenskap, Öron- näs- och halssjukdomar.
    Nylander, Karin
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Patologi.
    AP001056.1, A Prognosis-Related Enhancer RNA in Squamous Cell Carcinoma of the Head and Neck2019Inngår i: Cancers, ISSN 2072-6694, Vol. 11, nr 3, artikkel-id 347Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    A growing number of long non-coding RNAs (lncRNAs) have been linked to squamous cell carcinoma of the head and neck (SCCHN). A subclass of lncRNAs, termed enhancer RNAs (eRNAs), are derived from enhancer regions and could contribute to enhancer function. In this study, we developed an integrated data analysis approach to identify key eRNAs in SCCHN. Tissue-specific enhancer-derived RNAs and their regulated genes previously predicted using the computational pipeline PreSTIGE, were considered as putative eRNA-target pairs. The interactive web servers, TANRIC (the Atlas of Noncoding RNAs in Cancer) and cBioPortal, were used to explore the RNA levels and clinical data from the Cancer Genome Atlas (TCGA) project. Requiring that key eRNAs should show significant associations with overall survival (Kaplan-Meier log-rank test, p < 0.05) and the predicted target (correlation coefficient r > 0.4, p < 0.001), we identified five key eRNA candidates. The most significant survival-associated eRNA was AP001056.1 with ICOSLG encoding an immune checkpoint protein as its regulated target. Another 1640 genes also showed significant correlation with AP001056.1 (r > 0.4, p < 0.001), with the "immune system process" being the most significantly enriched biological process (adjusted p < 0.001). Our results suggest that AP001056.1 is a key immune-related eRNA in SCCHN with a positive impact on clinical outcome.

  • 287. Guan, Jikui
    et al.
    Fransson, Susanne
    Siaw, Joachim Tetteh
    Treis, Diana
    Van den Eynden, Jimmy
    Chand, Damini
    Umapathy, Ganesh
    Ruuth, Kristina
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för molekylärbiologi (Teknisk-naturvetenskaplig fakultet).
    Svenberg, Petter
    Wessman, Sandra
    Shamikh, Alia
    Jacobsson, Hans
    Gordon, Lena
    Stenman, Jakob
    Svensson, Pär-Johan
    Hansson, Magnus
    Larsson, Erik
    Martinsson, Tommy
    Palmer, Ruth H.
    Kogner, Per
    Hallberg, Bengt
    Clinical response of the novel activating ALK-I1171T mutation in neuroblastoma o the ALK inhibitor ceritinib2018Inngår i: Cold Spring Harbor Molecular Case Studies, ISSN 2373-2873, Vol. 4, nr 4, artikkel-id a002550Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Tumors with anaplastic lymphoma kinase (ALK) fusion rearrangements, including non-small-cell lung cancer and anaplastic large cell lymphoma, are highly sensitive to ALK tyrosine kinase inhibitors (TKIs), underscoring the notion that such cancers are addicted to ALK activity. Although mutations in ALK are heavily implicated in childhood neuroblastoma, response to the ALK TKI crizotinib has been disappointing. Embryonal tumors in patients with DNA repair defects such as Fanconi anemia (FA) often have a poor prognosis, because of lack of therapeutic options. Here we report a child with underlying FA and ALK mutant high-risk neuroblastoma responding strongly to precision therapy with the ALK TKI ceritinib. Conventional chemotherapy treatment caused severe, life-threatening toxicity. Genomic analysis of the initial biopsy identified germline FANCA mutations as well as a novel ALK-I1171T variant. ALK-I1171T generates a potent gain-of-function mutant, as measured in PC12 cell neurite outgrowth and NIH3T3 transformation. Pharmacological inhibition profiling of ALK-I1171T in response to various ALK TKIs identified an 11-fold improved inhibition of ALK-I1171T with ceritinib when compared with crizotinib. Immunoaffinity-coupled LC-MS/MS phosphoproteomics analysis indicated a decrease in ALK signaling in response to ceritinib. Ceritinib was therefore selected for treatment in this child. Monotherapy with ceritinib was well tolerated and resulted in normalized catecholamine markers and tumor shrinkage. After 7.5 mo treatment, the residual primary tumor shrunk, was surgically removed, and exhibited hallmarks of differentiation together with reduced Ki67 levels. Clinical follow-up after 21 mo treatment revealed complete clinical remission including all metastatic sites. Therefore, ceritinib presents a viable therapeutic option for ALK-positive neuroblastoma.

  • 288. Guan, Jikui
    et al.
    Umapathy, Ganesh
    Yamazaki, Yasuo
    Wolfstetter, Georg
    Mendoza-Garcia, Patricia
    Department of Medical Biochemistry and Cell Biology, Instititute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
    Pfeifer, Kathrin
    Mohammed, Ateequrrahman
    Hugosson, Fredrik
    Zhang, Hongbing
    Hsu, Amy W
    Halenbeck, Robert
    Hallberg, Bengt
    Palmer, Ruth H
    FAM150A and FAM150B are activating ligands for anaplastic lymphoma kinase2015Inngår i: eLIFE, E-ISSN 2050-084X, Vol. 4, artikkel-id e09811Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Aberrant activation of anaplastic lymphoma kinase (ALK) has been described in a range of human cancers, including non-small cell lung cancer and neuroblastoma (Hallberg and Palmer, 2013). Vertebrate ALK has been considered to be an orphan receptor and the identity of the ALK ligand(s) is a critical issue. Here we show that FAM150A and FAM150B are potent ligands for human ALK that bind to the extracellular domain of ALK and in addition to activation of wild-type ALK are able to drive 'superactivation' of activated ALK mutants from neuroblastoma. In conclusion, our data show that ALK is robustly activated by the FAM150A/B ligands and provide an opportunity to develop ALK-targeted therapies in situations where ALK is overexpressed/activated or mutated in the context of the full length receptor.

  • 289.
    Guan, Jikui
    et al.
    Institute of Biomedicine,Dept of Medical Biochem and Cell Biol, Sahlgrenska Academy, University of Gothenburg..
    Yamazaki, Yasuo
    Institute of Biomedicine,Dept of Medical Biochem and Cell Biol, Sahlgrenska Academy, University of Gothenburg..
    Chand, Damini
    Institute of Biomedicine,Dept of Medical Biochem and Cell Biol, Sahlgrenska Academy, University of Gothenburg..
    van Dijk, Jesper R.
    Institute of Biomedicine,Dept of Medical Biochem and Cell Biol, Sahlgrenska Academy, University of Gothenburg..
    Ruuth, Kristina
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Palmer, Ruth H.
    Institute of Biomedicine,Dept of Medical Biochem and Cell Biol, Sahlgrenska Academy, University of Gothenburg..
    Hallberg, Bengt
    Institute of Biomedicine,Dept of Medical Biochem and Cell Biol, Sahlgrenska Academy, University of Gothenburg..
    Novel mechanisms of ALK activation revealed by analysis of the Y1278S neuroblastoma mutation2017Inngår i: Cancers, ISSN 2072-6694, Vol. 9, nr 11, artikkel-id 149Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Numerous mutations have been observed in the Anaplastic Lymphoma Kinase (ALK) receptor tyrosine kinase (RTK) in both germline and sporadic neuroblastoma. Here, we have investigated the Y1278S mutation, observed in four patient cases, and its potential importance in the activation of the full length ALK receptor. Y1278S is located in the 1278-YRASYY-1283 motif of the ALK activation loop, which has previously been reported to be important in the activation of the ALK kinase domain. In this study, we have characterized activation loop mutations within the context of the full length ALK employing cell culture and Drosophila melanogaster model systems. Our results show that the Y1278S mutant observed in patients with neuroblastoma harbors gain-of-function activity. Secondly, we show that the suggested interaction between Y1278 and other amino acids might be of less importance in the activation process of the ALK kinase than previously proposed. Thirdly, of the three individual tyrosines in the 1278-YRASYY-1283 activation loop, we find that Y1283 is the critical tyrosine in the activation process. Taken together, our observations employing different model systems reveal new mechanistic insights on how the full length ALK receptor is activated and highlight differences with earlier described activation mechanisms observed in the NPM-ALK fusion protein, supporting a mechanism of activation more in line with those observed for the Insulin Receptor (InR).

  • 290.
    Gudey, Shyam Kumar
    et al.
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap.
    Sundar, Reshma
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap.
    Heldin, Carl-Henrik
    Ludwig Institute for Cancer Research, Uppsala.
    Landström, Marene
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap.
    Pro-invasive Snail1 targets TGFbeta receptor I to promote epithelial to mesenchymal transition in prostate cancerManuskript (preprint) (Annet vitenskapelig)
  • 291. Guerra, Lina
    et al.
    Albihn, Ami
    Tronnersjö, Susanna
    Yan, Qinzi
    Guidi, Riccardo
    Stenerlöw, Bo
    Sterzenbach, Torsten
    Josenhans, Christine
    Fox, James G
    Schauer, David B
    Thelestam, Monica
    Larsson, Lars-Gunnar
    Henriksson, Marie
    Frisan, Teresa
    Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden.
    Myc is required for activation of the ATM-dependent checkpoints in response to DNA damage2010Inngår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 5, nr 1, artikkel-id e8924Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    BACKGROUND: The MYC protein controls cellular functions such as differentiation, proliferation, and apoptosis. In response to genotoxic agents, cells overexpressing MYC undergo apoptosis. However, the MYC-regulated effectors acting upstream of the mitochondrial apoptotic pathway are still unknown.

    PRINCIPAL FINDINGS: In this study, we demonstrate that expression of Myc is required to activate the Ataxia telangiectasia mutated (ATM)-dependent DNA damage checkpoint responses in rat cell lines exposed to ionizing radiation (IR) or the bacterial cytolethal distending toxin (CDT). Phosphorylation of the ATM kinase and its downstream effectors, such as histone H2AX, were impaired in the myc null cell line HO15.19, compared to the myc positive TGR-1 and HOmyc3 cells. Nuclear foci formation of the Nijmegen Breakage Syndrome (Nbs) 1 protein, essential for efficient ATM activation, was also reduced in absence of myc. Knock down of the endogenous levels of MYC by siRNA in the human cell line HCT116 resulted in decreased ATM and CHK2 phosphorylation in response to irradiation. Conversely, cell death induced by UV irradiation, known to activate the ATR-dependent checkpoint, was similar in all the cell lines, independently of the myc status.

    CONCLUSION: These data demonstrate that MYC contributes to the activation of the ATM-dependent checkpoint responses, leading to cell death in response to specific genotoxic stimuli.

  • 292. Guerra, Lina
    et al.
    Cortes-Bratti, Ximena
    Guidi, Riccardo
    Frisan, Teresa
    Department of Cell and Molecular Biology, Karolinska Institute, Stockholm, Sweden.
    The biology of the cytolethal distending toxins2011Inngår i: Toxins, ISSN 2072-6651, E-ISSN 2072-6651, Vol. 3, nr 3, s. 172-190Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    The cytolethal distending toxins (CDTs), produced by a variety of Gram-negative pathogenic bacteria, are the first bacterial genotoxins described, since they cause DNA damage in the target cells. CDT is an A-B(2) toxin, where the CdtA and CdtC subunits are required to mediate the binding on the surface of the target cells, allowing internalization of the active CdtB subunit, which is functionally homologous to the mammalian deoxyribonuclease I. The nature of the surface receptor is still poorly characterized, however binding of CDT requires intact lipid rafts, and its internalization occurs via dynamin-dependent endocytosis. The toxin is retrograde transported through the Golgi complex and the endoplasmic reticulum, and subsequently translocated into the nuclear compartment, where it exerts the toxic activity. Cellular intoxication induces DNA damage and activation of the DNA damage responses, which results in arrest of the target cells in the G1 and/or G2 phases of the cell cycle and activation of DNA repair mechanisms. Cells that fail to repair the damage will senesce or undergo apoptosis. This review will focus on the well-characterized aspects of the CDT biology and discuss the questions that still remain unanswered.

  • 293. Guerra, Lina
    et al.
    Guidi, Riccardo
    Frisan, Teresa
    Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden.
    Do bacterial genotoxins contribute to chronic inflammation, genomic instability and tumor progression?2011Inngår i: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658, Vol. 278, nr 23, s. 4577-4588Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Cytolethal distending toxin, produced by several Gram-negative bacteria, and colibactin, secreted by several commensal and extraintestinal pathogenic Escherichia coli strains, are the first bacterial genotoxins to be described to date. Exposure to cytolethal distending toxin and colibactin induces DNA damage, and consequently activates the DNA damage response, resulting in cell cycle arrest of the intoxicated cells and DNA repair. Irreversible DNA damage will lead to cell death by apoptosis or to senescence. It is well established that chronic exposure to DNA damaging agents, either endogenous (reactive oxygen species) or exogenous (ionizing radiation), may cause genomic instability as a result of the alteration of genes coordinating the DNA damage response, thus favoring tumor initiation and progression. In this review, we summarize the state of the art of the biology of cytolethal distending toxin and colibactin, focusing on the activation of the DNA damage response and repair pathways, and discuss the cellular responses induced in intoxicated cells, as well as how prolonged intoxication may lead to chronic inflammation, the accumulation of genomic instability, and tumor progression in both in vitro and in vivo models.

  • 294. Guerra, Lina
    et al.
    Guidi, Riccardo
    Slot, Ilse
    Callegari, Simone
    Sompallae, Ramakrishna
    Pickett, Carol L
    Åström, Stefan
    Eisele, Frederik
    Wolf, Dieter
    Sjögren, Camilla
    Masucci, Maria G
    Frisan, Teresa
    Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm.
    Bacterial genotoxin triggers FEN1-dependent RhoA activation, cytoskeleton remodeling and cell survival2011Inngår i: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 124, nr 16, s. 2735-2742Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The DNA damage response triggered by bacterial cytolethal distending toxins (CDTs) is associated with activation of the actin-regulating protein RhoA and phosphorylation of the downstream-regulated mitogen-activated protein kinase (MAPK) p38, which promotes the survival of intoxicated (i.e. cells exposed to a bacterial toxin) cells. To identify the effectors of this CDT-induced survival response, we screened a library of 4492 Saccharomyces cerevisiae mutants that carry deletions in nonessential genes for reduced growth following inducible expression of CdtB. We identified 78 genes whose deletion confers hypersensitivity to toxin. Bioinformatics analysis revealed that DNA repair and endocytosis were the two most overrepresented signaling pathways. Among the human orthologs present in our data set, FEN1 and TSG101 regulate DNA repair and endocytosis, respectively, and also share common interacting partners with RhoA. We further demonstrate that FEN1, but not TSG101, regulates cell survival, MAPK p38 phosphorylation, RhoA activation and actin cytoskeleton reorganization in response to DNA damage. Our data reveal a previously unrecognized crosstalk between DNA damage and cytoskeleton dynamics in the regulation of cell survival, and might provide new insights on the role of chronic bacteria infection in carcinogenesis.

  • 295. Guidi, R
    et al.
    Belluz, L Del Bell
    Frisan, Teresa
    Dept. of Cell and Molecular Biology, Karolinska Institute, Stockholm Sweden.
    Bacterial genotoxin functions as immune-modulator and promotes host survival2016Inngår i: Microbial cell (Graz, Austria), ISSN 2311-2638, Vol. 3, nr 8, s. 355-357Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Bacterial genotoxins are effectors that cause DNA damage in target cells. Many aspects of the biology of these toxins have been characterised in vitro, such as structure, cellular internalisation pathways and effects on the target cells. However, little is known about their function in vivo. Salmonella enterica serovar Typhi (S. Typhi) is a Gram-negative, intracellular bacterium that causes typhoid fever, a debilitating disease infecting more than 20 million people every year. S. Typhiproduce a genotoxin named typhoid toxin (TT), but its role in the contest of host infection is poorly characterized. The major obstacle in addressing this issue is that S. Typhi is exclusively a human pathogen. To overcome this limitation, we have used as model bacterium S. Typhimurium, and engineered it to produce endogenous levels of an active and inactive typhoid toxin, hereby named as TT (or genotoxic) and cdtB (or control), respectively. To our surprise, infection with the genotoxin strain strongly suppressed intestinal inflammation, leading to a better survival of the host during the acute phase of infection, suggesting typhoid toxin may exert a protective role. The presence of a functional genotoxin was also associated with an increased frequency of asymptomatic carriers.

