Molecular genetic analysis of the
sur-2 gene defined by suppressors of a gain of function
let-60 ras allele Natasha Singh, John Yochem and Min Han Dept of MCD Biology, University of Colorado, Boulder, CO 80309 The
sur-2 gene is the most mutable locus identified in a screen for suppressors of a
let-60 ras gain of function allele(nlO46). The
sur-2 gene is currently defined by eight mutations that map 1 map unit away from
unc-54 on IR. Each of the eight alleles completely suppresses the Multivulva phenotype of
let-60(nlO46). Seven of the eight alleles display a completely penetrant egg laying defect (Egl) while one allele is partially Egl. The Egl defect can be explained by an abberant vulval lineage. Although the vulval lineages vary, the most common defect is the induction of P6.p to form a primary lineage while P5.p and P7.p remain syncicial. In addition to the vulval defect,
sur-2 mutants display other pleiotropic defects including a partial larval lethality, a partial dauer constitutive phenotype, gonad abnormalities and a male mating defect, suggesting that this gene is likely to play a key role in a number of important developmental decisions. The
sur-2(
ku9) allele is likely hypomorphic because vulval induction is reduced when
ku9 is placed over a deficiency. The
sur-2(
ku9) allele is also able to suppress the Muv phenotype of lin 15 (
n765) suggesting that
sur-2 acts downstream of, or parallel to,
let-60 and
lin-15. The interaction between
sur-2(
ku9) and
lin-1 is more difficult to interpret: the double mutant is extremely sick and most worms are sterile; however there is a partial suppression of the
lin-1 Muv phenotype suggesting that
sur-2, like
lin-1 acts very downstream in the pathway. The
sur-2 gene was cloned by deficiency mapping and DNA- mediated transformation. Rescue of the
ku9 egg laying defect requires nearly 14kb of genomic DNA. Developmental Northern blot analysis showed that the transcript is enriched during early embryonic and Ll stages of development. cDNA libraries constructed from these stages (Ahringer and Kimble) were screened and corresponding cDNA clones were obtained. DNA sequencing of the 14kb of genomic DNA and partial sequencing of the cDNAs indicate that
sur-2 encodes a novel protein; thus our genetic screen has enabled us to identify a new member of the ras signalling pathway that has eluded biochemical investigation. Further genetic and molecular analysis of
sur-2 is in progress.