Activation of the mitogen-activated protein kinase ERK1 during meiotic progression of mouse pachytene spermatocytes.
Okadaic acid (OA) causes meiotic progression and chromosome condensation in cultured pachytene spermatocytes and an increase in maturation promoting factor (cyclin B1/cdc2 kinase) activity, as evaluated by H1 phosphorylative activity in anti-cyclin B1 immunoprecipitates. OA also induces a strong increase of phosphorylative activity toward the mitogen-activated protein kinase substrate myelin basic protein (MBP). Immunoprecipitation experiments with anti-extracellular signal-regulated kinase 1 (ERK1) or anti-ERK2 antibodies followed by MBP kinase assays, and direct in-gel kinase assays for MBP, show that p44/ERK1 but not p42/ERK2 is stimulated in OA-treated spermatocytes. OA treatment stimulates phosphorylation of ERK1, but not of ERK2, on a tyrosine residue involved in activation of the enzyme. ERK1 immunoprecipitated from extracts of OA-stimulated spermatocytes induces a stimulation of H1 kinase activity in extracts from control pachytene spermatocytes, whereas immunoprecipitated ERK2 is uneffective. We also show that natural G(2)/M transition in spermatocytes is associated to intracellular redistribution of ERKs, and their association with microtubules of the metaphase spindle. Preincubation of cultured pachytene spermatocytes with PD98059 (a selective inhibitor of ERK-activating kinases MEK1/2) completely blocks the ability of OA to induce chromosome condensation and progression to meiotic metaphases. These results suggest that ERK1 is specifically activated during G(2)/M transition in mouse spermatocytes, that it contributes to the mechanisms of maturation promoting factor activation, and that it is essential for chromosome condensation associated with progression to meiotic metaphases.
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Dipartimento di Sanità Pubblica e Biologia Cellulare, Sezione di Anatomia, Università di Roma "Tor Vergata," Via O. Raimondo 8, 00173, Rome, Italy.
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D000818:Animals; D016203:CDC2 Protein Kinase / Q000378:metabolism; D002478:Cells, Cultured; D002875:Chromosomes; D019926:Cyclin B / Q000378:metabolism; D056744:Cyclin B1; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016195:G2 Phase; D008297:Male; D008540:Meiosis; D051379:Mice; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008938:Mitosis; D019319:Okadaic Acid / Q000494:pharmacology; D010766:Phosphorylation; D013090:Spermatocytes / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology; D013347:Subcellular Fractions / Q000201:enzymology; D014443:Tyrosine / Q000378:metabolism
C534700:Ccnb1 protein, mouse; D019926:Cyclin B; D056744:Cyclin B1; D004791:Enzyme Inhibitors; D019319:Okadaic Acid; D014443:Tyrosine; D016203:CDC2 Protein Kinase; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases