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Activation of mitogen activated protein (MAP) kinases by vanadate is independent of insulin receptor autophosphorylation
Author(s) -
D'Onofrio Franco,
Le Minh Quynh Uyen,
Chiasson Jean-Louis,
Srivastava Ashok K.
Publication year - 1994
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(94)80152-5
Subject(s) - autophosphorylation , insulin receptor , vanadate , chinese hamster ovary cell , mapk14 , mitogen activated protein kinase , mapk7 , mitogen activated protein kinase kinase , irs2 , phosphorylation , receptor tyrosine kinase , biology , map2k7 , protein kinase a , biochemistry , tyrosine phosphorylation , grb10 , map kinase kinase kinase , microbiology and biotechnology , cyclin dependent kinase 2 , insulin , endocrinology , receptor , insulin resistance
Treatment of Chinese hamster ovary (CHO) cells over‐expressing the human insulin receptor (CHO‐HIRc) with the insulin mimetic agent, vanadate, resulted in a dose‐ and time‐dependent tyrosine phosphorylation of two proteins with apparent molecular sizes of 42 kDa (p42) and 44 kDa (p44). However, vanadate was unable to stimulate the tyrosyi phosphorylation of theβ‐subunit of the insulin receptor. By using myelin basic protein (MBP) as the substrate to measure mitogen‐activated protein (MAP) kinase activity in whole cell lysates, vanadate‐stimulated tyrosyl phosphorylation of p42 and p44 was associated with a dose‐ and time‐dependent activation of MAP kinase activity. Furthermore, affinity purification of cell lysates on anti‐phosphotyrosine agarose column followed by immunoblotting with a specific antibody to MAP kinases demonstrated that vanadate treatment increased the tyrosyl phosphorylation of both p44 mapk and p42 mapk by several folds, as compared to controls, in concert with MAP kinase activation. In addition, retardation in gel mobility further confirmed that vanadate treatment increased the phosphorylation of p44 mapk and p42 mapk in CHO‐HIRc. A similar effect of vanadate on MAP kinase tyrosyl phosphorylation and activation was also observed in CHO cells over‐expressing a protein tyrosine kinase‐deficient insulin receptor (CHO‐1018). These results demonstrate that the protein tyrosine kinase activity of the insulin receptor may not be required in the signaling pathways leading to the vanadate‐mediated tyrosyl phosphorylation and activation of MAP kinases.