Tyrosine phosphorylation and activation of homologous protein kinases during oocyte maturation and mitogenic activation of fibroblasts.

Abstract

Meiotic maturation of Xenopus and sea star oocytes involves the activation of a number of protein-serine/ threonine kinase activities, including a myelin basic protein (MBP) kinase. A 44-kDa MBP kinase (p44 pk) purified from mature sea star oocytes is shown here to be phosphorylated at tyrosine. Antiserum to purified sea star p44 Pk was used to identify antigenically related proteins in Xenopus oocytes. Two tyrosine-phosphorylated 42-kDa proteins (p42) were detected with this antiserum in Xenopus eggs. Xenopus p42 chromatographs with MBP kinase activity on a Mono Q ion-exchange column. Tyrosine phosphorylation ofXenopus p42 approximately parallels MBP kinase activity during meiotic maturation. These results suggest that related MBP kinases are activated during meiotic maturation of Xenopus and sea star oocytes. Previous studies have suggested that Xenopus p42 is related to the mitogen-activated protein (MAP) kinases of cultured mammalian cells. We have cloned a MAP kinase relative from a Xenopus ovary cDNA library and demonstrate that this clone encodes the Xenopus p42 that is tyrosine phosphorylated during oocyte maturation. Comparison of the sequences of Xenopus p42 and a rat MAP kinase (ERKI) and peptide sequences from sea star p44"!Ik indicates that these proteins are close relatives. The family members appear to be tyrosine phosphorylated, and activated, in different contexts, with the murine MAP kinase active during the transition from quiescence to the GI stage of the mitotic cell cycle and the sea star and Xenopus kinases being active during M phase of the meiotic cell cycle.Progesterone induces Xenopus laevis oocytes to maturethat is, to leave first meiotic prophase and progress part way through meiosis and arrest as eggs, in second meiotic metaphase. A burst in the phosphorylation of numerous proteins is triggered near the time of nuclear envelope (germinal vesicle) breakdown (GVBD) (46). Similarly, with sea star oocytes, 1-methyladenine induces both maturation and protein phosphorylation (19). Only a few of the phosphoproteins in maturing oocytes have been identified, such as the 40S ribosomal protein S6 (33,38,39,51), nucleoplasmin (17), nuclear lamins (48), and possibly histone Hi. Using these and other proteins as substrates, several maturation-activated protein-serine/threonine kinases have been characterized from sea star oocytes (6, 42, 52, 55) and Xenopus oocytes (12, 44, 47; reviewed in reference 53).The major histone Hi kinases that are activated during Xenopus (20, 28) and sea star (6, 42) oocyte maturation are the amphibian and echinoderm homologs of the 34-kDa protein kinase encoded by the Schizosaccharomyces pombe mitotic control gene cdc2. Microinjection of active preparations of histone Hi kinase into immature oocytes is sufficient to induce GVBD (44). Two ribosomal S6 kinases (S6K-I and S6K-II) are also activated at GVBD in progesterone-treated Xenopus oocytes (12,44,47) or following microinjection of the frog homolog of p34cdc2 (68). Both S6 kinases appear to be activated as a consequ...

Inhibition of c-Jun DNA binding by mitogen-activated protein kinase.

“…Both sequences are found in the predicted protein sequences of two Xenopus ERK2-like cDNA clones (Gotoh et al, 1991b;Posada et al, 1991). Purified Xenopus p42 was recognized by antisera 837 and DC3, both of which recognize p42 ERK2 proteins.…”

“…MAP kinases are also activated downstream from M-phase promoting factor during oocyte maturation (Cicirelli et al, 1988;Cooper, 1989;Sanghera et al, 1990;Ferrell et al, 1991;Gotoh et al, 1991a,b;Posada et al, 1991;Posada and Cooper, 1992). MAP kinases may therefore be involved in mitogenesis, in activation and differentiation responses in neuronal cells, and in the G2-M transition in oocytes.…”

“…A 13 amino acid peptide containing these sites (LTEYVATRWYRAPE) is perfectly conserved in the predicted protein sequences of ERK1, ERK2, and two more distant relatives, the Saccharomyces cerevisiae KSS1 and FUS3 gene products (Courchesne et al, 1989;Elion et al, 1989;Boulton et al, 1990Boulton et al, , 1991bGotoh et al, 1991b;Her et al, 1991;Posada et al, 1991), raising the possibility that all of these kinases share a common mechanism for their activation.…”

“…These include p44mPk, a sea star MAP kinase that lacks the canonical threonine phosphorylation site (Sanghera et al, 1990;Posada et al, 1991); p54 MAP kinase, an activity detected in the livers of cycloheximide-treated rats (Kyriakis and Avruch, 1990); the as yet uncharacterized product of the ERK3 gene (Boulton et al, 1991b); and a 45-kDa putative ERK4 protein identified by antibody cross-reactivity (Boulton et al, 1991b).…”

STAT activation by the PDGF receptor requires juxtamembrane phosphorylation sites but not Src tyrosine kinase activation

“…Controls (co) were treated with diluent for 10 min. Cells were harvested, lysed in RIPA bu er, separated by 10% SDS ± PAGE and analysed by Western blot analysis using rabbit anti-ERK2 serum (1913) (Posada et al, 1991). (b) c-fos expression.…”