Adenosine Activates AMPK to Phosphorylate Bcl-XL Responsible for Mitochondrial Damage and DIABLO Release in HuH-7 Cells

Background/Aims: Accumulating evidence has pointed to AMP-activated protein kinase (AMPK) as an inducer of apoptosis in a variety of cancer cells. The present study aimed at understanding AMPK signals for adenosine-induced HuH-7 cell apoptosis. Methods: Cell viability, AMPK activity, mitochondrial membrane potentials, phosphorylation of Bcl-X-L, in situ DIABLO mobilizations, and caspase-3 activity were monitored in HuH-7 cells. Plasmid DNAs for DIABLO-GFP, mutant Bcl-X-L, dominant negative mutant AMPK alpha 2 and the siRNAs to silence the AMPK alpha 1 or AMPK alpha 2 targeted gene were constructed and transfected. Results: Adenosine or the AMPK activator AICAR induced apoptosis in HuH-7 cells, and no synergistic effect was obtained with co-treatment. Adenosine activated AMPK, to phosphorylate Bcl-X-L. Adenosine or AICAR disrupted mitochondrial membrane potentials, and the effect was inhibited by knocking-down AMPK alpha 1 and/or AMPK alpha 2, expressing dominant negative mutant AMPK alpha 2 or mutant Bcl-X-L lacking Ser/Thr phosphorylation sites. AICAR stimulated DIABLO release from the mitochondria, and the release was suppressed by expressing the mutant Bcl-X-L. AICAR activated caspase-3, which was also inhibited by expressing the mutant Bcl-X-L. Conclusion: Adenosine activates AMPK, to disrupt mitochondrial membrane potentials through Bcl-X-L phosphorylation, allowing DIABLO release from the mitochondria, as a factor for caspase-3 activation to induce HuH-7 cell apoptosis. Copyright (C) 2011 S. Karger AG, Basel