Pertussis toxin-catalyzed ADP-ribosylation of a G protein in mouse oocytes, eggs, and preimplantation embryos: developmental changes and possible functional roles.
G proteins, which in many somatic cells serve as mediators of signal transduction, were identified in preimplantation mouse embryos by their capacity to undergo pertussis toxin-catalyzed ADP-ribosylation. Two pertussis toxin (PT) substrates with Mr = 38,000 and 39,000 (alpha 38 and alpha 39) are present in approximately equal amounts. Relative to the amount in freshly isolated germinal vesicle (GV)-intact oocytes, the amount of PT-catalyzed ADP-ribosylation of alpha 38-39 falls during oocyte maturation, rises between the one- and two-cell stages, falls by the eight-cell and morula stages, and increases again by the blastocyst stage. The decrease in PT-catalyzed ADP-ribosylation of alpha 38-39 that occurs during oocyte maturation, however, does not require germinal vesicle breakdown (GVBD), since inhibiting GVBD with 3-isobutyl-1-methyl xanthine (IBMX) does not prevent the decrease in the extent of PT-catalyzed ADP-ribosylation. A biologically active phorbol diester (12-O-tetradecanoyl phorbol 13-acetate, TPA), but not an inactive one (4 alpha-phorbol 12,13-didecanoate, 4 alpha-PDD), totally inhibits the increase in PT-catalyzed ADP-ribosylation of alpha 38-39 that occurs between the one- and two-cell stage; TPA inhibits cleavage, but not transcriptional activation, which occurs in the two-cell embryo (Poueymirou and Schultz, 1987. Dev. Biol. 121, 489-498). In contrast, cytochalasin D, genistein, or aphidicolin, each of which inhibits cleavage of one-cell embryos, or alpha-amanitin or H8, each of which inhibits transcriptional activation but not cleavage of one-cell embryos, have little or much smaller inhibitory effects on the increase in PT-catalyzed ADP-ribosylation of alpha 38-39. Results of immunoblotting experiments using an antibody that is highly specific for alpha il-3 reveal the presence of a cross-reactive species of Mr = 38,000 (alpha 38) in the GV-intact oocyte, metaphase II-arrested egg, and one-, two-cell embryos. Relative to these stages, a reduced amount of this species is present in the eight-cell, morula, and blastocyst stages. Treatment of oocytes with PT results in a small but significant acceleration in the rate of GVBD, but has no effect on the extent of polar body emission. Treatment of one-cell embryos with PT has no effect on the extent of cleavage, onset of transcriptional activation at the two-cell stage, or development of two-cell embryos to the hatching blastocyst stage.
Jones
J
J
Department of Biology, University of Pennsylvania, Philadelphia 19104-6018.
Schultz
R M
RM
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism*; D000818:Animals; D001755:Blastocyst / Q000166:cytology / Q000378:metabolism*; D002455:Cell Division; D002462:Cell Membrane / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D019204:GTP-Binding Proteins / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008715:Methionine / Q000378:metabolism; D051379:Mice; D008815:Mice, Inbred Strains; D008970:Molecular Weight; D009243:NAD / Q000378:metabolism*; D009865:Oocytes / Q000166:cytology / Q000378:metabolism*; D010063:Ovum / Q000166:cytology / Q000378:metabolism*; D037342:Pertussis Toxin; D013462:Sulfur Radioisotopes; D014158:Transcription, Genetic / Q000187:drug effects; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*; D015044:Zona Pellucida / Q000502:physiology
D013462:Sulfur Radioisotopes; D010566:Virulence Factors, Bordetella; D009243:NAD; D000246:Adenosine Diphosphate Ribose; D008715:Methionine; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins