Epigenetic information is frequently erased near the start of each new generation (1). In some cases, however, epigenetic information can be transmitted from parent to progeny (epigenetic inheritance) (2). A particularly striking example of epigenetic inheritance is dsRNA-mediated gene silencing (RNAi) in C. elegans, which can be inherited for more than five generations (3–8). To understand this process we conducted a genetic screen for animals defective for transmitting RNAi silencing signals to future generations. This screen identified the gene heritable RNAi defective (hrde)-1. hrde-1 encodes an Argonaute (Ago) that associates with small interfering (si)RNAs in germ cells of the progeny of animals exposed to dsRNA. In nuclei of these germ cells, HRDE-1 engages the Nrde nuclear RNAi pathway to direct H3K9me3 at RNAi targeted genomic loci and promote RNAi inheritance. Under normal growth conditions, HRDE-1 associates with endogenously expressed siRNAs, which direct nuclear gene silencing in germ cells. In hrde-1 or nuclear RNAi deficient animals, germline silencing is lost over generational time. Concurrently, these animals exhibit steadily worsening defects in gamete formation and function that ultimately lead to sterility. These results establish that the Ago HRDE-1 directs gene-silencing events in germ cell nuclei, which drive multi-generational RNAi inheritance and promote immortality of the germ cell lineage. We propose that C. elegans uses the RNAi inheritance machinery to transmit epigenetic information, accrued by past generations, into future generations to regulate important biological processes.

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The intestinal cells of Caenorhabditis elegans embryos contain prominent, birefringent gut granules that we show are lysosome-related organelles. Gut granules are labeled by lysosomal markers, and their formation is disrupted in embryos depleted of AP-3 subunits, VPS-16, and VPS-41. We define a class of gut granule loss (glo) mutants that are defective in gut granule biogenesis. We show that the glo-1 gene encodes a predicted Rab GTPase that localizes to lysosome-related gut granules in the intestine and that glo-4 encodes a possible GLO-1 guanine nucleotide exchange factor. These and other glo genes are homologous to genes implicated in the biogenesis of specialized, lysosome-related organelles such as melanosomes in mammals and pigment granules in Drosophila. The glo mutants thus provide a simple model system for the analysis of lysosome-related organelle biogenesis in animal cells. INTRODUCTIONLysosomes are ubiquitous membrane-bound organelles that function as major degradative sites within eukaryotic cells (Tappel, 1969). Lysosomes contain an assortment of aciddependent hydrolases that function in the breakdown of proteins, lipids, nucleic acids, and oligosaccharides. Lysosomes receive exogenous material through the endocytic pathway and are characterized as being the terminal compartment of the endocytic pathway. Lysosomes also receive material via the secretory pathway and directly from the cytoplasm (Kornfeld and Mellman, 1989;Mullins and Bonifacino, 2001;Luzio et al., 2003). Lysosomes function in diverse and important cellular processes including cell surface receptor turnover, destruction of pathogens, antigen processing, digestion, starvation responses, tissue remodeling, ion storage, autophagy, and plasma membrane repair.The yeast vacuole shares several characteristics with the lysosomes of higher animals. Genetic screens have led to the identification of Ͼ150 genes necessary for the transport and sorting of newly synthesized proteins to the yeast vacuole (Jones, 1977;Bankaitis et al., 1986;Rothman and Stevens, 1986;Bonangelino et al., 2002). These genes control two pathways of Golgi-to-vacuole transport, the carboxypeptidase Y (CPY) and alkaline phosphatase (ALP) sorting pathways (Burd et al., 1998;Conibear and Stevens, 1998;Mullins and Bonifacino, 2001). Proteins trafficked via the CPY pathway transit an endosomal prevacuolar compartment en route to the vacuole. The ALP pathway mediates transport to the vacuole independent of the prevacuolar compartment.Many of the genes involved in transport to the yeast vacuole have homologues in higher animals (Lemmon and Traub, 2000;Mullins and Bonifacino, 2001;Bonangelino et al., 2002). For example, the HOPS complex proteins (Vps11p, Vps16p, Vps18p, and Vps33p) regulate membrane fusion events necessary for lysosomal delivery within yeast (Rieder and Emr, 1997;Peterson and Emr, 2001), Drosophila melanogaster (Sevrioukov et al., 1999;Sriram et al., 2003), and mammalian (Poupon et al., 2003;Richardson et al., 2004) endosomal systems. Similarly, the proteins compos...

Central questions in regenerative biology include how stem cells are maintained and how they transition from self-renewal to differentiation. Germline stem cells (GSCs) in Caeno-rhabditis elegans provide a tractable in vivo model to address these questions. In this system, Notch signaling and PUF RNA binding proteins, FBF-1 and FBF-2 (collectively FBF), maintain a pool of GSCs in a naïve state. An open question has been how Notch signaling modulates FBF activity to promote stem cell self-renewal. Here we report that two Notch targets, SYGL-1 and LST-1, link niche signaling to FBF. We find that SYGL-1 and LST-1 proteins are cytoplasmic and normally restricted to the GSC pool region. Increasing the distribution of SYGL-1 expands the pool correspondingly, and vast overexpression of either SYGL-1 or LST-1 generates a germline tumor. Thus, SYGL-1 and LST-1 are each sufficient to drive “stemness” and their spatial restriction prevents tumor formation. Importantly, SYGL-1 and LST-1 can only drive tumor formation when FBF is present. Moreover, both proteins interact physically with FBF, and both are required to repress a signature FBF mRNA target. Together, our results support a model in which SYGL-1 and LST-1 form a repressive complex with FBF that is crucial for stem cell maintenance. We further propose that progression from a naïve stem cell state to a state primed for differentiation relies on loss of SYGL-1 and LST-1, which in turn relieves FBF target RNAs from repression. Broadly, our results provide new insights into the link between niche signaling and a downstream RNA regulatory network and how this circuitry governs the balance between self-renewal and differentiation.