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Control of Drosophila blood cell activation via toll signaling in the fat body
Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). (Hultmark)ORCID iD: 0000-0003-0772-3431
Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Institute of Biomedical Technology (BioMediTech), University of Tampere, Tampere, Finland.
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2014 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 9, no 8, e102568Article in journal (Refereed) Published
Abstract [en]

The Toll signaling pathway, first discovered in Drosophila, has a well-established role in immune responses in insects as well as in mammals. In Drosophila, the Toll-dependent induction of antimicrobial peptide production has been intensely studied as a model for innate immune responses in general. Besides this humoral immune response, Toll signaling is also known to activate blood cells in a reaction that is similar to the cellular immune response to parasite infections, but the mechanisms of this response are poorly understood. Here we have studied this response in detail, and found that Toll signaling in several different tissues can activate a cellular immune defense, and that this response does not require Toll signaling in the blood cells themselves. Like in the humoral immune response, we show that Toll signaling in the fat body (analogous to the liver in vertebrates) is of major importance in the Toll-dependent activation of blood cells. However, this Toll-dependent mechanism of blood cell activation contributes very little to the immune response against the parasitoid wasp, Leptopilina boulardi, probably because the wasp is able to suppress Toll induction. Other redundant pathways may be more important in the defense against this pathogen.

Place, publisher, year, edition, pages
Public library of science , 2014. Vol. 9, no 8, e102568
Keyword [en]
Drosophila, toll signaling, immunity, hemocytes, fat body
National Category
Cell and Molecular Biology Microbiology
Research subject
Molecular Biology; biology
Identifiers
URN: urn:nbn:se:umu:diva-92747DOI: 10.1371/journal.pone.0102568ISI: 000339993900003PubMedID: 25102059OAI: oai:DiVA.org:umu-92747DiVA: diva2:742736
Funder
Swedish Research Council
Available from: 2014-09-02 Created: 2014-09-02 Last updated: 2017-12-05Bibliographically approved
In thesis
1. Toll-mediated cellular immune response in Drosophila melanogaster
Open this publication in new window or tab >>Toll-mediated cellular immune response in Drosophila melanogaster
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Insects are amongst the most abundant and diversified multi-cellular organisms on earth. As pollinators of the vast majority of our food crops their socio-economic value is hard to overestimate. Although many pest and pathogens of the honeybee have been known for decades, we still fail to explain the huge losses of honeybee colonies in recent years.At the beginning of my PhD studies, I investigated the effect that senescence and the age-related caste dimorphisms have on two basic parameters of the adult honeybee’s immune system, namely blood cell concentration and the activity of the phenoloxidase cascade. Realizing the limitations of working on an organism for which (at the time) no sequenced genome or molecular tools were available, I switched labs to work on Drosophila melanogaster. The fruit fly has proven to be a particularly useful model system to identify and study genes critical for both the innate immune response itself, as well as the signaling pathways regulating it. For the main part of my thesis, I used the tissue-specific expression of fluorescent markers to visualize segmentally aligned bands of sessile blood cells in the Drosophila larva. This phenotype is disturbed in larvae heterozygote for a gain-of-function mutation in the Toll pathway called Tl10b. In a genetic screen, I scored the ability of genomic mutations to modify the Tl10b loss of bands phenotype. I identified five genomic regions that suppressed the disturbed band pattern of sessile blood cells, and in three of these regions I mapped down this phenotype to single gene level. Two genes are involved in intracellular vesicle trafficking (Rab23 and ird1) and one is activated at the onset of metamorphosis (hdc). To confirm the experimental model, I tested the role of another negative regulator of the Toll pathway. I used tissue specific GAL4 fly lines to express RNAi silencing constructs targeting Gprk2 expression in vivo. This led to an unexpected and novel discovery. Even though blood cells give rise to the most apparent phenotypes in the Tl10b larva, the main source for the immune signal is the fat body. This indicates that besides the humoral response, also in cell based immunity this organ plays a major role. Based on this finding, I could show that the modification of Tl10b blood cell phenotypes caused by loss of ird1 expression are due the role this gene plays in autophagy cell motility. The improved understanding of these basic and evolutionary highly conserved mechanisms will undoubtedly help in fending off infectious disease in both man and honeybees in the future.

