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Publications (9 of 9) Show all publications
Pu, L., Wang, J., Lu, Q., Nilsson, L., Philbrook, A., Pandey, A., . . . Chen, C. (2023). Dissecting the genetic landscape of GPCR signaling through phenotypic profiling in  C. elegans. Nature Communications, 14, Article ID 8410.
Open this publication in new window or tab >>Dissecting the genetic landscape of GPCR signaling through phenotypic profiling in  C. elegans
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2023 (English)In: Nature Communications, E-ISSN 2041-1723, Vol. 14, article id 8410Article in journal (Refereed) Published
Abstract [en]

G protein-coupled receptors (GPCRs) mediate responses to various extracellular and intracellular cues. However, the large number of GPCR genes and their substantial functional redundancy make it challenging to systematically dissect GPCR functions in vivo. Here, we employ a CRISPR/Cas9-based approach, disrupting 1654 GPCR-encoding genes in 284 strains and mutating 152 neuropeptide-encoding genes in 38 strains in C. elegans. These two mutant libraries enable effective deorphanization of chemoreceptors, and characterization of receptors for neuropeptides in various cellular processes. Mutating a set of closely related GPCRs in a single strain permits the assignment of functions to GPCRs with functional redundancy. Our analyses identify a neuropeptide that interacts with three receptors in hypoxia-evoked locomotory responses, unveil a collection of regulators in pathogen-induced immune responses, and define receptors for the volatile food-related odorants. These results establish our GPCR and neuropeptide mutant libraries as valuable resources for the C. elegans community to expedite studies of GPCR signaling in multiple contexts.

Place, publisher, year, edition, pages
Springer Nature, 2023
National Category
Neurosciences
Identifiers
urn:nbn:se:umu:diva-217489 (URN)10.1038/s41467-023-44177-z (DOI)38110404 (PubMedID)2-s2.0-85180225404 (Scopus ID)
Funder
Swedish Research Council, 2018-02914Swedish Research Council, 2021-06602Swedish Research Council, 2018-02216
Note

Originally included in thesis in manuscript form. 

Available from: 2023-12-05 Created: 2023-12-05 Last updated: 2024-01-05Bibliographically approved
Pakharukova, N., Malmi, H., Tuittila, M., Dahlberg, T., Ghosal, D., Chang, Y.-W., . . . Zavialov, A. V. (2022). Archaic chaperone-usher pili self-secrete into superelastic zigzag springs. Nature, 609(7926), 335-340
Open this publication in new window or tab >>Archaic chaperone-usher pili self-secrete into superelastic zigzag springs
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2022 (English)In: Nature, ISSN 0028-0836, E-ISSN 1476-4687, Vol. 609, no 7926, p. 335-340Article in journal (Refereed) Published
Abstract [en]

Adhesive pili assembled via the chaperone-usher pathway (CUP) are hair-like appendages that mediate host tissue colonization and biofilm formation of Gram-negative bacteria 1-3. Archaic CUP pili, the most diverse and widespread CUP adhesins, are promising vaccine and drug targets due to their prevalence in the most troublesome multidrug-resistant (MDR) pathogens 1,4,5. However, their architecture and assembly-secretion process remain unknown. Here, we present the 3.4 Å resolution cryo-electron microscopy structure of the prototypical archaic Csu pilus that mediates biofilm formation of Acinetobacter baumannii, a notorious MDR nosocomial pathogen. In contrast to the thick helical tubes of the classical type 1 and P pili, archaic pili assemble into a conceptually novel ultrathin zigzag architecture secured by an elegant clinch mechanism. The molecular clinch provides the pilus with high mechanical stability as well as superelasticity, a property observed now for the first time in biomolecules, while enabling a more economical and faster pilus production. Furthermore, we demonstrate that clinch formation at the cell surface drives pilus secretion through the outer membrane. These findings suggest that clinch-formation inhibitors might represent a new strategy to fight MDR bacterial infections.

