Regulation of AMP-activated protein kinase by multisite phosphorylation in response to agents that elevate cellular cAMP.
The AMP-activated protein kinase (AMPK) and cAMP signaling systems are both key regulators of cellular metabolism. In this study, we show that AMPK activity is attenuated in response to cAMP-elevating agents through modulation of at least two of its alpha subunit phosphorylation sites, viz. alpha-Thr(172) and alpha1-Ser(485)/alpha2-Ser(491), in the clonal beta-cell line INS-1 as well as in mouse embryonic fibroblasts and COS cells. Forskolin, isobutylmethylxanthine, and the glucose-dependent insulinotropic peptide inhibited AMPK activity and reduced phosphorylation of the activation loop alpha-Thr(172) via inhibition of calcium/calmodulin-dependent protein kinase kinase-alpha and -beta, but not LKB1. These agents also enhanced phosphorylation of alpha-Ser(485/491) by the cAMP-dependent protein kinase. AMPK alpha-Ser(485/491) phosphorylation was necessary but not sufficient for inhibition of AMPK activity in response to forskolin/isobutylmethylxanthine. We show that AMPK alpha-Ser(485/491) can be a site for autophosphorylation, which may play a role in limiting AMPK activation in response to energy depletion or other regulators. Thus, our findings not only demonstrate cross-talk between the cAMP/cAMP-dependent protein kinase and AMPK signaling modules, but also describe a novel mechanism by which multisite phosphorylation of AMPK contributes to regulation of its enzyme activity.
Hurley
Rebecca L
RL
Department of Medicine and Biochemistry, Dartmouth Medical School, Remsen 322, N. College St., Hanover, NH 03755, USA.
Barré
Laura K
LK
Wood
Sumintra D
SD
Anderson
Kristin A
KA
Kemp
Bruce E
BE
Means
Anthony R
AR
Witters
Lee A
LA
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D055372:AMP-Activated Protein Kinases; D000818:Animals; D019556:COS Cells; D054737:Calcium-Calmodulin-Dependent Protein Kinase Kinase; D002522:Chlorocebus aethiops; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D005947:Glucose / Q000378:metabolism; D051379:Mice; D009097:Multienzyme Complexes / Q000378:metabolism / Q000502:physiology*; D010455:Peptides / Q000737:chemistry; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism / Q000502:physiology*; D051381:Rats
D009097:Multienzyme Complexes; D010455:Peptides; D010726:Phosphodiesterase Inhibitors; D005576:Colforsin; D000242:Cyclic AMP; D017346:Protein Serine-Threonine Kinases; D054737:Calcium-Calmodulin-Dependent Protein Kinase Kinase; C517701:Camkk1 protein, rat; D055372:AMP-Activated Protein Kinases; D005947:Glucose; D015056:1-Methyl-3-isobutylxanthine