FBF-1 and FBF-2 (collectively known as FBF) are two highly similar PUF domain RNA-binding proteins required for stem cell maintenance in the C. elegans germline1. To maintain germline stem cells, FBF inhibit the expression of positive regulators of meiosis, including the two RNA binding proteins GLD-1 and GLD-32. We have obtained evidence that FBF also inhibits the expression of HIM-3, a component of meiotic chromosome axes (homolog of S. cerevisiae Hop1p3). We found that the
him-3 promoter is active in germline progenitors, even in immature gonads that have not yet initiated meiosis. Proper regulation of HIM-3 depends on its 3''UTR: a GFP:H2B:
him-3 3''UTR fusion driven by a pan-germline promoter is repressed in progenitors and activated only in cells that have initiated the meiotic program. This regulation depends on FBF: depletion of FBF-1/2 by RNAi or by mutation activates the GFP:H2B:
him-3 3''UTR reporter in progenitors. Endogenous HIM-3 protein is also mis-expressed in progenitors in
fbf-1/2 mutants. FBF-1 and FBF-2 bind in vitro to a sequence motif termed FBF-binding element (FBE)4. There are two putative FBEs in the
him-3 3'' UTR and mutations in one of these activates the GFP:H2B:
him-3 3''UTR reporter in progenitors, suggesting that FBF may regulate
him-3 directly. To test whether FBF might regulate the expression of other meiotic proteins, we created 3''UTR fusions for 14 other genes coding for structural components of meiotic chromosomes. We found that the 3'' UTRs of
syp-2,
syp-3 and
htp-1 are down-regulated in progenitors in a pattern similar to
him-3. Depletion of FBF and mutation in a putative FBE both activate the
syp-3 3''UTR reporter in progenitors. Similar experiments are in progress for
syp-2 and
htp-1. To determine whether FBF-1/2 affect RNA levels, we compared the level of endogenous
him-3,
syp-2, and
syp-3 mRNAs in L2 larvae depleted/not depleted of
fbf-1/2 by RNAi. We found that depletion of FBF causes a 2-3 fold increase in mRNA levels compared to controls. We are currently testing if these mRNAs exist in a complex with FBF in vivo. Our data suggest that FBF maintain progenitors by directly silencing mRNAs coding for structural components of meiotic chromosomes. 1. Crittenden, et al., Nature 417 (6889), 660 (2002). 2. Kimble and Crittenden, WormBook, 1 (2005); Reinke, WormBook, 1 (2006). 3. Zetka, et al., Genes Dev. 13 (17), 2258 (1999). 4. Bernstein, et al., RNA 11 (4), 447 (2005).