Worm Breeder's Gazette 13(3): 66 (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.

Suppressors of daf-11 identify new daf-d genes

Wendy Schackwitz, James H. Thomas

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

C. elegans forms a dauer larva when exposed to high levels of dauer pheromone. Several genes involved in this process have been identified. Mutations in most of these genes result in one of two phenotypes, dauer defective (Daf-d) or dauer constitutive (Daf-c). The genes have been placed in a regulatory pathway (see below) based on genetic interactions. daf-d genes are efficiently identified by looking for suppressors of daf-c genes. Although this type of screen had been performed, screens for suppressors of daf-11 had not been done, therefore there was the potential of identifying new daf-d genes by looking for suppressors of daf-11 .We have identified new daf-d genes by using EMS to generate suppressors of the daf-11 Daf-c phenotype. The screen identified 20 strong suppressor mutations (>90% suppression of the daf-11 Daf-c phenotype) that appear to identify new genes. These 20 suppressors were mapped to linkage groups and complementation tests within each group revealed that they represent l6 genes. Eight of the strongest suppressors (>97% suppression of the daf-11 Daf-c phenotype) were chosen for further analysis. These eight suppressors are being mapped in more detail. Summary of results to date: sa143 maps to LG I between dpy-5 and unc-13 ; sa151 maps to LG I between aex-6 and unc-54 ; sa169 maps to LG I close to egl-30 ; sa172 , sa173 ,and sa159 (allelic) map to LG 1 to the right or very close to unc-13 ; sa150 and sa158 (allelic) map to LG III between unc-93 and unc-32 ; sa171 maps to LG IV close to dpy-4 ; sa176 maps to LG V close to unc-34 .;and sa163 maps to LG X.

To test the phenotype of the suppressor mutations on their own, strains that no longer carry daf-11 ( m87 )have been constructed for sa143 , sa150 , sa151 , sa169 ,and sa171 .To test whether the suppressor mutations conveyed a Daf-d phenotype, the ability to form dauers on starved plates at 25 C was semi-quantitatively assayed. Most plates of the wild type formed thousands of dauers under our test conditions. Suppressors sa150 and sa169 formed many dauers on starved plates (l00- l000s )and thus do not confer a Daf-d phenotype. The sa143 , sa151 ,and sa171 strains formed very few dauers on starved plates (0-50 dauers). Thus these mutations confer an incompletely penetrant Daf-d phenotype. Interestingly daf-3 and daf-5 mutations also confer an incompletely penetrant Daf-d phenotype (0-50 dauers), whereas daf-12 is completely penetrant (0 dauers).

Limited epistasis analysis has been performed for sa143 with several daf-c mutations. The resulting daf-d; daf-c double mutants fall into three phenotypic classes: fully suppressed, weakly suppressed, and not suppressed. The Daf-c phenotype of daf-11 ( m87 ), daf-11 ( m84 ), daf-11 ( sa195 ),and daf-21 ( p673 )are all fully suppressed by sa143 (0-0.6% dauer formation). The Daf-c phenotypes of daf-7 ( e1372 ), daf-14 ( m77 ),and daf-4 ( e1364 )are weakly suppressed by sa143 (76-84% dauer formation), and daf- l9 ( m86 )is not suppressed by sa143 (100% dauer formation). This pattern of suppression can be explained by the parallel pathway model shown below. sa143 lies downstream of the group 1 daf-c genes, and thus fully suppresses mutations in them. sa143 only weakly suppresses the group 2 daf-c mutations because, in the double mutant, one branch of the pathway is in a dauer repressing state whereas the other branch is in a dauer stimulating state. When the two branches are summed together the result is a weak Daf-4 phenotype. Further characterization of the suppressors are underway.

Figure 1