Novel LOTUS-domain proteins are organizational hubs that recruit C. elegans Vasa to germ granules

We describe MIP-1 and MIP-2, novel paralogous C. elegans germ granule components that interact with the intrinsically disordered MEG-3 protein. These proteins promote P granule condensation, form granules independently of MEG-3 in the postembryonic germ line, and balance each other in regulating P granule growth and localization. MIP-1 and MIP-2 each contain two LOTUS domains and intrinsically disordered regions and form homo- and heterodimers. They bind and anchor the Vasa homolog GLH-1 within P granules and are jointly required for coalescence of MEG-3, GLH-1, and PGL proteins. Animals lacking MIP-1 and MIP-2 show temperature-sensitive embryonic lethality, sterility, and mortal germ lines. Germline phenotypes include defects in stem cell self-renewal, meiotic progression, and gamete differentiation. We propose that these proteins serve as scaffolds and organizing centers for ribonucleoprotein networks within P granules that help recruit and balance essential RNA processing machinery to regulate key developmental transitions in the germ line

Feo, the Drosophila homolog of PRC1, is required for central spindle formation and cytokinesis

We performed a functional analysis of fascetto (feo), a Drosophila gene that encodes a protein homologous to the Ase1p/PRC1/MAP65 conserved family of microtubule-associated proteins (MAPs) [1-5]. These MAPs are enriched at the spindle midzone in yeast and mammals and at the fragmoplast in plants, and are essential for the organization and function of these microtubule arrays [1-5]. Here we show that the Feo protein is specifically enriched at the central spindle midzone, and its depletion by either mutation or RNAi results in aberrant central spindles. In Feo-depleted cells, late anaphases showed normal overlap of the antiparallel MTs at the cell equator, but telophases displayed thin MT bundles of uniform width instead of robust hourglass-shaped central spindles. These thin central spindles exhibited diffuse localizations of both the Pav and Asp proteins, suggesting that these spindles are comprised of improperly oriented MTs. Feo-depleted cells also displayed defects in the contractile apparatus that correlated with those in the central spindle: late anaphase cells formed regular contractile structures, but these structures did not constrict during telophase, leading to failures in cytokinesis. The central spindle phenotype of Feo-depleted cells suggests that Feo interacts with the antiparallel MTs of the central spindle midzone, maintaining their precise overlap during MT elongation and antiparallel sliding

The landscape of C. elegans 3'UTRs

Three-prime untranslated regions (3'UTRs) of metazoan mRNAs contain numerous regulatory elements, yet remain largely uncharacterized. Using polyA capture, 3'RACE, full-length cDNAs, and RNA-seq, we define ~26,000 distinct 3'UTRs in Caenorhabditis elegans for ~85% of the 18,328 experimentally supported protein coding genes and revise ~40% of gene models. Alternative 3'UTR isoforms are frequent, often differentially expressed during development. Average 3'UTR length decreases with animal age. Surprisingly, no polyadenylation signal (PAS) is detected for 13% of polyA sites, predominantly among shorter alternative isoforms. Trans-spliced (vs. non-trans-spliced) mRNAs possess longer 3'UTRs and frequently contain no PAS or variant PAS. We identify conserved 3'UTR motifs, isoform-specific predicted microRNA target sites, and polyadenylation of most histone genes. Our data reveal a rich complexity of 3'UTRs genome-wide and throughout development

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

A Balance of Capping Protein and Profilin Functions Is Required to Regulate Actin Polymerization in Drosophila Bristle

Profilin is a well-characterized protein known to be important for regulating actin filament assembly. Relatively few studies have addressed how profilin interacts with other actin-binding proteins in vivo to regulate assembly of complex actin structures. To investigate the function of profilin in the context of a differentiating cell, we have studied an instructive genetic interaction between mutations in profilin (chickadee) and capping protein (cpb). Capping protein is the principal protein in cells that caps actin filament barbed ends. When its function is reduced in the Drosophila bristle, F-actin levels increase and the actin cytoskeleton becomes disorganized, causing abnormal bristle morphology. chickadee mutations suppress the abnormal bristle phenotype and associated abnormalities of the actin cytoskeleton seen in cpb mutants. Furthermore, overexpression of profilin in the bristle mimics many features of the cpb loss-of-function phenotype. The interaction between cpb and chickadee suggests that profilin promotes actin assembly in the bristle and that a balance between capping protein and profilin activities is important for the proper regulation of F-actin levels. Furthermore, this balance of activities affects the association of actin structures with the membrane, suggesting a link between actin filament dynamics and localization of actin structures within the cell