Place, publisher, year, edition, pages
Nature Publishing Group, 2022
National Category
Microbiology in the medical area Other Physics Topics Structural Biology
Research subject
Microbiology
Identifiers
urn:nbn:se:umu:diva-198528 (URN)10.1038/s41586-022-05095-0 (DOI)000844487100001 ()35853476 (PubMedID)2-s2.0-85136986109 (Scopus ID)
Funder
Swedish Research Council, 2019-04016The Kempe Foundations, JCK-1724Swedish Research Council, 2019-01720Swedish Research Council, 2016-04451
Available from: 2022-08-08 Created: 2022-08-08 Last updated: 2023-03-24Bibliographically approved
Nadeem, A., Berg, A., Pace, H., Alam, A., Toh, E., Ådén, J., . . . Wai, S. N. (2022). Protein-lipid interaction at low pH induces oligomerization of the MakA cytotoxin from Vibrio cholerae. eLIFE, 11, Article ID e73439.
Open this publication in new window or tab >>Protein-lipid interaction at low pH induces oligomerization of the MakA cytotoxin from Vibrio cholerae
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2022 (English)In: eLIFE, E-ISSN 2050-084X, Vol. 11, article id e73439Article in journal (Refereed) Published
Abstract [en]

The α-pore-forming toxins (α-PFTs) from pathogenic bacteria damage host cell membranes by pore formation. We demonstrate a remarkable, hitherto unknown mechanism by an α-PFT protein from Vibrio cholerae. As part of the MakA/B/E tripartite toxin, MakA is involved in membrane pore formation similar to other α-PFTs. In contrast, MakA in isolation induces tube-like structures in acidic endosomal compartments of epithelial cells in vitro. The present study unravels the dynamics of tubular growth, which occurs in a pH-, lipid-, and concentration-dependent manner. Within acidified organelle lumens or when incubated with cells in acidic media, MakA forms oligomers and remodels membranes into high-curvature tubes leading to loss of membrane integrity. A 3.7 Å cryo-electron microscopy structure of MakA filaments reveals a unique protein-lipid superstructure. MakA forms a pinecone-like spiral with a central cavity and a thin annular lipid bilayer embedded between the MakA transmembrane helices in its active α-PFT conformation. Our study provides insights into a novel tubulation mechanism of an α-PFT protein and a new mode of action by a secreted bacterial toxin.

Place, publisher, year, edition, pages
eLife Sciences Publications, Ltd, 2022
Keywords
Vibrio cholerae, MakA, lipid
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-192300 (URN)10.7554/eLife.73439 (DOI)2-s2.0-85124321786 (Scopus ID)
Funder
Swedish Research Council, 2018–02914Swedish Research Council, 2016–05009Swedish Research Council, 2019–01720Swedish Research Council, 2016–06963Swedish Research Council, 2019–02011Swedish Cancer Society, 2017–419Swedish Cancer Society, 2020–711The Kempe Foundations, JCK-1728The Kempe Foundations, SMK-1756.2The Kempe Foundations, SMK-1553The Kempe Foundations, JCK-1724The Kempe Foundations, SMK-1961Knut and Alice Wallenberg FoundationFamiljen Erling-Perssons Stiftelse
Available from: 2022-02-08 Created: 2022-02-08 Last updated: 2024-01-12Bibliographically approved
Nadeem, A., Nagampalli, R., Toh, E., Alam, A., Myint, S. L., Heidler, T., . . . Persson, K. (2021). A tripartite cytolytic toxin formed by Vibrio cholerae proteins with flagellum-facilitated secretion. Proceedings of the National Academy of Sciences of the United States of America, 118(47), Article ID e2111418118.
Open this publication in new window or tab >>A tripartite cytolytic toxin formed by Vibrio cholerae proteins with flagellum-facilitated secretion
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2021 (English)In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 118, no 47, article id e2111418118Article in journal (Refereed) Published
Abstract [en]

Vibrio cholerae, responsible for outbreaks of cholera disease, is a highly motile organism by virtue of a single flagellum. We describe how the flagellum facilitates the secretion of three V. cholerae proteins encoded by a hitherto-unrecognized genomic island. The proteins MakA/B/E can form a tripartite toxin that lyses erythrocytes and is cytotoxic to cultured human cells. A structural basis for the cytolytic activity of the Mak proteins was obtained by X-ray crystallography. Flagellum-facilitated secretion ensuring spatially coordinated delivery of Mak proteins revealed a role for the V. cholerae flagellum considered of particular significance for the bacterial environmental persistence. Our findings will pave the way for the development of diagnostics and therapeutic strategies against pathogenic Vibrionaceae.

National Category
Biochemistry and Molecular Biology
Research subject
Biochemistry
Identifiers
urn:nbn:se:umu:diva-191257 (URN)10.1073/pnas.2111418118 (DOI)000727697700014 ()34799450 (PubMedID)2-s2.0-85121209218 (Scopus ID)
Funder
Swedish Research Council, 2016-05009Swedish Research Council, 2018-02914Swedish Research Council, 2019-01720Swedish Research Council, 2007-08673The Kempe Foundations, SMK-1756.2The Kempe Foundations, SMK-1553The Kempe Foundations, JCK-1728Swedish Cancer Society, 2017-419The Kempe Foundations, SMK-1961Swedish Research Council
Available from: 2022-01-12 Created: 2022-01-12 Last updated: 2023-05-11Bibliographically approved
Myint, S. L., Zlatkov, N., Aung, K. M., Toh, E., Sjöström, A. E., Nadeem, A., . . . Wai, S. N. (2021). Ecotin and LamB in Escherichia coli influence the susceptibility to Type VI secretion-mediated interbacterial competition and killing by Vibrio cholerae. Biochimica et Biophysica Acta - General Subjects, 1865(7), Article ID 129912.
Open this publication in new window or tab >>Ecotin and LamB in Escherichia coli influence the susceptibility to Type VI secretion-mediated interbacterial competition and killing by Vibrio cholerae
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2021 (English)In: Biochimica et Biophysica Acta - General Subjects, ISSN 0304-4165, E-ISSN 1872-8006, Vol. 1865, no 7, article id 129912Article in journal (Refereed) Published
Abstract [en]

Background: A prevailing action of the Type VI secretion system (T6SS) in several Gram-negative bacterial species is inter-bacterial competition. In the past several years, many effectors of T6SS were identified in different bacterial species and their involvement in inter-bacterial interactions were described. However, possible defence mechanisms against T6SS attack among prey bacteria were not well clarified yet. Methods: Escherichia coli was assessed for susceptibility to T6SS-mediated killing by Vibrio cholerae. TheT6SS-mediated bacterial killing assays were performed in absence or presence of different protease inhibitors and with different mutant E. coli strains. Expression levels of selected proteins were monitored using SDS-PAGE and immunoblot analyses. Results: The T6SS-mediated killing of E. coli by V. cholerae was partly blocked when the serine protease inhibitor Pefabloc was present. E. coli lacking the periplasmic protease inhibitor Ecotin showed enhanced susceptibility to killing by V. cholerae. Mutations affecting E. coli membrane stability also caused increased susceptibility to killing by V. cholerae. E. coli lacking the maltodextrin porin protein LamB showed reduced susceptibility to killing by V. cholerae whereas E. coli with induced high levels of LamB showed reduced survival in inter-bacterial competition. Conclusions: Our study identified two proteins in E. coli, the intrinsic protease inhibitor Ecotin and the outer membrane porin LamB, that influenced E. coli susceptibility to T6SS-mediated killing by V. cholerae. General significance: We envision that it is feasible to explore these findings to target and modulate their expression to obtain desired changes in inter-bacterial competition in vivo, e.g. in the gastrointestinal microbiome.

