Open AccessThe phorbol ester-dependent activator of the mitogen-activated protein kinase p42mapk is a kinase with specificity for the threonine and tyrosine regulatory sites.
Author(s) -
Anthony Rossomando,
Jie Wu,
Michael J. Weber,
Thomas W. Sturgill
Publication year - 1992
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.89.12.5221
Subject(s) - mapk14 , map2k7 , mitogen activated protein kinase kinase , autophosphorylation , c raf , map kinase kinase kinase , protein tyrosine phosphatase , cyclin dependent kinase 9 , mitogen activated protein kinase , ask1 , cyclin dependent kinase 2 , biochemistry , biology , microbiology and biotechnology , protein kinase a , protein kinase c , tyrosine kinase , mapkapk2 , chemistry , kinase , signal transduction
Mitogen-activated protein kinases (MAP kinases) are activated by dual tyrosine and threonine phosphorylations in response to various stimuli, including phorbol esters. To define the mechanism of activation, recombinant wild-type 42-kDa MAP kinase (p42mapk) and a kinase-defective mutant of p42mapk (K52R) were used to assay both activator activity for p42mapk and kinase activity toward K52R in stimulated EL4.I12 mouse thymoma cells. Phorbol 12,13-dibutyrate (10 min, 650 nM) stimulated a single peak of MAP kinase activator that was coeluted from Mono Q at pH 7.5 and 8.9 with K52R kinase activity. Both activities were inactivated by the serine/threonine-specific phosphatase 2A but not by the tyrosine-specific phosphatase CD45. Phosphorylation of K52R occurred specifically on Thr-183 and Tyr-185, as determined by tryptic phosphopeptide mapping in comparison with synthetic marker phosphopeptides. These findings indicate that phorbol ester-stimulated MAP kinase kinase can activate p42mapk by threonine and tyrosine phosphorylations, and that p42mapk thus does not require an autophosphorylation reaction.