The P granule component PGL-1 promotes the localization and silencing activity of the PUF protein FBF-2 in germline stem cells

Abstract

SUMMARYIn the C. elegans germline, maintenance of undifferentiated stem cells depends on the PUF family RNA-binding proteins FBF-1 and FBF-2. FBF-1 and FBF-2 are 89% identical and are required redundantly to silence the expression of mRNAs that promote meiosis. Here we show that, despite their extensive sequence similarity, FBF-1 and FBF-2 have different effects on target mRNAs. FBF-1 promotes the degradation and/or transport of meiotic mRNAs out of the stem cell region, whereas FBF-2 prevents translation. FBF-2 activity depends on the P granule component PGL-1. PGL-1 is required to localize FBF-2 to perinuclear P granules and for efficient binding of FBF-2 to its mRNA targets. We conclude that multiple regulatory mechanisms converge on meiotic RNAs to ensure silencing in germline stem cells. Our findings also support the view that P granules facilitate mRNA silencing by providing an environment in which translational repressors can encounter their mRNA targets immediately upon exit from the nucleus.

The PUF binding landscape in metazoan germ cells

“…Rescued animals were phenotypically wild-type and FLAG-tagged FBF proteins were enriched in the distal germline, which includes the GSCs (Supplemental Fig. S1), consistent with their endogenous expression patterns (Lamont et al 2004;Voronina et al 2012). Also, their expression was essentially limited to the germline (Supplemental Fig.…”

“…These paralogs possess 91% identical amino acid sequences, essentially the same binding properties, and are functionally redundant for GSC maintenance (Zhang et al 1997;Crittenden et al 2002;Bernstein et al 2005). Yet FBF-1 and FBF-2 have subtle differences in phenotype and subcellular localization, suggesting they may have distinct targets (Crittenden et al 2002;Lamont et al 2004;Voronina et al 2012). An early RIP-chip study, relying on overexpressed GFP-tagged FBF-1, identified 1350 putative targets (Kershner and Kimble 2010).…”

Dynein light chain DLC-1 promotes localization and function of the PUF protein FBF-2 in germline progenitor cells

“…In the distal mitotic cells, FBF-2 is localized to perinuclear foci overlapping with P granules, and this localization is required for FBF-2 activity (Voronina et al, 2012). Because DLC-1 is required for FBF-2 regulatory activity, we tested whether DLC-1 played a role in FBF-2 localization to P granules.…”

“…Despite high similarity between FBF-1 and FBF-2 (89% identity at the amino acid level) and apparent redundancy in their control of the switch from spermatogenesis to oogenesis, single fbf-1 and fbf-2 mutants have distinct phenotypes, suggesting that these genes play unique roles (Lamont et al, 2004). By examining the effects of single mutants on target mRNAs, it has been shown that FBF-1 inhibits accumulation of the target mRNAs in the mitotic zone and FBF-2 primarily represses mRNA translation (Voronina et al, 2012). In addition, only FBF-2 localizes to the germ cell-specific subtype of RNA granules called P granules, and this localization is required for the function of FBF-2 (Voronina et al, 2012).…”

“…By examining the effects of single mutants on target mRNAs, it has been shown that FBF-1 inhibits accumulation of the target mRNAs in the mitotic zone and FBF-2 primarily represses mRNA translation (Voronina et al, 2012). In addition, only FBF-2 localizes to the germ cell-specific subtype of RNA granules called P granules, and this localization is required for the function of FBF-2 (Voronina et al, 2012). By contrast, FBF-1 does not localize to P granules and functions independently of these structures.…”