Place, publisher, year, edition, pages
Elsevier, 2021
Keywords
Ecotin, Escherichia coli, Interbacterial competition, LamB, T6SS, Vibrio cholerae
National Category
Microbiology in the medical area
Identifiers
urn:nbn:se:umu:diva-182908 (URN)10.1016/j.bbagen.2021.129912 (DOI)000652016000005 ()33892013 (PubMedID)2-s2.0-85104614899 (Scopus ID)
Available from: 2021-05-28 Created: 2021-05-28 Last updated: 2023-09-05Bibliographically approved
Nadeem, A., Alam, A., Toh, E., Myint, S. L., Ur Rehman, Z., Liu, T., . . . Wai, S. N. (2021). Phosphatidic acid-mediated binding and mammalian cell internalization of the Vibrio cholerae cytotoxin MakA. PLoS Pathogens, 17(3), Article ID 1009414.
Open this publication in new window or tab >>Phosphatidic acid-mediated binding and mammalian cell internalization of the Vibrio cholerae cytotoxin MakA
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2021 (English)In: PLoS Pathogens, ISSN 1553-7366, E-ISSN 1553-7374, Vol. 17, no 3, article id 1009414Article in journal (Refereed) Published
Abstract [en]

Vibrio cholerae is a noninvasive intestinal pathogen extensively studied as the causative agent of the human disease cholera. Our recent work identified MakA as a potent virulence factor of V. cholerae in both Caenorhabditis elegans and zebrafish, prompting us to investigate the potential contribution of MakA to pathogenesis also in mammalian hosts. In this study, we demonstrate that the MakA protein could induce autophagy and cytotoxicity of target cells. In addition, we observed that phosphatidic acid (PA)-mediated MakA-binding to the host cell plasma membranes promoted macropinocytosis resulting in the formation of an endomembrane-rich aggregate and vacuolation in intoxicated cells that lead to induction of autophagy and dysfunction of intracellular organelles. Moreover, we functionally characterized the molecular basis of the MakA interaction with PA and identified that the N-terminal domain of MakA is required for its binding to PA and thereby for cell toxicity. Furthermore, we observed that the ΔmakA mutant outcompeted the wild-type V. cholerae strain A1552 in the adult mouse infection model. Based on the findings revealing mechanistic insights into the dynamic process of MakA-induced autophagy and cytotoxicity we discuss the potential role played by the MakA protein during late stages of cholera infection as an anti-colonization factor.

Place, publisher, year, edition, pages
Public Library of Science, 2021
National Category
Microbiology in the medical area
Identifiers
urn:nbn:se:umu:diva-181991 (URN)10.1371/journal.ppat.1009414 (DOI)000631027700007 ()33735319 (PubMedID)2-s2.0-85103129339 (Scopus ID)
Available from: 2021-04-12 Created: 2021-04-12 Last updated: 2023-05-11Bibliographically approved
Ahmad, I., Nygren, E., Khalid, F., Myint, S. L. & Uhlin, B. E. (2020). A Cyclic-di-GMP signalling network regulates biofilm formation and surface associated motility of Acinetobacter baumannii 17978. Scientific Reports, 10(1), Article ID 1991.
Open this publication in new window or tab >>A Cyclic-di-GMP signalling network regulates biofilm formation and surface associated motility of Acinetobacter baumannii 17978
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2020 (English)In: Scientific Reports, E-ISSN 2045-2322, Vol. 10, no 1, article id 1991Article in journal (Refereed) Published
Abstract [en]

Acinetobacter baumannii has emerged as an increasing multidrug-resistant threat in hospitals and a common opportunistic nosocomial pathogen worldwide. However, molecular details of the pathogenesis and physiology of this bacterium largely remain to be elucidated. Here we identify and characterize the c-di-GMP signalling network and assess its role in biofilm formation and surface associated motility. Bioinformatic analysis revealed eleven candidate genes for c-di-GMP metabolizing proteins (GGDEF/EAL domain proteins) in the genome of A. baumannii strain 17978. Enzymatic activity of the encoded proteins was assessed by molecular cloning and expression in the model organisms Salmonella typhimurium and Vibrio cholerae. Ten of the eleven GGDEF/EAL proteins altered the rdar morphotype of S. typhimurium and the rugose morphotype of V. cholerae. The over expression of three GGDEF proteins exerted a pronounced effect on colony formation of A. baumannii on Congo Red agar plates. Distinct panels of GGDEF/EAL proteins were found to alter biofilm formation and surface associated motility of A. baumannii upon over expression. The GGDEF protein A1S_3296 appeared as a major diguanylate cyclase regulating macro-colony formation, biofilm formation and the surface associated motility. AIS_3296 promotes Csu pili mediated biofilm formation. We conclude that a functional c-di-GMP signalling network in A. baumannii regulates biofilm formation and surface associated motility of this increasingly important opportunistic bacterial pathogen.

