Weiqing Li

In Caenorhabditis elegans, the decision to enter a developmentally arrested dauer larval stage is triggered by a combination of signals from sensory neurons in response to environmental cues, which include a dauer pheromone. These sensory inputs are coupled to the parallel DAF-2/insulin receptor-like and DAF-7/TGFbeta-like signaling pathways. Although sensory inputs have been shown to physiologically regulate DAF-7/TGFbeta expression, no such regulation of insulin-like ligands in the DAF-2 pathway has been reported. We show here that daf-28 encodes an insulin-like protein, which when mutated causes dauer arrest and down-regulation of DAF-2/IR signaling. A daf-28GFP fusion gene is expressed in ASI and ASJ, two sensory neurons that regulate dauer arrest. daf-28GFP expression in ASI and ASJ is down-regulated under dauer-inducing conditions and in mutants of DAF-11/guanylyl cyclase, a predicted component of the dauer-pheromone-sensing pathway. Thus, daf-28 expression in sensory neurons is regulated by the environmental cues that normally trigger dauer arrest. Among the 38 C. elegans insulin genes, daf-28 is so far the only insulin mutant to affect dauer arrest. daf-28 was revealed from this functional redundancy by a dominant-negative allele that disrupts a probable proteolytic processing site required for insulin maturation. This DAF-28 mutant is likely to be poisonous to wild-type DAF-28 and other insulins.

Perrault syndrome is a genetically heterogeneous recessive disorder characterized by ovarian dysgenesis and sensorineural hearing loss. In a nonconsanguineous family with five affected siblings, linkage analysis and genomic sequencing revealed the genetic basis of Perrault syndrome to be compound heterozygosity for mutations in the mitochondrial histidyl tRNA synthetase HARS2 at two highly conserved amino acids, L200V and V368L. The nucleotide substitution creating HARS2 p.L200V also created an alternate splice leading to deletion of 12 codons from the HARS2 message. Affected family members thus carried three mutant HARS2 transcripts. Aminoacylation activity of HARS2 p.V368L and HARS2 p.L200V was reduced and the deletion mutant was not stably expressed in mammalian mitochondria. In yeast, lethality of deletion of the single essential histydyl tRNA synthetase HTS1 was fully rescued by wild-type HTS1 and by HTS1 p.L198V (orthologous to HARS2 p.L200V), partially rescued by HTS1 p.V381L (orthologous to HARS2 p.V368L), and not rescued by the deletion mutant. In Caenorhabditis elegans, reduced expression by RNAi of the single essential histydyl tRNA synthetase hars-1 severely compromised fertility. Together, these data suggest that Perrault syndrome in this family was caused by reduction of HARS2 activity. These results implicate aberrations of mitochondrial translation in mammalian gonadal dysgenesis. More generally, the relationship between HARS2 and Perrault syndrome illustrates how causality may be demonstrated for extremely rare inherited mutations in essential, highly conserved genes.

To sense its population density and to trigger entry into the stress-resistant dauer larval stage, Caenorhabditis elegans uses the dauer pheromone, which consists of ascaroside derivatives with short, fatty acid-like side chains. Although the dauer pheromone has been studied for 25 years, its biosynthesis is completely uncharacterized. The daf-22 mutant is the only known mutant defective in dauer pheromone production. Here, we show that daf-22 encodes a homolog of human sterol carrier protein SCPx, which catalyzes the final step in peroxisomal fatty acid beta-oxidation. We also show that dhs-28, which encodes a homolog of the human d-bifunctional protein that acts just upstream of SCPx, is also required for pheromone production. Long-term daf-22 and dhs-28 cultures develop dauer-inducing activity by accumulating less active, long-chain fatty acid ascaroside derivatives. Thus, daf-22 and dhs-28 are required for the biosynthesis of the short-chain fatty acid-derived side chains of the dauer pheromone and link dauer pheromone production to metabolic state.