Distinct binding determinants for ERK2/p38 alpha and JNK MAP kinases mediate catalytic activation and substrate selectivity of MAP kinase phosphatase-1

Mitogen-activated protein (MAP) kinase phosphatase 1 (MKP-1/CL100) is an in ducible nuclear dual specificity protein phosphatase that can dephosphoryla te and inactivate both mitogen- and stress-activated protein kinases in vit ro and in vivo, However, the molecular mechanism responsible for the substr ate selectivity of MKP-1 is unknown, In addition, it has been suggested tha t the signal transducers and activators of transcription 1 (STAT1) transcri ption factor is a physiological non-MAP kinase substrate for MKP-1, We have used the yeast two-hybrid assay to demonstrate that MKP-1 is able to inter act selectively with the extracellular signal-regulated kinase 1/2 (ERK1/2) , p38 alpha, and c-Jun NH2-terminal kinase (JNK) MAP kinase isoforms, Furth ermore, this binding is accompanied by catalytic activation of recombinant MKP-1 protein in vitro, and these end points show an absolute correlation w ith MKP-1 substrate selectivity in vivo. In contrast, MKP-1 does not intera ct with STAT1, Recombinant STAT1 does not cause catalytic activation of MKP -1; nor does MKP-1 block tyrosine phosphorylation of STAT1 in vivo. Both bi nding and catalytic activation of MKP-1 are abrogated by mutation of a cons erved docking site in ERK2, p38 alpha, and JNK1 MAP kinases, Within MKP-1, MAP kinase binding is mediated by the amino-terminal noncatalytic domain of the protein. However, mutation of a conserved cluster of positively charge d residues within this domain abolishes the binding and activation of MKP-1 by ERK2 and p38 alpha but not JNK1, indicating that there are distinct bin ding determinants for these MAP kinase isoforms, We conclude that the subst rate selectivity of MKP-1 is determined by specific protein-protein interac tions coupled with catalytic activation of the phosphatase and that these i nteractions are restricted to members of the MAP kinase family of enzymes.