alpha 1-AMP-Activated Protein Kinase Regulates Hypoxia-Induced Na,K-ATPase Endocytosis via Direct Phosphorylation of Protein Kinase C zeta

作者：Gusarova Galina A; Dada Laura A; Kelly Aileen M; Brodie Chaya; Witters Lee A; Chandel Navdeep S; Sznajder Jacob I^*

来源：Molecular and Cellular Biology, 2009, 29(13): 3455-3464.

DOI：10.1128/MCB.00054-09

Hypoxia promotes Na,K-ATPase endocytosis via protein kinase C zeta (PKC zeta)-mediated phosphorylation of the Na,K-ATPase alpha subunit. Here, we report that hypoxia leads to the phosphorylation of 5'-AMP-activated protein kinase ( AMPK) at Thr172 in rat alveolar epithelial cells. The overexpression of a dominant-negative AMPK alpha subunit (AMPK-DN) construct prevented the hypoxia-induced endocytosis of Na,K-ATPase. The overexpression of the reactive oxygen species (ROS) scavenger catalase prevented hypoxia-induced AMPK activation. Moreover, hypoxia failed to activate AMPK in mitochondrion-deficient rho(0)-A549 cells, suggesting that mitochondrial ROS play an essential role in hypoxia-induced AMPK activation. Hypoxia-induced PKC zeta translocation to the plasma membrane and phosphorylation at Thr410 were prevented by the pharmacological inhibition of AMPK or by the overexpression of the AMPK-DN construct. We found that AMPK alpha phosphorylates PKC zeta on residue Thr410 within the PKC zeta activation loop. Importantly, the activation of AMPK alpha was necessary for hypoxia-induced AMPK-PKC zeta binding in alveolar epithelial cells. The overexpression of T410A mutant PKC zeta prevented hypoxia-induced Na,K-ATPase endocytosis, confirming that PKC zeta Thr410 phosphorylation is essential for this process. PKC zeta activation by AMPK is isoform specific, as small interfering RNA targeting the alpha 1 but not the alpha 2 catalytic subunit prevented PKC zeta activation. Accordingly, we provide the first evidence that hypoxia-generated mitochondrial ROS lead to the activation of the AMPK alpha 1 isoform, which binds and directly phosphorylates PKC zeta at Thr410, thereby promoting Na,K-ATPase endocytosis.

• 出版日期2009-7-1
• 单位西北大学