Cytological and biochemical analyses of Wolbachia pipienitis, causative reagent of cytoplasmic incompatibility in Drosophila simulans
Kose, Hiroyuki
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https://hdl.handle.net/2142/18963
Description
Title
Cytological and biochemical analyses of Wolbachia pipienitis, causative reagent of cytoplasmic incompatibility in Drosophila simulans
Author(s)
Kose, Hiroyuki
Issue Date
1995
Doctoral Committee Chair(s)
Karr, Timothy L.
Department of Study
Biochemistry
Discipline
Biochemistry
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Biology, Cell
Chemistry, Biochemistry
Language
eng
Abstract
Cytoplasmic incompatibility (CI) refers to the male-specific sterility widely observed among various insect species. CI has been correlated to an intracellular microorganism called Wolbachia pipientis. The expression of CI results when males infected with the symbionts are crossed with uninfected females; whereas reciprocal crosses between infected females and uninfected males are compatible, as well as crosses between flies of same infection types. CI was discovered in Drosophila simulans(DSR) in 1989, however virtually nothing is known regarding molecular mechanisms of CI.
In order to investigate the host-symbiont relationships, the symbionts were isolated in pure form from fly embryos and subsequently used to generate monoclonal antibodies specific to Wolbachia. Using one of those Wolbachia antibodies, the behavior of the symbiont during early embryogenesis was described. Indirect immunofluorescence staining, combined with confocal microscopy, revealed that during mitosis Wolbachia are localized near spindle poles and centrosomes. Direct interactions between the bacteria and centrosome-organized microtubules were implicated.
Furthermore, counting the number of Wolbachia revealed that there was no indication of bacterial cell division during preblastoderm period, which suggests that the localization as well as the growth of Wolbachia is under the tight regulation of host system.
"Six monoclonal anti-Wolbachia antibodies were classified based on nuclei staining patterns. Class I Mabs stain nuclei in both DSR (infected) & DSRT (uninfected), class II in DSRT, but not in DSR, and class III in neither DSR nor DSRT. It is surprising that immunoblot analysis showed that all of these anti-Wolbachia antibodies revealed identical bands in infected and uninfected extracts, suggesting that Wolbachia bind to a number of host proteins. One of the antigens by these antibodies was identified as histone H1, and N-terminal amino acid sequence was determined for one other Wol-antigen. The sequence showed mild homology with kinesin-like protein (KLP) of Drosophila melonogaster. Based upon these observations, I propose a ""sink"" model, which may explain the molecular mechanisms of CI."
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