Mechanism of activation of the Caenorhabditis elegans ras homologue let-60 by a novel, temperature-sensitive, gain-of-function mutation.
The Caenorhabditis elegans let-60 gene encodes a Ras protein that mediates induction of the hermaphrodite vulva. To better understand how mutations constitutively activate Ras and cause unregulated cell division, we have characterized ga89, a temperature-sensitive, gain-of-function mutation in let-60 ras. At 25 degrees, ga89 increases let-60 activity resulting in a multivulva phenotype. At 15 degrees, ga89 decreases let-60 activity resulting in a vulvaless phenotype in let-60(ga89)/Df animals. The ga89 mutation causes a leucine (L) to phenylalanine (F) substitution at amino acid 19, a residue conserved in all Ras proteins. We introduced the L19F change into human H-Ras protein and found that the in vitro GTPase activity of H-Ras became temperature-dependent. Genetic experiments suggest that LET-60 (L19F) interacts with GAP and GNEF, since mutations that decrease GAP and GNEF activity affect the multivulva phenotype of let-60(ga89) animals. These results suggest that the L19F mutation primarily affects the intrinsic rate of GTP hydrolysis by Ras, and that this effect may be sufficient to account for the activated-Ras phenotype caused by let-60(ga89). Our results suggest that a mutation in a human ras gene analogous to ga89 might contribute to oncogenic transformation.
Eisenmann
D M
DM
Department of Developmental Biology, Stanford University Medical Center, California 94305-5427, USA.
Kim
S K
SK
D000818:Animals; D017173:Caenorhabditis elegans / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D029742:Caenorhabditis elegans Proteins; D002454:Cell Differentiation / Q000235:genetics; D002455:Cell Division; D005260:Female; D020558:GTP Phosphohydrolases / Q000235:genetics / Q000378:metabolism; D020690:GTPase-Activating Proteins; D018507:Gene Expression Regulation, Developmental; D011905:Genes, ras; D020662:Guanine Nucleotide Exchange Factors; D015801:Helminth Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D009154:Mutation; D010641:Phenotype; D011506:Proteins / Q000378:metabolism; D015398:Signal Transduction; D013696:Temperature; D014844:Vulva / Q000254:growth & development; D020703:ras GTPase-Activating Proteins; D020663:ras Guanine Nucleotide Exchange Factors; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism
D029742:Caenorhabditis elegans Proteins; D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors; D015801:Helminth Proteins; D011506:Proteins; D020703:ras GTPase-Activating Proteins; D020663:ras Guanine Nucleotide Exchange Factors; C066646:let-60 protein, C elegans; D020558:GTP Phosphohydrolases; D018631:ras Proteins