JCB: Report

In many organisms, germ cells have electron-dense cytoplasmic organelles, generally referred to as “germ granules,” which are believed to play roles in germ cell specification and differentiation (Eddy, 1975; Saffman and Lasko, 1999). They are large RNA-enriched nonmembranous organelles, and historically called by diverse names such as P granules in Caenorhabditis elegans, polar granules in Drosophila, and germinal granules in Xenopus (Eddy, 1975). In animals in which the germline is preformed (including the above three organisms), germ granules are maternally inherited by the fertilized egg and then specifically segregated to the germ lineage during early embryogenesis. Germ granules contain specific mRNAs and proteins, some of which are conserved among species (e.g., Drosophila Vasa [Hay et al., 1988a,b; Lasko and Ashburner, 1988] and its homologues [Raz, 2000]), but many appear to be species specific. Despite the evolutionary divergence of their molecular composition, the majority of the protein components of germ granules are implicated in various aspects of RNA metabolism, which has led to the prediction that the common biochemical function of germ granules is to regulate the translation efficiency and/or stability of mRNAs in the germline (Seydoux and Braun, 2006; Strome and Lehmann, 2007). In Caenorhabditis elegans, germ granules are called P granules, and some of the specific mRNAs and 40 protein components (mostly RNA-binding proteins) of P granules have been identified (Strome, 2005). Among them, GLH-1–4 (Drosophila Vasa homologues; with DEAD-box helicase motifs; Gruidl et al., 1996; Kuznicki et al., 2000), PGL-1 and PGL-3 (C. elegans–specific germ granule components; each with an RGG box [multiple Arg-Gly-Gly repeats]; Kawasaki et al., 1998, 2004), and DEPS-1 (Spike et al., 2008) are the only known components that exclusively localize to P granules at all developmental stages; other components localize to P granules transiently, and many of them are present in somatic lineages as well (Strome, 2005). Although many P-granule components have been identified, how the granules are assembled (and disassembled in somatic cells) is not well understood. Here we established an assay to analyze the granule-forming ability of P-granule components using cultured mammalian Germ granules are germ lineage–specific ribo­ nucleoprotein (RNP) complexes, but how they are assembled and specifically segregated to germ lineage cells remains unclear. Here, we show that the PGL proteins PGL­1 and PGL­3 serve as the scaffold for germ granule formation in Caenorhabditis elegans. Using cul­ tured mammalian cells, we found that PGL proteins have the ability to self­associate and recruit RNPs. Depletion of PGL proteins from early C. elegans embryos caused dis­ persal of other germ granule components in the cytoplasm, suggesting that PGL proteins are essential for the architec­ ture of germ granules. Using a structure–function analysis in vivo, we found that two functional domains of PGL pro­ teins contribute to germ granule assembly: an RGG box for recruiting RNA and RNA­binding proteins and a self­ association domain for formation of globular granules. We propose that self­association of scaffold proteins that can bind to RNPs is a general mechanism by which large RNP granules are formed. PGL proteins self associate and bind RNPs to mediate germ granule assembly in C. elegans