TAGC poster 2020

Ramesh Krishnan, Anna Bakrhat, Andreas Bausch, Uri Abdu

Research output: Contribution to conferencePoster


Javelin, a novel protein is essential in Drosophila bristle actin bundles formation 1 2 In several specialized cell types, such as intestinal microvilli, stereocilia of the inner ear and Drosophila bristles, the structural integrity requires a specific actin filament organization brought about by a set of actin bundling proteins. In Drosophila bristle development, two major actin bundling proteins-Forked and Fascin are involved in forming actin networks but only little is known about the utilization of these proteins in a time-and space-dependent manner. In our present study, using RNA sequence analysis a new gene, Javelin (Jv), was identified that directly affect actin bundles formation. Javelin mutant bristles do not taper like bristles in wild-type flies, instead they present a small enlargement before the tip. In-vitro and in-vivo studies using Javelin in the presence of Forked and Fascin showed that jv plays a role in actin bundling in Drosophila.. 3 5 6 Purification of Javelin (Jv) and Co-immunoprecipitation assay In-vivo actin bundling-Jv interacts with Forked and Fascin when expressed ectopically in Drosophila oocyte 4 Jv full length, Javelin N-terminus and C-terminus exhibit unique localization pattern in Drosophila oocyte When expressed with Forked, Jv N-terminus inhibits actin bundling whereas Jv C-terminus favours actin bundling Confocal projections showing the localization pattern of Javelin full length (A), N-terminus of javelin (B) and C-terminus of Javelin (C) when driven with a maternal Gal4-3. Confocal projections showing the effect of Javelin full length (A-C), Javelin N-terminus (D-E) and Javelin C-terminus (G-H) on the actin bundles when expressed with mCherry Forked. Schematic representation of the domain structures of Javelin full length, Javelin N-terminus and Javelin C-terminus respectively. Jv consists of only one functional domain-CCD, which has no actin bundling activity. Inspite of this, Jv has a role in actin network thus making it an interesting candidate. Javelin was cloned into two parts-N terminus and C-terminus. Both contained the Coiled Coil Domain (CCD) and was fused with EGFP Confocal projections showing the ectopic expression of GFP Forked, Fascin and Javelin and their different combinations in Drosophila oocyte and the resulting actin networks. Jv 170 kDa Gel image showing the purified Javelin (MW 170 kDa) from SF-9 cells jv 50μm WT 50μm jv WT WT Javelin (jv) mutants show aberrant bristle morphology Bristle tip becomes swollen, resembling a spear Irregular groove pattern with unparallel and shallower ridges Western blot showing Co-IP of Flag Javelin and GFP Forked by anti-Flag and anti-GFP antibody. Co-immunoprecipitation (Co-IP) shows no interaction between Flag Javelin and GFP Forked in Drosophila ovary lysate. (A) represents the purified flag-javelin protein, (B) represents the GFP Forked ovary lysate and (C) represents the GFP beads bound with both Flag Javelin and GFP Forked.
Original languageEnglish
StatePublished - 7 May 2020


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