Place, publisher, year, edition, pages
Nature Publishing Group, 2020
National Category
Microbiology in the medical area
Identifiers
urn:nbn:se:umu:diva-170711 (URN)10.1038/s41598-020-58522-5 (DOI)000559759000022 ()32029764 (PubMedID)2-s2.0-85079062852 (Scopus ID)
Funder
Swedish Research Council, 2015-03007Swedish Research Council, 2015-06824Swedish Research Council, 2016-06598Swedish Research Council, 349-2007-8673Swedish Research Council, 829-2006-7431The Kempe Foundations, JCK-1527The Kempe Foundations, JCK-1724
Available from: 2020-05-13 Created: 2020-05-13 Last updated: 2023-03-24Bibliographically approved
Pakharukova, N., Malmi, H., Tuittila, M., Dahlberg, T., Ghosal, D., Chang, Y.-W., . . . Zavialov, A. V.Archaic chaperone-usher pilus self-secretes into a superelastic zigzag spring architecture.
Open this publication in new window or tab >>Archaic chaperone-usher pilus self-secretes into a superelastic zigzag spring architecture
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(English)Manuscript (preprint) (Other academic)
National Category
Biophysics Biochemistry and Molecular Biology Other Physics Topics
Identifiers
urn:nbn:se:umu:diva-192464 (URN)10.21203/rs.3.rs-936177/v1 (DOI)
Available from: 2022-02-14 Created: 2022-02-14 Last updated: 2022-02-15
Toh, E., Baryalai, P., Nadeem, A., Aung, K. M., Myint, S. L., Uhlin, B. E. & Wai, S. N.Sublytic activity of a pore-forming protein from commensal bacteria causes epigenetic modulation of tumor-affiliated protein expression.
Open this publication in new window or tab >>Sublytic activity of a pore-forming protein from commensal bacteria causes epigenetic modulation of tumor-affiliated protein expression
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Cytolysin A (ClyA) is a pore-forming protein expressed at sublytic levels by a strongly silenced gene in non-pathogenic Escherichia coli, including typical commensal isolates in the intestinal microbiome of healthy mammalian hosts. Upon overproduction, the ClyA-expressing bacteria display a cytolytic phenotype. However, it remains unclear whether sublytic amounts of native ClyA play a role in commensal E. coli-host interactions in vivo. Here, we show that sublytic amounts of ClyA are released via outer membrane vesicles (OMVs) and can affect host cells in a profound and remarkable manner. OMVs isolated from ClyA+ E. coli were rapidly internalised into cultured colon cancer cells. The OMV-associated ClyA inhibited the expression of cancer-activating proteins such as H3K27me3, CXCR4, STAT3, and MDM2 via the EZH2/H3K27me3/miR622/CXCR4 signalling axis. Our results demonstrate that sublytic amounts of ClyA in OMVs from non-pathogenic E. coli can target the stability of the EZH2 protein to modulate epigenetics of colon cancer cells 

Keywords
outer membrane vesicles, pore-forming toxin ClyA, cancer cell epigenetics, non-pathogenic E. coli
National Category
Microbiology in the medical area
Research subject
Medical Cell Biology
Identifiers
urn:nbn:se:umu:diva-208205 (URN)
Available from: 2023-05-11 Created: 2023-05-11 Last updated: 2023-05-11
Organisations
Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0001-5384-3691

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