The regulation of AMP-activated protein kinase by H(2)O(2).
AMP-activated protein kinase (AMPK), a heterotrimeric serine/threonine kinase, is activated by conditions leading to an increase of the intracellular AMP:ATP ratio. However, how AMPK is regulated under the oxidative stress is completely unknown. In the present study, we examined effects of the oxidative agent H(2)O(2) on AMPK. AMPK was transiently and concentration-dependently activated by H(2)O(2) in NIH-3T3 cells. This activation was tightly associated with an increased AMP:ATP ratio, an electrophoretic mobility shift of AMPK alpha1 catalytic subunit, and an increased phosphorylation level of AMPK alpha1 threonine 172, which is a major in vitro phosphorylation site by the upstream AMPK kinase. All of these events were significantly blocked by the pretreatment of 0.5% dimethyl sulfoxide, a potent hydroxyl radical scavenger, indicating that AMPK cascades are highly sensitive to the oxidative stress. Interestingly, a specific tyrosine kinase inhibitor, genistein, further stimulated the H(2)O(2)-induced AMPK activity by 70% without altering the AMP:ATP. Taken together, our results suggest that AMP:ATP ratio is the major parameter to which AMPK responds under the oxidative stress, but AMPK may be regulated in part by a tyrosine kinase-dependent pathway, which is independent of the cellular adenosine nucleotides level.
Choi
S L
SL
Department of Molecular Biology, East-West Medical Research Center, College of Medicine, Seoul 130-701, Korea.
Kim
S J
SJ
Lee
K T
KT
Kim
J
J
Mu
J
J
Birnbaum
M J
MJ
Soo Kim
S
S
Ha
J
J
D016475:3T3 Cells; D055372:AMP-Activated Protein Kinases; D000249:Adenosine Monophosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002384:Catalysis; D004789:Enzyme Activation / Q000187:drug effects; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D051379:Mice; D009097:Multienzyme Complexes / Q000378:metabolism*; D018384:Oxidative Stress / Q000502:physiology*; D010766:Phosphorylation / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D013912:Threonine / Q000378:metabolism
D009097:Multienzyme Complexes; D017382:Reactive Oxygen Species; D013912:Threonine; D000249:Adenosine Monophosphate; D000255:Adenosine Triphosphate; D006861:Hydrogen Peroxide; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases