Worm Breeder's Gazette 13(1): 79 (October 1, 1993)

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.

Screening For New Dauer Related Mutations

Ian M. Caldicott, Donald L. Riddle

Division of Biological Sciences, University of Missouri, Columbia, MO 65211

The numerous screens performed to isolate dauer constitutive mutants have identified 15 genes, mutations in two of which ( daf-9 and daf-15 )result in a dauer-like phenotype. These screens have always involved SDS selection or screening at 25°C (Riddle et al. Nature, 290:668-671, 1981), even when recovery at 15°C was not required (Riddle, unpublished; Malone and Thomas WBG V12 #4p.62). Almost all dauer-constitutive mutants are temperature-sensitive (Golden and Riddle, PNAS 81:819-823, 1984). To minimize the likelihood of isolating alleles of previously defined genes we performed a visual F2 screen for dauer and dauer-like larvae at 15°C. Three of the previously defined dauer constitutive genes ( daf-2 III, daf-15 IV and daf-9 X) were known to be targets for this screen because nonconditional alleles had been previously isolated. To more quickly identify mutations in these genes, the screen was initially performed in a dpy-1 III; unc-24 IV background.

As reported at the C. elegans meeting, we identified 38 mutations including alleles of daf-2 , daf-15 ,severe but conditional alleles of daf-4 and daf-11 ,and approximately 10 new genes. None of the mutations in new genes resulted in nonconditional formation of true dauers, indicating that the target size for this class of mutant is quite small. Since this screen provided a large number of new genes, all of which have a dauer-like phenotype when mutated, we repeated the screen. We used other genetic backgrounds to compensate for the possibility that particular markers might introduce biases into the screen. To date we have screened 9,000 genomes in the dpy-1 unc-24 background, 2,000 in an unc-36 dpy-20 background, and 5,500 genomes in a wild-type background. We have identified a total of 81 mutants. Five of the mutants isolated since the first screen have a true dauer constitutive phenotype (complementation testing in progress) and the rest have a non-conditional, constitutive dauer-like phenotype. The dauer-like larval arrest mutants vary considerably in size and "dauerness".

We are conducting molting analyses and pheromone sensitivity assays to divide the dauer-like mutations into two groups. We are assigning daf gene names to those that are unable to form dauers in pheromone, yet execute only 2 molts, following the precedent of daf-9 and daf-15 .These mutations are clearly affected in their ability to form dauers. The other mutations will be assigned let names. Of the ten new genes identified so far one is on chromosome II, 1 is on chromosome V, 2 are on chromosome III, and 5 are on chromosome IV. Initial analysis is centering on the cluster of genes on chromosome IV. These mutations define a set of genes required for non-dauer development, at least in some tissues or stages. Loss of gene function results in non-conditional dauer-like morphogenesis that prevents development to the reproductive adult.