  • 296. Guidi, Riccardo
    et al.
    Guerra, Lina
    Levi, Laura
    Stenerlöw, Bo
    Fox, James G
    Josenhans, Christine
    Masucci, Maria G
    Frisan, Teresa
    Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden.
    Chronic exposure to the cytolethal distending toxins of Gram-negative bacteria promotes genomic instability and altered DNA damage response2013Inngår i: Cellular Microbiology, ISSN 1462-5814, E-ISSN 1462-5822, Vol. 15, nr 1, s. 98-113Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Epidemiological evidence links chronic bacterial infections to the increased incidence of certain types of cancer but the molecular mechanisms by which bacteria contribute to tumour initiation and progression are still poorly characterized. Here we show that chronic exposure to the genotoxin cytolethal distending toxin (CDT) of Gram-negative bacteria promotes genomic instability and acquisition of phenotypic properties of malignancy in fibroblasts and colon epithelial cells. Cells grown for more than 30 weeks in the presence of sublethal doses of CDT showed increased mutation frequency, and accumulation of chromatin and chromosomal aberrations in the absence of significant alterations of cell cycle distribution, decreased viability or senescence. Cell survival was dependent on sustained activity of the p38 MAP kinase. The ongoing genomic instability was associated with impaired activation of the DNA damage response and failure to efficiently activate cell cycle checkpoints upon exposure to genotoxic stress. Independently selected sublines showed enhanced anchorage-independent growth as assessed by the formation of colonies in semisolid agarose. These findings support the notion that chronic infection by CDT-producing bacteria may promote malignant transformation, and point to the impairment of cellular control mechanisms associated with the detection and repair of DNA damage as critical events in the process.

  • 297. Guidi, Riccardo
    et al.
    Levi, Laura
    Rouf, Syed Fazle
    Puiac, Speranta
    Rhen, Mikael
    Frisan, Teresa
    Department of Cell and Molecular Biology, Karolinska Institute, Stockholm, Sweden..
    Salmonella enterica delivers its genotoxin through outer membrane vesicles secreted from infected cells2013Inngår i: Cellular Microbiology, ISSN 1462-5814, E-ISSN 1462-5822, Vol. 15, nr 12, s. 2034-2050Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Cytolethal-distending toxins (CDTs) belong to a family of DNA damage inducing exotoxins that are produced by several Gram-negative bacteria. Salmonella enterica serovar Typhi expresses its CDT (named as Typhoid toxin) only in the Salmonella-containing vacuole (SCV) of infected cells, which requires its export for cell intoxication. The mechanisms of secretion, release in the extracellular space and uptake by bystander cells are poorly understood. We have addressed these issues using a recombinant S. Typhimurium strain, MC71-CDT, where the genes encoding for the PltA, PltB and CdtB subunits of the Typhoid toxin are expressed under control of the endogenous promoters. MC71-CDT grown under conditions that mimic the SCV secreted the holotoxin in outer membrane vesicles (OMVs). Epithelial cells infected with MC71-CDT also secreted OMVs-like vesicles. The release of these extracellular vesicles required an intact SCV and relied on anterograde transport towards the cellular cortex on microtubule and actin tracks. Paracrine internalization of Typhoid toxin-loaded OMVs by bystander cells was dependent on dynamin-1, indicating active endocytosis. The subsequent induction of DNA damage required retrograde transport of the toxin through the Golgi complex. These data provide new insights on the mode of secretion of exotoxins by cells infected with intracellular bacteria.

