Identification of PLC210, a Caenorhabditis elegans phospholipase C, as a putative effector of Ras.
Mammalian Ras proteins regulate multiple effectors including Raf, Ral guanine nucleotide dissociation stimulator (RalGDS), and phosphoinositide 3-kinase. In the nematode Caenorhabditis elegans, LIN-45 Raf has been identified by genetic analyses as an effector of LET-60 Ras. To search for other effectors in C. elegans, we performed a yeast two-hybrid screening for LET-60-binding proteins. The screening identified two cDNA clones encoding a phosphoinositide-specific phospholipase C (PI-PLC) with a predicted molecular mass of 210 kDa, designated PLC210. PLC210 possesses two additional functional domains unseen in any known PI-PLCs. One is the C-terminal Ras-associating domain bearing a structural homology with those of RalGDS and AF-6. This domain, which could be narrowed down to 100 amino acid residues, associated in vitro with human Ha-Ras in a GTP-dependent manner and competed with yeast adenylyl cyclase for binding Ha-Ras. The binding was abolished by specific mutations within the effector region of Ha-Ras. The other functional domain is the N-terminal CDC25-like domain, which possesses a structural homology to guanine nucleotide exchange proteins for Ras. These results strongly suggest that PLC210 belongs to a novel class of PI-PLC, which is a putative effector of Ras.
Shibatohge
M
M
Department of Physiology II, Kobe University School of Medicine, 7-5-1 Kusunoki-cho, Chuo-ku, Kobe 650, Japan.
Kariya
K i
Ki
Liao
Y
Y
Hu
C D
CD
Watari
Y
Y
Goshima
M
M
Shima
F
F
Kataoka
T
T
D000595:Amino Acid Sequence; D000818:Animals; D017173:Caenorhabditis elegans / Q000201:enzymology*; D029742:Caenorhabditis elegans Proteins; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006801:Humans; D008969:Molecular Sequence Data; D016296:Mutagenesis; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D011485:Protein Binding / Q000187:drug effects; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D010738:Type C Phospholipases / Q000235:genetics / Q000378:metabolism*; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism*
D029742:Caenorhabditis elegans Proteins; D018076:DNA, Complementary; C066646:let-60 protein, C elegans; D006160:Guanosine Triphosphate; D010738:Type C Phospholipases; D054801:Phosphoinositide Phospholipase C; D018631:ras Proteins; D043265:Phosphatidylinositol Diacylglycerol-Lyase