The class I PITP giotto is required for Drosophila cytokinesis.

Phosphatidylinositol transfer proteins (PITPs) are highly conserved polypeptides that bind phosphatidylinositol or phosphatidylcholine monomers, facilitating their transfer from one membrane compartment to another . Although PITPs have been implicated in a variety of cellular functions, including lipid-mediated signaling and membrane trafficking, the precise biological roles of most PITPs remain to be elucidated . Here we show for the first time that a class I PITP is involved in cytokinesis. We found that giotto (gio), a Drosophila gene that encodes a class I PITP, serves an essential function required for both mitotic and meiotic cytokinesis. Neuroblasts and spermatocytes from gio mutants both assemble regular actomyosin rings. However, these rings fail to constrict to completion, leading to cytokinesis failures. Moreover, gio mutations cause an abnormal accumulation of Golgi-derived vesicles at the equator of spermatocyte telophases, suggesting that Gio is implicated in membrane-vesicle fusion. Consistent with these results, we found that Gio is enriched at the cleavage furrow, the ER, and the spindle envelope. We propose that Gio mediates transfer of lipid monomers from the ER to the equatorial membrane, causing a specific local enrichment in phosphatidylinositol. This change in membrane composition would ultimately facilitate vesicle fusion, allowing membrane addition to the furrow and/or targeted delivery of proteins required for cytokinesis.

Genetic dissection of meiotic cytokinesis in Drosophila males.

We have used Drosophila male meiosis as a model system for genetic dissection of the cytokinesis mechanism. Drosophila mutants defective in meiotic cytokinesis can be easily identified by their multinucleate spermatids. Moreover, the large size of meiotic spindles allows characterization of mutant phenotypes with exquisite cytological resolution. We have screened a collection of 1955 homozygous mutant male sterile lines for those with multinucleate spermatids, and thereby identified mutations in 19 genes required for cytokinesis. These include 16 novel loci and three genes, diaphanous, four wheel drive, and pebble, already known to be involved in Drosophila cytokinesis. To define the primary defects leading to failure of cytokinesis, we analyzed meiotic divisions in male mutants for each of these 19 genes. Examination of preparations stained for tubulin, anillin, KLP3A, and F-actin revealed discrete defects in the components of the cytokinetic apparatus, suggesting that these genes act at four major points in a stepwise pathway for cytokinesis. Our results also indicated that the central spindle and the contractile ring are interdependent structures that interact throughout cytokinesis. Moreover, our genetic and cytological analyses provide further evidence for a cell type-specific control of Drosophila cytokinesis, suggesting that several genes required for meiotic cytokinesis in males are not required for mitotic cytokinesis.

The Drosophila Cog5 homologue is required for cytokinesis, cell elongation, and assembly of specialized Golgi architecture during spermatogenesis.

The multisubunit conserved oligomeric Golgi (COG) complex has been shown previously to be involved in Golgi function in yeast and mammalian tissue culture cells. Despite this broad conservation, several subunits, including Cog5, were not essential for growth and showed only mild effects on secretion when mutated in yeast, raising questions about what functions these COG complex subunits play in the life of the cell. Here, we show that function of the gene four way stop (fws), which encodes the Drosophila Cog5 homologue, is necessary for dramatic changes in cellular and subcellular morphology during spermatogenesis. Loss-of-function mutations in fws caused failure of cleavage furrow ingression in dividing spermatocytes and failure of cell elongation in differentiating spermatids and disrupted the formation and/or stability of the Golgi-based spermatid acroblast. Consistent with the lack of a growth defect in yeast lacking Cog5, animals lacking fws function were viable, although males were sterile. Fws protein localized to Golgi structures throughout spermatogenesis. We propose that Fws may directly or indirectly facilitate efficient vesicle traffic through the Golgi to support rapid and extensive increases in cell surface area during spermatocyte cytokinesis and polarized elongation of differentiating spermatids. Our study suggests that Drosophila spermatogenesis can be an effective sensitized genetic system to uncover in vivo functions for proteins involved in Golgi architecture and/or vesicle transport.