Molecular and genetic studies on a SipD/IpaD-related virulence spike protein from immune-suppressive particles of a Drosophila parasitoid

<p>Parasitoid wasps are effective biological control agents that have evolved intricate host immune suppression strategies to overcome host immune defenses. The most important anti-parasite defense mechanism is encapsulation, where the host’s blood cells surround the wasp egg. In the <em>Drosophila</em>/<em>Leptopilina</em> host-parasitoid system, extracellular vesicles (EVs) are oviposited along with the wasp egg. The EVs possess spikes and contain hundreds of proteins, some of which enable wasp eggs to escape encapsulation by changing the morphology and function of hemocytes, or killing them. Previous studies have identified an abundant surface/spike protein (SSp40) from <em>Leptopilina heterotoma</em> EVs. While p40, a predicted membrane protein, is synthesized in the secretory cells of the venom gland apparatus, it is delivered into the host via EVs and taken up by both the macrophage and lamellocyte hemocyte types. Previous experiments suggested that p40, while still on the EV, may mediate direct EV-lamellocyte interaction and promote lamellocyte shape change and lysis. Structurally, p40 resembles the needle-tip proteins SipD and IpaD on the bacterial type 3 secretion systems (T3SS) of Gram negative bacteria <em>Salmonella</em> and <em>Shigella</em>. The bacterial T3SS delivers effector proteins into mammalian cells. To understand p40’s effects on host hemocytes, we expressed p40-RFP fusion transgenes in wild type and<em> hopscotch^{Tumoous-lethal}</em>(<em>hop^Tum-l</em>) mutant <em>Drosophila </em>larvae. <em>hop^Tum-l</em> mutant larvae have overabundant hemocytes, that form small tumors. The mutation is semi-lethal. Expression of the membrane-bound and secreted forms of p40 containing the SipD/IpaD-like domain increased viability of mutants by more than six-fold and relieved their tumor burden by more than 50%—the latter parameter being a measure of cellular immunity. Our structural analysis of the conserved p40 domain from <em>L. heterotoma</em> Gotheron strain from Southern France revealed only 84% sequence identity with p40 from the Lh14 strain, with 29 of the 41 amino acid substitutions being non-conservative. Homology modeling revealed that while the Lh14 and LhGoth p40 proteins show complete and precise alignment, the LhGoth protein has four additional 3₁₀ helices. These results suggest possible differences in protein folding and protein-membrane interactions in the two strains. Altogether, these observations confirm an immune-suppressive function for p40, independently of other EV proteins, possibly via direct effects on host hemocytes. They contribute to our understanding of p40’s role in parasitism by <em>L. heterotoma </em>and in the development of its host range.</p>

Name thesis:degree_name

Master of Science (M.S.)

Level thesis:degree_level

Thesis

Discipline thesis:degree_discipline

Biology

Year dc:date.available

2021

Author dc:creator

• Sevilla, Carlo Miguel A

Contributors dc:contributor

• Shubha Govind
• Shaneen Singh
• Amy Berkov

• host-parasitoid interactions
• IpaD/SipD-like virulence proteins
• immune suppressive extracellular vesicles
• Drosophila melanogaster hemocytes
• wasp egg encapsulation
• innate immunity
• Biochemistry
• Bioinformatics
• Biotechnology
• Cell Biology
• Genomics
• Immunity
• Molecular Biology
• Molecular Genetics
• Other Ecology and Evolutionary Biology
• Pathogenic Microbiology

Repository record dc:identifier

https://academicworks.cuny.edu/cc_etds_theses/916

OAI identifier oai:identifier

oai:academicworks.cuny.edu:cc_etds_theses-1987