Protein modification by phosphorylation during the process of nuclear membrane dissolution in puromycin-treated mouse oocytes.
The present study was undertaken to elucidate the mechanism of nuclear membrane dissolution (NMD) in puromycin-treated mouse oocytes. Treatment of germinal vesicle breakdown (GVBD) oocytes with puromycin (50 micrograms/ml) induced chromosome decondensation with formation of a polar body; these are designated nuclear membrane (NM) oocytes. After withdrawal of puromycin, NM oocytes underwent NMD (approximately 70%) during a 12-h culture period. Either dibutyryl cyclic AMP (dbcAMP, 25-100 micrograms/ml) or isobutylmethylxanthine (IBMX, 0.1-1.0 mM) inhibited the process of NMD in a dose-dependent manner, suggesting the involvement of cAMP in the process of NMD. To determine which protein(s) participated in the transition from interphase to metaphase II during NMD, NM oocytes were labeled with [35S]methionine, and one- and two-dimensional gel electrophoresis were performed. Although the synthesis of stage-specific proteins during NMD was not found, two specific proteins of Mr 27,000 and 46,000, which were synthesized at interphase following removal of puromycin, were modified during NMD. Phosphatase treatment and 32PO4-labeling experiments indicated that phosphorylation was responsible for these modifications, which were inhibited by either dbcAMP or IBMX. Therefore, it appears that phosphorylation of specific proteins may play an important role in the transition from interphase to metaphase II.
Kang
H M
HM
Department of Zoology, College of Natural Sciences, Seoul National University, Korea.
Cho
C
C
Lee
K K
KK
Kwon
H B
HB
Kim
K
K
Cho
W K
WK
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000818:Animals; D003994:Bucladesine / Q000494:pharmacology; D005260:Female; D066298:In Vitro Techniques; D051379:Mice; D008813:Mice, Inbred ICR; D008970:Molecular Weight; D009685:Nuclear Envelope / Q000187:drug effects* / Q000378:metabolism; D009865:Oocytes / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D010766:Phosphorylation; D011506:Proteins / Q000302:isolation & purification / Q000378:metabolism*; D011691:Puromycin / Q000494:pharmacology*
D011506:Proteins; D011691:Puromycin; D003994:Bucladesine; D015056:1-Methyl-3-isobutylxanthine