Antagonistic Smad transcription factors control the dauer/non-dauer switch inC. elegans

Abstract

SUMMARYThe C. elegans daf-8 gene encodes an R-Smad that is expressed in a subset of head neurons, the intestine, gonadal distal tip cells and the excretory cell. We found that DAF-8, which inhibits the DAF-3 Co-Smad, is associated with DAF-3 and the DAF-14 Smad in vivo and in vitro. Overexpression of daf-8 conferred a dauer-defective phenotype and suppressed constitutive dauer formation in daf-8 and daf-14 mutants. In contrast to mammalian systems described thus far, active DAF-3 drives a feedback regulatory loop that represses transcription of daf-7 (a TGF ligand) and daf-8 by directly binding to their regulatory regions. Hence, DAF-8 and DAF-3 are mutually antagonistic. The feedback repression may reinforce the developmental switch by allowing DAF-3 to freely activate dauer transcription in target tissues, unless sufficiently inhibited by DAF-8 and DAF-14. In the adult, DAF-8 downregulates lag-2 expression in the distal tip cells, thus promoting germ line meiosis. This function does not involve DAF-3, thereby avoiding the feedback loop that functions in the dauer switch.

Linking the environment, DAF-7/TGFβ signaling and LAG-2/DSL ligand expression in the germline stem cell niche

“…In apparent conflict with our results, and as noted by Dalfó et al, (2012), Park et al (2010) reported that daf-8/R-Smad negatively regulates lag-2 high-copy reporter expression in the DTC and germline proliferative zone size, independently of daf-3. Dalfó et al (2012) observed positive regulation (i.e.…”

“…Within the 3 kb region upstream of lag-2, we found three instances of the 5 bp DAF-3 binding motif (Fig. 3A) previously defined in the myo-2 promoter (Thatcher et al, 1999), and that are also present within a DAF-3-bound region in the promoters of daf-7 and daf-8, as determined by whole-animal ChIP (Park et al, 2010). If these sites mediate DAF-3 repressor activity, mutating them should render lag-2 expression insensitive to the loss of daf-7 or daf-1.…”

“…Furthermore, DAF-3 was not enriched in the lag-2-coding region, similar to our negative control, daf-14 ( Fig. S5A) (Park et al, 2010). Together, these results indicate that the lag-2 upstream regulatory region contains the lag-2p-DBS and lag-2p-Cons sequence components of the PD motif, both of which have enriched DAF-3 binding in the ChIP assay.…”

“…S5A). As positive controls, we verified DAF-3 enrichment at its characterized binding sites in the myo-2, daf-7 and daf-8 promoters (Park et al, 2010;Thatcher et al, 1999). Furthermore, DAF-3 was not enriched in the lag-2-coding region, similar to our negative control, daf-14 ( Fig.…”

“…Work by others has shown that DAF-3 can also bind to mdl-1, daf-7 and daf-8 promoters in non-dauer animals (Deplancke et al, 2006;Park et al, 2010), and Sims et al (2016) demonstrated that DAF-3 bound the osm-9 promoter in continuously developing adult animals, which was dependent upon functional ZFP-1/AF10 protein and endogenous RNAi pathways. Spacing may also be important for DAF-3 binding of DBS and Cons, as binding in the bacterial one-hybrid assay was maintained when the sites were individually scrambled (rather than deleted) but was lost when both were scrambled (Fig.…”