TYROSYL-PHOSPHORYLATED PROTEINS ARE INVOLVED IN REGULATION OF MEIOSISIN THE RAT EGG

Fertilization in invertebrates results in tyrosine (Tyr) phosphorylati on of several egg proteins. However, the involvement of Tyr phosphoryl ation in mediating mammalian egg activation has not yet been investiga ted. Using an antibody specific for phosphotyrosine (P-Tyr), immunoblo tting, and densitometric analysis, we found that maturation of the ooc yte is accompanied by a generalized increase in the P-Tyr content of a lmost all egg proteins detected. After sperm penetration, 5 of the 17 protein bands detected demonstrated a small increase in their P-Tyr co ntent, while at the pronuclear (PN) stage the signal was markedly redu ced. Ionomycin emulated the changes observed at fertilization in most protein bands detected, demonstrating a small increase in their P-Tyr content within 15 min of exposure. Analysis of the involvement of the tyrosyl-phosphorylated, mitogen-activated protein (MAP) kinase during meiosis revealed comigration of the phosphotyrosyl bands with the prot ein and a good correlation with its enzyme activity. Maturation was ac companied by an increase in MAP kinase activity. The activity dropped partially after sperm penetration and furthermore later at the PN stag e. A larger quantity accompanied by a more significant, change in the P-Tyr content implies for extracellular regulated kinase (ERK) 2 being the dominant isoform present in the rat egg. Our results indicate tha t fertilization in mammals involves changes in activity of protein tyr osine kinases (PTKs) or in the balance between PTKs and protein tyrosi ne phosphatases. The single, ionomycin-induced Ca2+ rise is sufficient to imitate fertilization-induced changes in MBP kinase activity, as w ell as in tyrosine phosphorylation of other proteins within the egg. ( C) 1998 Wiley-Liss, Inc.