Worm Breeder's Gazette 16(5): 41 (February 1, 2001)

These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.

All available mutations in flp-1 also delete daf-10 sequences, confounding phenotype interpretation

Michael Ailion, James H. Thomas

Department of Genetics, University of Washington, Seattle, WA 98195

Based on the details explained below, several of the phenotypes previously attributed to flp-1 mutants are more likely to be caused by loss of daf-10. These include defects in osmotic avoidance (Osm), thermal avoidance (Tav), nose-touch response (Not), wandering behavior, regulation of dauer formation, and possibly regulation of egg-laying behavior in response to food signals. Defects in these behaviors are characteristic of cilium-structure mutants, and could be easily accounted for by loss of daf-10.

In surveying a variety of behavioral mutants for a dauer phenotype at 27, we found that both flp-1 deletion mutants, yn2 and yn4, have a moderate Daf-c phenotype at 27 and poor dauer recovery. Since many 27 Daf-c mutants are hypersensitive to dauer pheromone, we tested flp-1(yn2) for response to pheromone at 25. Surprisingly, it formed no dauers in response to pheromone. Mutations in the dyf genes similarly lead to a Daf-d phenotype at 25 and a Daf-c phenotype at 27. These mutants have defects in the structure of the ciliated endings of sensory neurons, a defect which can be assayed by the failure of these neurons to fill with fluorescent dyes. Thus, we performed FITC dye-filling assays on flp-1(yn2) and flp-1(yn4) and found that both mutants failed to exhibit dye-filling of any of the amphid or phasmid neurons.

flp-1 is not expressed in any of the amphid sensory neurons. Thus, we suspected that the flp-1 Dyf phenotype was due to a background mutation present in both flp-1 strains. The two alleles are not independent since they were isolated as deletions from the same parent strain which carries a Tc1 insertion just upstream of flp-1. We mapped the Dyf phenotype and found that it mapped to the right of unc-5 and was not separable from flp-1 (based on flp-1 movement phenotypes). The only candidate dyf mutant in this region is daf-10. We found that flp-1(yn2) failed to complement daf-10(e1387) for the Dyf phenotype, indicating that the flp-1 mutant strain also carries a mutation in daf-10. The flp-1 strains exhibit a more severe dye-filling defect than daf-10(e1387), indicating that they are stronger alleles.

daf-10 is encoded by F23B2.4 (Steve Stone and Jocelyn Shaw, personal communication), the gene immediately upstream of flp-1. Further inspection of the flp-1 deletions indicates that yn2 and yn4 both delete the daf-10 promoter and from two to four exons of the predicted daf-10 coding sequence. The original Tc1 used to isolate the flp-1 deletions appears to reside in the daf-10 gene, and both flp-1 deletion alleles are likely null for both genes.

Although several of the sensory phenotypes of flp-1 mutants can be explained by loss of daf-10, it is likely that several other phenotypes previously ascribed to flp-1 are bona fide flp-1 phenotypes. Specifically, flp-1 mutants have defects in movement and regulation of the active state of egg-laying that are not easily explained by loss of daf-10. Furthermore, both of these phenotypes were rescued by flp-1(+) transgenes.