Worm Breeder's Gazette 13(3): 65 (June 1, 1994)
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.
Dauer formation and dauer recovery are regulated by the same three external conditions: the concentration of dauer pheromone, temperature and the abundance of food. Previously all mutations that caused constitutive dauer formation (Daf-c) also inhibited dauer recovery. Together, these facts show that many genes are involved in regulating both dauer formation and dauer recovery. We recently described sa191 ,a semi-dominant, temperature-sensitive Daf-c mutation that identifies a new gene, daf-28 (Malone and Thomas, Genetics 136: 879-886). This mutation is unique because the dauers recover rapidly at all temperatures. We hypothesize that daf-28 ( sa191 )perturbs only dauer formation, and not recovery. Because sa191 is dominant, and because a deficiency of the locus does not cause a dominant Daf-c phenotype, we conclude that sa191 is a gain-of-function mutation.
As a first step in understanding the role of daf-28 in dauer formation, we have constructed double mutants containing daf-28 ( sa191 )and mutations in other genes that affect dauer formation. The mutations analyzed fall into two classes: mutations that prevent dauer formation (Daf-d) and other Daf-c mutations. Previous double mutant analysis suggests that these genes act in a complex, branched pathway (please see abstract by Schackwitz and Thomas for an abbreviated model). All but one of the daf-28 ;daf-d double mutants are dauer constitutive, including those containing daf-22 , daf-6 , daf-3 , daf-5 ,and mutations in several genes that affect the structure of amphid sensory cilia. These results suggest that daf-28 functions downstream of these Daf-d genes. In contrast, daf-12 completely suppresses the Daf-c phenotype of daf-28 ,suggesting that daf-12 acts downstream of daf-28 .
Like daf-28 ( sa191 ),all of the Daf-c mutations that we tested are temperature sensitive. The phenotypes of the daf-28 ;daf-c double mutants were examined at 15°, the permissive temperature at which all of the single mutants form dauers at low frequency (0 to 77 %). Under these conditions, all but one of the daf-28 ;daf-c double mutants formed 100% dauers, including those containing daf-11 , daf-21 , daf-1 , daf-4 , daf-7 , daf-8 ,and daf-14 .These synergistic interactions suggest that daf-28 functions in parallel with these genes rather than in series. In contrast, the daf-2 ; daf-28 double mutant forms few dauers at 15°. It is possible that daf-28 functions in a sequential pathway with daf-2 .However, two results distinguish daf-28 from daf-2 .First, a daf-16 mutation suppresses daf-2 but not daf-28 .Second, daf-2 ; daf-12 double mutants arrest in larval development, whereas the daf-28 ; daf-12 double mutant is viable and fertile. In summary, daf-28 ( sa191 )isunique in its interactions with other mutations that affect dauer formation, and it may define a new branch of the network of genes that impinge on daf-12 .
To further our understanding of the function of daf-28 ,we reverted daf-28 ( sa191 )with the aim of obtaining loss-of-function mutations and possible extragenic suppressors. Taking advantage of the fact that unc-31 ; daf-28 mutants recovery poorly from the dauer stage, we mutagenized these animals with EMS and screened for F2 progeny that did not form dauers. In a screen of 9,200 genomes, we identified 25 mutants, seven of which represent daf-12 alleles. Of the remaining 18 revertants, we have determined that at least seven do not contain daf-12 mutations. Since daf-12 mutations are the only known Daf-d mutations that suppress daf-28 ( sa191 ),we expect that these revertants represent either loss-of-function mutations in daf-28 or extragenic suppressors defining new genes.