Place, publisher, year, edition, pages
Umeå: Umeå universitet, 2014. 81 p.
Series
Umeå University medical dissertations, ISSN 0346-6612 ; 1670
National Category
Biochemistry and Molecular Biology
Research subject
Molecular Biology
Identifiers
urn:nbn:se:umu:diva-92751 (URN)978-91-7601-116-4 (ISBN)
Public defence
2014-09-26, Major Groove, building 6L, molekylärbiologi, Umeå University, Umeå, 09:00 (English)
Opponent
Supervisors
Available from: 2014-09-05 Created: 2014-09-02 Last updated: 2014-09-08Bibliographically approved
2. Activation of the Cellular Immune Response in Drosophila melanogaster Larvae
Open this publication in new window or tab >>Activation of the Cellular Immune Response in Drosophila melanogaster Larvae
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

During the last 40 years, Drosophila melanogaster has become an invaluable tool in understanding innate immunity. The innate immune system of Drosophila consists of a humoral and a cellular component. While many details are known about the humoral immune system, our knowledge about the cellular immune system is comparatively small. Blood cells or hemocytes constitute the cellular immune system. Three blood types have been described for Drosophila larvae. Plasmatocytes are phagocytes with a plethora of functions. Crystal cells mediate melanization and contribute to wound healing. Plasmatocytes and crystal cells constitute the blood cell repertoire of a healthy larva, whereas lamellocytes are induced in a demand-adapted manner after infection with parasitoid wasp eggs. They are involved in the melanotic encapsulation response against parasites and form melanotic nodules that are also referred to as tumors.

In my thesis, I focused on unraveling the mechanisms of how the immune system orchestrates the cellular immune response. In particular, I was interested in the hematopoiesis of lamellocytes.

In Article I, we were able to show that ectopic expression of key components of a number of signaling pathways in blood cells induced the development of lamellocytes, led to a proliferative response of plasmatocytes, or to a combination of lamellocyte activation and plasmatocyte proliferation.

In Article II, I combined newly developed fluorescent enhancer-reporter constructs specific for plasmatocytes and lamellocytes and developed a “dual reporter system” that was used in live microscopy of fly larvae. In addition, we established flow cytometry as a tool to count total blood cell numbers and to distinguish between different blood cell types. The “dual reporter system” enabled us to differentiate between six blood cell types and established proliferation as a central feature of the cellular immune response. The combination flow cytometry and live imaging increased our understanding of the tempo-spatial events leading to the cellular immune reaction.

In Article III, I developed a genetic modifier screen to find genes involved in the hematopoiesis of lamellocytes. I took advantage of the gain-of-function phenotype of the Tl10b mutation characterized by an activated cellular immune system, which induced the formation blood cell tumors. We screened the right arm of chromosome 3 for enhancers and suppressors of this mutation and uncovered ird1.

Finally in Article IV, we showed that the activity of the Toll signaling pathway in the fat body, the homolog of the liver, is necessary to activate the cellular immune system and induce lamellocyte hematopoiesis.

Place, publisher, year, edition, pages
Umeå: Umeå University, 2015. 41 p.
Series
Umeå University medical dissertations, ISSN 0346-6612 ; 1741
Keyword
Drosophila melanogaster, immunity, blood cells, hematopoiesis, flow cytometry, in vivo imaging, genetic screens, Tl10b, fat body, Toll signaling
National Category
Immunology
Research subject
Molecular Biology
Identifiers
urn:nbn:se:umu:diva-106981 (URN)978-91-7601-317-5 (ISBN)
Public defence
2015-09-07, Major Groove, Byggnad 6L, Umeå, 12:30 (English)
Opponent
Supervisors
Available from: 2015-08-17 Created: 2015-08-13 Last updated: 2015-08-13Bibliographically approved

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Schmid, Martin RudolfAnderl, InesHultmark, Dan

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