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Signaling and transcriptional regulation of antimicrobial peptide genes in Drosophila melanogaster
Stockholm University, Faculty of Science, Department of Molecular Biology and Functional Genomics.
2006 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Insects rely solely on innate immune reactions for protection against infect-ing microbes in their environment. In Drosophila, one major defense mechanism is the production of a battery of antimicrobial peptides (AMPs). The expression of AMPs is primarily regulated at the level of transcription and constitutes both constitutive expression in a tissue-specific manner and inducible systemic expression in response to infection. The aim of my thesis has been to investigate the regulation of AMP gene expression at different levels. I have studied a novel cis-regulatory element, Region 1 (R1) found in the proximal promoter of all Cecropin genes in Drosophila melanogaster, as well as in other species of Drosophila. We found that the R1 element was important for the expression of CecropinA1 (CecA1) both in vitro and in vivo. A signaling-dependent R1-binding activity (RBA) was identified in nuclear extracts from Drosophila cells and flies. The molecular nature of the RBA, has despite considerable effort, not yet been identified. I also have studied the role of the JNK pathway in transcriptional regulation of AMP genes. The role of the JNK pathway in the regulation of AMP genes has long been elusive, however, in this study we showed that the pathway is directly involved in the expression of AMP genes. Analysis of cells mutant for JNK pathway components showed severely reduced AMP gene expression. Fur-thermore, over-expression of a JNK pathway-inhibitor also inhibited AMP gene expression. Lastly, I have studied transcription factors that have not previously been implicated in transcriptional regulation of AMP genes. In a yeast screen, three members of the POU family of transcription factors were identified as regulators of CecA1. Two of them, Drifter (Dfr) and POU do-main protein 1 (Pdm1) were further characterized. We showed that Dfr was able to promote AMP gene expression in the absence of infection, suggest-ing it to play a role in constitutive expression of AMP genes. Indeed, down-regulation of Dfr expression using RNAi severely reduced the constitutive expression of AMP genes in the male ejaculatory duct. We also identified an enhancer element important for Dfr-mediated expression of CecA1. Pdm1, on the other hand, was shown to be important for the systemic expression of AMP genes. In Pdm1 mutant flies, several AMP genes are systemically expressed even in the absence of infection, suggesting that Pdm1 works as a repressor of those genes. However, at least on AMP gene, AttacinA (AttA) requires Pdm1 for its expression, suggesting that Pdm1 works as an activator for this gene. Upon infection, Pdm1 was rapidly degraded, but, regenerated shortly after infection. We propose that the degradation of Pdm1 is important for the activation of the Pdm1-repressed genes and that regeneration sup-ports the expression of AttA.

Place, publisher, year, edition, pages
Stockholm: Institutionen för molekylärbiologi och funktionsgenomik , 2006. , 70 p.
Keyword [en]
antimicrobial peptides, drosophila, immunity, transcriptiona regulation
National Category
Biochemistry and Molecular Biology
Identifiers
URN: urn:nbn:se:su:diva-1051ISBN: 91-7155-263-4 (print)OAI: oai:DiVA.org:su-1051DiVA: diva2:189316
Public defence
2006-06-08, sal G, Arrheniuslaboratorierna, Svante Arrhenius väg 14-18, Stockholm, 10:00
Opponent
Supervisors
Available from: 2006-05-01 Created: 2006-05-01 Last updated: 2011-03-28Bibliographically approved
List of papers
1. Isolation of regulators of Drosophila immune defense genes by a double interaction screen in yeast
Open this publication in new window or tab >>Isolation of regulators of Drosophila immune defense genes by a double interaction screen in yeast
2007 (English)In: Insect Biochemistry and Molecular Biology, ISSN 0965-1748, E-ISSN 1879-0240, Vol. 37, no 3, 202-212 p.Article in journal (Refereed) Published
Abstract [en]

Innate immunity is a universal and ancient defense system in metazoans against microorganisms. Antimicrobial peptides, which are synthesized both in insects and humans, constitute an endogenous, gene-encoded defense arsenal. In Drosophila, antimicrobial peptides, such as the potent cecropins, are expressed both constitutively in barrier epithelia, as well as systemically in response to infection. Rel/NF-κB proteins are well-known regulators of antimicrobial peptide genes, but very few Rel/NF-κB co-factors and/or tissue-specific regulators have been identified. We performed a double interaction screen in yeast to isolate Drosophila cDNAs coding for direct regulators, as well as Dif co-regulators, of the CecropinA1 gene. Three classes of positive cDNA clones corresponding to 15 Drosophila genes were isolated and further characterized. One of the Dif-independent cDNAs encoded the Rel/NF-κB protein Relish; a well-known activator of antimicrobial peptide genes in Drosophila, demonstrating the applicability of this type of screen for isolating regulators of immune defense. Most interestingly, three transcription factors belonging to the POU domain class of homeodomain proteins, Pdm1, Pdm2 and Dfr/Vvl were isolated as Dif-interacting partners, and subsequently verified as regulators of CecA1 expression in Drosophila cells. The importance of POU proteins in development and differentiation in Drosophila and mammals is well documented, but their role in regulation of Drosophila immune defense genes is a new and essential finding.

Keyword
Innate immunity, Cecropin, Gene regulation, POU transcription factors, NF-kappaB, Antimicrobial peptides, Yeast screen
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:su:diva-24100 (URN)10.1016/j.ibmb.2006.10.008 (DOI)
Available from: 2007-02-01 Created: 2007-02-01 Last updated: 2011-05-23Bibliographically approved
2. Functional characterization of a novel promoter element required for an innate immune response in Drosophila
Open this publication in new window or tab >>Functional characterization of a novel promoter element required for an innate immune response in Drosophila
2003 In: Molecular and Cellular Biology, ISSN 0270-7306, Vol. 23, no 22, 8272-8281 p.Article in journal (Refereed) Published
Identifiers
urn:nbn:se:su:diva-22716 (URN)
Note
Part of urn:nbn:se:su:diva-1051Available from: 2006-05-01 Created: 2006-05-01Bibliographically approved
3. Cooperative Control of Drosophila Immune Responses by the JNK and NF-κB Signaling Pathways
Open this publication in new window or tab >>Cooperative Control of Drosophila Immune Responses by the JNK and NF-κB Signaling Pathways
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Manuscript (Other academic)
Identifiers
urn:nbn:se:su:diva-22717 (URN)
Note
Part of urn:nbn:se:su:diva-1051Available from: 2006-05-01 Created: 2006-05-01 Last updated: 2010-01-13Bibliographically approved
4. The POU protein Drifter activates antimicrobial peptide gene expression in Drosophila
Open this publication in new window or tab >>The POU protein Drifter activates antimicrobial peptide gene expression in Drosophila
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Manuscript (Other academic)
Identifiers
urn:nbn:se:su:diva-22719 (URN)
Note
Part of urn:nbn:se:su:diva-1051Available from: 2006-05-01 Created: 2006-05-01 Last updated: 2011-03-28Bibliographically approved
5. The Drosophila transcription factor Pdm1 inhibits antimicrobial peptide gene expression
Open this publication in new window or tab >>The Drosophila transcription factor Pdm1 inhibits antimicrobial peptide gene expression
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Manuscript (Other academic)
Identifiers
urn:nbn:se:su:diva-22720 (URN)
Note
Part of urn:nbn:se:su:diva-1051Available from: 2006-05-01 Created: 2006-05-01 Last updated: 2011-03-28Bibliographically approved

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