Asparagine 26, glutamic acid 31, valine 45, and tyrosine 64 of Ras proteins are required for their oncogenicity.
Ras and Rap1 proteins are related GTP-dependent signal transducers which require Gly-12, the effector domain (residues 32-40), and Ala-59 for stimulation of their GTPase activities by GAP1 and GAP3, respectively. The replacement of Gly-12 by Val or Ala-59 by Thr potentiates the Ras oncogenicity and Rap1A antioncogenicity. However, the mutations in the effector domain, in particular the replacement of Thr-35 by Ala, abolish both Ras oncogenicity and Rap1A antioncogenicity, indicating that the effector domain is involved in interactions of these signal transducers with their targets as well as the GAPs. In this paper, we demonstrate that (i) replacement of Tyr-64 of the Ha-Ras protein or Phe-64 of the Rap1A protein by Glu or other non-hydrophobic amino acids reduces their intrinsic GTPase activities and abolishes their stimulation by GAP1 or GAP3, respectively, (ii) replacement of Tyr-64 by Gly and other non-hydrophobic amino acids results in complete loss of the oncogenicity of the v-Ha-Ras protein, indicating that the hydrophobic residue 64, in addition to the known effector domain, is essential for the Ras protein to interact with its target as well as GAP1. In addition we have found that Asn-26, Glu-31, and Val-45 of the v-Ha-Ras protein are required for its oncogenicity. Replacement of the Ras residues at either positions 26, 31, or 45 by the corresponding Rap1A residues abolishes the Ras oncogenicity.
Nur-E-Kamal
M S
MS
Melbourne Tumor Biology Branch, Ludwig Institute for Cancer Research, Royal Melbourne Hospital, Victoria, Australia.
Sizeland
A
A
D'Abaco
G
G
Maruta
H
H
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D002471:Cell Transformation, Neoplastic; D002678:Chimera; D003001:Cloning, Molecular / Q000379:methods; D004926:Escherichia coli / Q000235:genetics; D020558:GTP Phosphohydrolases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D011905:Genes, ras; D005982:Glutathione Transferase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015689:Oncogene Protein p21(ras) / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D016133:Polymerase Chain Reaction / Q000379:methods; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D015183:Restriction Mapping; D015398:Signal Transduction; D020668:rap GTP-Binding Proteins
D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; C501009:HRAS protein, human; D015689:Oncogene Protein p21(ras); D016283:Proto-Oncogene Proteins p21(ras); D020668:rap GTP-Binding Proteins