title: Phosphorylation of the kinase interaction motif in mitogen-activated protein (MAP) kinase phosphatase-4 mediates cross-talk between protein kinase A and MAP kinase signaling pathways.
abstract: MAP kinase phosphatase 4 (DUSP9/MKP-4) plays an essential role during placental development and is one of a subfamily of three closely related cytoplasmic dual-specificity MAPK phosphatases, which includes the ERK-specific enzymes DUSP6/MKP-3 and DUSP7/MKP-X. However, unlike DUSP6/MKP-3, DUSP9/MKP-4 also inactivates the p38α MAP kinase both in vitro and in vivo. Here we demonstrate that inactivation of both ERK1/2 and p38α by DUSP9/MKP-4 is mediated by a conserved arginine-rich kinase interaction motif located within the amino-terminal non-catalytic domain of the protein. Furthermore, DUSP9/MKP-4 is unique among these cytoplasmic MKPs in containing a conserved PKA consensus phosphorylation site (55)RRXSer-58 immediately adjacent to the kinase interaction motif. DUSP9/MKP-4 is phosphorylated on Ser-58 by PKA in vitro, and phosphorylation abrogates the binding of DUSP9/MKP-4 to both ERK2 and p38α MAP kinases. In addition, although mutation of Ser-58 to either alanine or glutamic acid does not affect the intrinsic catalytic activity of DUSP9/MKP-4, phospho-mimetic (Ser-58 to Glu) substitution inhibits both the interaction of DUSP9/MKP-4 with ERK2 and p38α in vivo and its ability to dephosphorylate and inactivate these MAP kinases. Finally, the use of a phospho-specific antibody demonstrates that endogenous DUSP9/MKP-4 is phosphorylated on Ser-58 in response to the PKA agonist forskolin and is also modified in placental tissue. We conclude that DUSP9/MKP-4 is a bona fide target of PKA signaling and that attenuation of DUSP9/MKP-4 function can mediate cross-talk between the PKA pathway and MAPK signaling through both ERK1/2 and p38α in vivo.
authors: CR-UK Stress Response Laboratory, Medical Research Institute, Ninewells Hospital and Medical School, University of Dundee, Dundee DD1 9SY, United Kingdom.
authors: Robin J
authors: RJ
authors: Dickinson
authors: CR-UK Stress Response Laboratory, Medical Research Institute, Ninewells Hospital and Medical School, University of Dundee, Dundee DD1 9SY, United Kingdom.
authors: Laurent
authors: L
authors: Delavaine
authors: Centro de Investigación Príncipe Felipe, Avda. Autopista del Saler 16-3, 46013 Valencia, Spain.
authors: Rocío
authors: R
authors: Cejudo-Marín
authors: CR-UK Stress Response Laboratory, Medical Research Institute, Ninewells Hospital and Medical School, University of Dundee, Dundee DD1 9SY, United Kingdom.
authors: Graeme
authors: G
authors: Stewart
authors: CR-UK Stress Response Laboratory, Medical Research Institute, Ninewells Hospital and Medical School, University of Dundee, Dundee DD1 9SY, United Kingdom.
authors: Christopher J
authors: CJ
authors: Staples
authors: CR-UK Stress Response Laboratory, Medical Research Institute, Ninewells Hospital and Medical School, University of Dundee, Dundee DD1 9SY, United Kingdom.
authors: Mark P
authors: MP
authors: Didmon
authors: Instituto de Biología y Genética Molecular, Consejo Superior de Investigaciones Científicas-Universidad de Valladolid, c/Sanz y Forés s/n, 47003 Valladolid, Spain.
authors: Antonio Garcia
authors: AG
authors: Trinidad
authors: Instituto de Biología y Genética Molecular, Consejo Superior de Investigaciones Científicas-Universidad de Valladolid, c/Sanz y Forés s/n, 47003 Valladolid, Spain.
authors: Andrés
authors: A
authors: Alonso
authors: Centro de Investigación Príncipe Felipe, Avda. Autopista del Saler 16-3, 46013 Valencia, Spain.
authors: Rafael
authors: R
authors: Pulido
authors: CR-UK Stress Response Laboratory, Medical Research Institute, Ninewells Hospital and Medical School, University of Dundee, Dundee DD1 9SY, United Kingdom. Electronic address: s.m.keyse@dundee.ac.uk.
authors: Stephen M
authors: SM
authors: Keyse
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D019556:COS Cells; D002384:Catalysis; D020134:Catalytic Domain; D002522:Chlorocebus aethiops; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D054637:Dual-Specificity Phosphatases / Q000378:metabolism*; D006801:Humans; D051379:Mice; D054639:Mitogen-Activated Protein Kinase Phosphatases / Q000378:metabolism*; D010766:Phosphorylation; D011994:Recombinant Proteins / Q000737:chemistry; D015398:Signal Transduction; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D017868:Cyclic AMP-Dependent Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D054639:Mitogen-Activated Protein Kinase Phosphatases; C104821:DUSP9 protein, human; D054637:Dual-Specificity Phosphatases; C484491:Dusp9 protein, mouse