Cyclic AMP stimulates Mrp2 translocation by activating p38{alpha} MAPK in hepatic cells.
Cyclic AMP (cAMP) induces translocation of multidrug resistant protein 2 (Mrp2) to the canalicular membrane and activates p38 MAPK in rat hepatocytes. In this study, we tested the hypothesis that cAMP-induced Mrp2 translocation may be mediated via p38 MAPK. Studies were conducted in rat hepatocytes and in a human hepatoma cell line, HuH-7. In rat hepatocytes, cAMP increased Mrp2 translocation and p38 MAPK activity. These effects of cAMP were inhibited by SB203580, an inhibitor of p38 MAPK. Wortmannin, a specific inhibitor of phosphoinositide-3-kinase (PI3K), did not inhibit cAMP induced activation of p38 MAPK, indicating PI3K-independent activation of p38 MAPK by cAMP. To further define the role of p38 MAPK, molecular approaches were used to up- or downregulate p38 MAPK activity in HuH-7 cells using constitutively active (CA) and dominant-negative (DN) MAPK kinase 3 and 6 (MKK3/6). MKK3/6 are upstream kinases responsible for the activation of p38 MAPK. Cells transfected with CAMKK6 showed increased p38 MAPK activity and MRP2 translocation compared with empty vector. cAMP-induced activation of p38 MAPK was inhibited in cells transfected with DNMKK3/6 and DNMKK3, but not with DNMKK6. DNMKK3/6 and DNMKK3 also inhibited cAMP-induced MRP2 translocation. cAMP selectively activated p38alpha MAPK in HuH-7 cells. Knockdown of p38alpha MAPK by short heterodimer RNA resulted in decreased level of p38 MAPK and failure of cAMP to stimulate MRP2 translocation. Taken together, these results suggest that cAMP-induced MRP2 translocation in hepatic cells is mediated via PI3K-independent and MKK3-mediated activation of p38alpha MAPK.
Schonhoff
Christopher M
CM
Dept. of Biomedical Sciences, Cummings School of Veterinary Medicine at Tufts University, North Grafton, Massachusetts 01536, USA.
Webster
Cynthia R L
CR
Anwer
M Sawkat
MS
D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D045744:Cell Line, Tumor; D000242:Cyclic AMP / Q000502:physiology*; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D048669:MAP Kinase Kinase 6 / Q000378:metabolism; D020930:MAP Kinase Kinase Kinases / Q000378:metabolism; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects*; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D014162:Transfection; D000077191:Wortmannin; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
D000730:Androstadienes; D000242:Cyclic AMP; D007093:Imidazoles; D048669:MAP Kinase Kinase 6; D020930:MAP Kinase Kinase Kinases; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D011725:Pyridines; D000077191:Wortmannin; D048051:p38 Mitogen-Activated Protein Kinases; C000715087:ABCC2 protein, human; C116394:MAP kinase kinase kinase 6; C093642:SB 203580