  • 298.
    Guo, Xiong
    et al.
    Institute of Endemic Diseases, Medical College of Xi'an Jiaotong University, Xi'an, China.
    Aigner, Thomas
    Department of Pathology, University of Erlangen-Nürnberg, Erlangen, Germany.
    Lammi, Pirkko
    Institute of Molecular Medicine, University of Erlangen-Nürnberg, Erlangen, Germany.
    Lammi, Mikko
    Institute of Molecular Medicine, University of Erlangen-Nürnberg, Erlangen, Germany.
    Zhang, Ju Ren
    Institute of Endemic Diseases, Medical College of Xi'an Jiaotong University, Xi'an, China.
    Wang, Jian Ming
    Institute of Endemic Diseases, Medical College of Xi'an Jiaotong University, Xi'an, China.
    Zhang, Fu Qiang
    Institute of Endemic Diseases, Medical College of Xi'an Jiaotong University, Xi'an, China.
    von der Mark, Klaus
    Institute of Molecular Medicine, University of Erlangen-Nürnberg, Erlangen, Germany.
    Investigation of abnormal chondrocyte differentiation and differential expression of collagen types I, II, III, VI and X in articular cartilage from patients with Kashin-Beck disease1998Inngår i: Chinese Journal of Pathology, Vol. 27, s. 19-21Artikkel i tidsskrift (Fagfellevurdert)
    Abstract
  • 299.
    Guo, Xiong
    et al.
    Institute of Endemic Diseases, Medical College of Xi'an Jiaotong University, Xi'an, China.
    Lammi, Mikko
    Institute of Molecular Medicine, University of Erlangen-Nurnberg, Erlangen, Germany.
    Aigner, Thomas
    Department of Pathology, University of Erlangen-Nurnberg, Erlangen, Germany.
    Lammi, Pirkko
    Institute of Molecular Medicine, University of Erlangen-Nurnberg, Erlangen, Germany.
    Vornehm, Silvia
    Institute of Molecular Medicine, University of Erlangen-Nurnberg, Erlangen, Germany.
    Yu, Zhidao
    Institute of Endemic Diseases, Medical College of Xi'an Jiaotong University, Xi'an, China.
    Xiong, Yongmin
    Institute of Endemic Diseases, Medical College of Xi'an Jiaotong University, Xi'an, China.
    von der Mark, Klaus
    Institute of Molecular Medicine, University of Erlangen-Nurnberg, Erlangen, Germany.
    Effect of low selenium on chondrocyte differentation and differential expression of collagen types I, II and X in articular cartilage from mini-pigs2000Inngår i: Journal of Xi'an Medical University, ISSN 1671-8259, Vol. 12, s. 108-112Artikkel i tidsskrift (Annet vitenskapelig)
  • 300. Guo, Yijie
    et al.
    Zhou, Yuan
    Yan, Siqi
    Qu, Chengjuan
    Umeå universitet, Medicinska fakulteten, Institutionen för integrativ medicinsk biologi (IMB).
    Wang, Liyun
    Guo, Xiong
    Han, Jing
    Decreased expression of CHST-12, CHST-13, and UST in the proximal interphalangeal joint cartilage of school-age children with Kashin-Beck disease: an endemic osteoarthritis in China caused by selenium deficiency2019Inngår i: Biological Trace Element Research, ISSN 0163-4984, E-ISSN 1559-0720, Vol. 191, nr 2, s. 276-285Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The objective of this study is to investigate changes in the expression of enzymes involved in chondroitin sulfate (CS) sulfation in distal articular surface of proximal interphalangeal joint isolated from school-age children patients with Kashin-Beck disease (KBD), using normal children as controls. Articular cartilage samples were collected from four normal and four KBD children (7-12 years old), and these children were assigned to control and KBD groups. Hematoxylin and eosin (H&E), toluidine blue (TB), and immunohistochemical (IHC) stainings were utilized to evaluate changes in joint pathology and expression of enzymes involved in CS sulfation, including carbohydrate sulfotransferase 12 (CHST-12), carbohydrate sulfotransferase 13 (CHST-13), and uronyl 2-O-sulfotransferase (UST). The correspondence results were examined by semi-quantitative analysis. Compared with the control group, the KBD group showed the following: a significant decrease of total chondrocytes in superficial, middle, and deep layers and deposition of sulfated glycosaminoglycans in extracellular matrix of KBD cartilage were observed; positive staining chondrocytes of CHST-12, CHST-13, and UST were significantly less in superficial zone of KBD cartilage; and CHST-13 positive staining chondrocytes was reduced in deep zone of KBD cartilage. In contrast, the positive staining rates of CHST-12, CHST-13, and UST in KBD were significantly higher than those in the control group. The decreased expression of these enzymes and the physiologic compensatory reaction may be the signs of early-stage KBD. The alterations of CS structure modifying sulfotransferases in finger articular cartilage might play an important role in the onset and pathogenesis of school-age KBD children.

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