No abstract

p38α Mitogen-activated Protein Kinase (p38α) is activated by a variety of mechanisms, including autophosphorylation initiated by TGFβ-activated kinase 1 binding protein 1 (TAB1) during myocardial ischemia and other stresses. Chemical genetic approaches and co-expression in mammalian, bacterial and cell-free systems revealed that mouse p38α autophosphorylation occurs in cis by direct interaction with TAB1(371-416). In isolated rat cardiac myocytes and perfused mouse hearts TAT-TAB1(371-416) rapidly activates p38 and profoundly perturbs function. Crystal structures and characterization in solution revealed a bipartite docking site for TAB1 in the p38α C-terminal kinase lobe. TAB1 binding stabilizes active p38α and induces rearrangements within the activation segment by helical extension of the Thr-Gly-Tyr motif that allows auto-phosphorylation in cis. Interference with p38α recognition by TAB1 abolishes its cardiac toxicity. Potentially, such intervention could circumvent the drawbacks seen when pharmacological inhibitors of p38 catalytic activity are used clinically.

Abstract-The ischemic activation of p38␣ mitogen-activated protein kinase (p38␣-MAPK) is thought to contribute to myocardial injury. Under other circumstances, activation is through dual phosphorylation by MAPK kinase 3 (MKK3). Therefore, the mkk3 Ϫ/Ϫ murine heart should be protected during ischemia. In retrogradely perfused mkk3 Ϫ/Ϫ and mkk3 ϩ/ϩ mouse hearts subjected to 30 minutes of global ischemia and 120 minutes of reperfusion, infarction/risk volume was similar (50Ϯ5 versus 51Ϯ4, Pϭ0.93, respectively), as was intraischemic p38-MAPK phosphorylation (10 minutes ischemia as percent basal, 608Ϯ224 versus 384Ϯ104, Pϭ0.43, respectively). This occurred despite undetectable activation of MKK3/6 in mkk3 Ϫ/Ϫ hearts. However, tumor necrosis factor (TNF)-induced p38-MAPK phosphorylation was markedly diminished in mkk3 Ϫ/Ϫ vs mkk3 ϩ/ϩ hearts (percent basal, 127Ϯ23 versus 540Ϯ267, respectively, Pϭ0.04), suggesting an MKK-independent activation mechanism by ischemia. Hence, we examined p38-MAPK activation by TAB1-associated autophosphorylation. In wild-type mice and mkk3 Ϫ/Ϫ mice, the p38-MAPK catalytic site inhibitor SB203580 (1 mol/L) diminished phosphorylation during ischemia versus control (10 minutes ischemia as percent basal, 143Ϯ2 versus 436Ϯ96, Pϭ0.003, and 122Ϯ25 versus 623Ϯ176, Pϭ0.05, respectively) and reduced infarction volume (infarction/risk volume, 57Ϯ5 versus 36Ϯ3, PϽ0.001, and 50Ϯ5 versus 29Ϯ3, Pϭ0.003, respectively) but did not alter TNF-induced activation, although in homogenates of ischemic hearts but not TNF-exposed hearts, p38-MAPK was associated with TAB1. Furthermore, adenovirally expressed wild-type and drug-resistant p38␣-MAPK, lacking the SB203580 binding site, was phosphorylated when H9c2 myoblasts were subjected to simulated ischemia. However, SB203580 (1 mol/L) did not prevent the phosphorylation of resistant p38␣-MAPK. These findings suggest the ischemic activation of p38-MAPK contributing to myocardial injury is by TAB1-associated autophosphorylation. Key Words: p38 mitogen-activated protein kinase Ⅲ myocardial infarction Ⅲ TAB1 Ⅲ ischemic preconditioning Ⅲ mitogen-activated protein kinase kinase 3 R ecent attention has focused on myocardial p38 mitogen-activated protein kinase (p38-MAPK) as a critical determinant of myocardial injury during ischemia and reperfusion. 1 In the absence of preconditioning, most investigators find that the activation of p38-MAPK that accompanies true and simulated ischemia/reperfusion contributes to injury. [2][3][4][5][6][7][8][9] Previously, using a variety of approaches, we have shown that the p38␣-MAPK isoform becomes activated and increases cell death during simulated ischemia in cell lines and in adult and neonatal primary cardiocytes. 1,3 Using one of these models, Wang et al 10 have shown that the upstream kinase, mitogen-activated protein kinase kinase-3 (MKK3), preferentially activates p38␣-MAPK by dual phosphorylation and increases cell death. This observation is in keeping with mutational analyses of MKK3 that have mapped the regions within its pr...