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Authors: Garg, Aprajita
Advisors: Wu, Louisa P
Department/Program: Cell Biology & Molecular Genetics
Type: Dissertation
Sponsors: Digital Repository at the University of Maryland
University of Maryland (College Park, Md.)
Subjects: Immunology
Cellular biology
Keywords: Phagosome maturation
Staphylococcus aureus
Issue Date: 2012
Abstract: Phagocytosis is a vital component of <italic>Drosophila</italic> cellular immunity. The sequential interaction of a phagosome with the endocytic compartments entails docking and fusion and requires the role of many RabGTPases. We identified Rab14 as a possible regulator of phagosome maturation in an <italic>in vivo</italic> screen. Expression studies indicated the presence of Rab14 in hemocytes in both larval and adult stages. To further characterize the role of Rab14 during phagocytosis, a <italic>Rab14<super>null</super></italic> mutant was generated. The <italic>Rab14</italic> mutant displayed no difference in the phagocytic uptake of <italic>E.coli</italic> or <italic>S.aureus</italic>, however the kinetics of phagosome maturation for both microbes was decreased. Cell biology studies indicated delayed acquisition of the late endosomal marker, Rab7 and the lysosomal marker, Spinster, on the <italic>S.aureus</italic> phagosome. Since recruitment of the early endosomal marker Rab5 was not affected, we concluded Rab14 functions downstream to Rab5 and is essential for efficient Rab7 recruitment on microbial phagosomes. Rab14 itself was recruited during both the early and late phagosomal stages and in accordance with that, it showed colocalization with both Rab5 and Rab7 on endosomes. <italic>Rab14</italic> mutants also showed a defect in endosomal maturation, suggesting Rab14 is also essential during endocytosis. <italic>Rab14</italic> mutants demonstrated defects in the induction of the specific antimicrobial peptides Cecropin and Defensin in response to <italic>S.aureus</italic> infection, indicating a possible crosstalk between the cellular and humoral response. Along with that, <italic>Rab14</italic> mutants also showed defects in the expression of the Toll ligand Spatzle, in response to infection. Hence we propose Rab14 regulates expression of specific antimicrobial peptides through the cytokine Spatzle. The inefficient cellular and humoral responses result in increased susceptibility of <italic>Rab14</italic> mutants to bacterial infection. This could be rescued by hemocyte-specific expression of <italic>Rab14</italic>, suggesting its role in the hemocytes is essential for the immune response against <italic>S. aureus</italic>. Our study identifying a role for Rab14 during phagosomal maturation gives insight on the pathogenesis of <italic>M.tuberculosis</italic>, a pathogen which manipulates Rab14 to subvert phagosome maturation. Moreover it identifies Rab14 function during cellular and humoral responses as an integral component of the Drosophila immune response against <italic>S.aureus</italic>.
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UMD Theses and Dissertations

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