DAF-12 is a nuclear hormone receptor that is a key component of the dauer formation decision in C. elegans . The current model for its action is as follows: - under reproductive growth conditions, a cholesterol-derived hormone is produced by DAF-9 that inhibits the dauer-promoting activity of DAF-12. However, under harsh environmental conditions, or in a
daf-9 mutant background, this hormone is not produced and the lack of hormone causes DAF-12 to promote dauer formation. We have been characterising the components of the dauer formation decision in the
ncr-2;
ncr-1 double mutant background.
ncr-1 and
ncr-2 are the C. elegans homologues of the human NPC1 gene, the major locus responsible for Niemann-Pick Type C disease. Like NPC1,
ncr-1&2 are likely to be involved in intracellular cholesterol trafficking.
ncr-2;
ncr-1 double mutants form dauers constitutively (Daf-c), and we propose that this is due to a lack of available cholesterol substrate for the production of the reproductive hormone required to inhibit the dauer-promoting activity of DAF-12. In support of this hypothesis, we have found that the Daf-c phenotype of the ncr mutants is sensitive to the cholesterol concentration of the media, and also that the ncr genes function upstream of
daf-9 in the dauer formation pathway (Li et al. 2004 submitted). We found through epistasis analysis and screening for suppressors of the ncr Daf-c phenotype that loss-of-function mutations in
daf-12 and the gene encoding its co-regulator DIN-1 (Andreas Ludwig, personal communication) suppress the Daf-c phenotype of ncr mutants. Whilst loss-of-function mutations in
daf-12 generally suppress the ncr phenotype in a recessive fashion, loss-of-function alleles of
din-1 act dominantly. We have shown this
din-1 dominance is due to haploinsufficiency in the ncr background. This haploinsufficiency of
din-1 in the ncr background is in contrast to the recessive suppression by
din-1 of other Daf-c mutants including
daf-9 . We propose a model whereby the dauer decision is the result of the relative levels of dauer-promoting complexes versus reproductive-growth complexes. The reproductive-growth complex consists of DAF-12 bound by the cholesterol-derived reproductive hormone (holoreceptor). In our model, the binding of this hormone prevents association of the co-regulator DIN-1 with DAF-12. In the absence of reproductive hormone, DIN-1 is able to associate with DAF-12 aporeceptor, forming the dauer-promoting complex. The relative abundance of these two types of complex therefore results from the relative levels of hormone and DIN-1. This explains why
din-1 is haploinsufficient in the ncr background, (where hormone levels are low but not zero), compared to its recessive nature in the
daf-9 mutant where hormone is completely absent Interestingly, a new
daf-12 allele,
sa1413 , which is deleted for most of the ligand binding domain of DAF-12 (including the DIN-1 interaction domain - Andreas Ludwig, personal communication) acts as a dominant negative allele. This provides evidence that DAF-12 functions as a dimer in the dauer-promoting complex and may prove to be a useful tool in future investigations of DAF-12 action.