Molecular cloning and characterization of an adaptor protein Shc isoform from Xenopus laevis oocytes.
In order to gain further insight into IGF-1 receptor signaling in Xenopus laevis oocytes and embryos, we have undertaken the characterization of the adapter protein Shc and studied its implication in oocyte maturation induced after IGF-1 receptor activation, especially since expression of this molecule has been indirectly evidenced in Xenopus oocytes, eggs and embryos. We report herein the cloning from Xenopus postvitellogenic oocytes of a complementary DNA encoding a protein of 470 amino acids which shows the higher identity with the mammalian adaptor protein p52(ShcA). Western blot analysis using homologous antibodies evidenced a 60-kDa protein, p60(Xl)(Shc), that is predominantly expressed in oocytes and in early embryos. We also demonstrate that, like p60(Xl)(Shc), Grb2 and the guanine nucleotide exchange factor Sos are expressed in oocytes throughout vitellogenesis and in early embryos and that overexpression of a dominant-negative form of Grb2 specifically inhibits insulin-induced resumption of meiosis. We finally show that Grb2 binds to p60(Shc) in oocytes specifically upon insulin treatment. Altogether, these results suggest that Shc and Grb2-Sos are implicated in ras-dependent Xenopus oocyte maturation induced by insulin/IGF-1; they also indicate that inability of insulin/IGF-1 to activate the Ras-MAPK cascade in vitellogenic oocytes does not result from an insufficient expression level of Shc, Grb2 and Sos.
Chesnel
Franck
F
UMR 6061-CNRS << Génétique et développement >>, IFR 97 - Université de Rennes 1 - 35042 Rennes cedex, France. Franck.chesnel@univ-rennes1.fr
Heligon
Christophe
C
Richard-Parpaillon
Laurent
L
Boujard
Daniel
D
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport / Q000235:genetics* / Q000502:physiology; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D004625:Embryo, Nonmammalian / Q000378:metabolism; D047108:Embryonic Development; D005260:Female; D051380:GRB2 Adaptor Protein; D040461:Gene Components; D018507:Gene Expression Regulation, Developmental; D007328:Insulin / Q000494:pharmacology; D008099:Liver / Q000378:metabolism; D008969:Molecular Sequence Data; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism*; D020033:Protein Isoforms / Q000235:genetics / Q000502:physiology; D011506:Proteins / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D055765:Shc Signaling Adaptor Proteins; D020664:Son of Sevenless Proteins / Q000378:metabolism; D014818:Vitellogenesis / Q000502:physiology; D029867:Xenopus Proteins / Q000235:genetics / Q000502:physiology; D014982:Xenopus laevis / Q000235:genetics*
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D018076:DNA, Complementary; D051380:GRB2 Adaptor Protein; D007328:Insulin; D020033:Protein Isoforms; D011506:Proteins; D012333:RNA, Messenger; D055765:Shc Signaling Adaptor Proteins; D020664:Son of Sevenless Proteins; D029867:Xenopus Proteins