N488I mutation of the gamma2-subunit results in bidirectional changes in AMP-activated protein kinase activity.
Mutations in the human gene encoding the nucleotide-binding region in the gamma-subunit of AMP-activated protein kinase (AMPK) cause cardiomyopathy with preexcitation syndrome. Mutant AMPK showed reduced binding affinity to nucleotides in vitro raising the possibility that altered regulation of AMPK activity by AMP/ATP could contribute to the disease phenotype. In this study, we determined the sensitivity of AMPK activity to AMP/ATP in the beating hearts using transgenic mice expressing a mutant (N488I, gamma2-mutant) or wild-type gamma2-subunit (gamma2-TG). The [ATP] and [AMP] were unaltered in all hearts but the AMPK activity was increased by 2.5-fold in gamma2-mutant hearts freeze-clamped at normal AMP/ATP compared with nontransgenic (WT) or gamma2-TG. The increased basal AMPK activity was caused by increased Thr-172 phosphorylation of the alpha-subunit (p-AMPK, by 4-fold) at normal [ATP] and was not changed by reducing glycogen content by 60% in the gamma2-mutant hearts. A reversal of AMP/ATP, caused by ATP degradation, increased p-AMPK by 7-fold in WT but caused no change in gamma2-mutant hearts. These results demonstrate that the mutation renders AMPK insensitive to the inhibitory and stimulatory effects of the regulatory nucleotides ATP and AMP, respectively, suggesting that the pathogenesis of the human disease may not be attributable to a simple loss- or gain-of-function.
Zou
Liqun
L
NMR Laboratory for Physiological Chemistry, Brigham and Women's Hospital, Boston, MA, USA.
Shen
Mei
M
Arad
Michael
M
He
Huamei
H
Løfgren
Bo
B
Ingwall
Joanne S
JS
Seidman
Christine E
CE
Seidman
Jon G
JG
Tian
Rong
R
D055372:AMP-Activated Protein Kinases; D000249:Adenosine Monophosphate / Q000032:analysis* / Q000502:physiology; D000255:Adenosine Triphosphate / Q000032:analysis* / Q000502:physiology; D000818:Animals; D006003:Glycogen / Q000032:analysis; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D009097:Multienzyme Complexes / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D009154:Mutation; D009206:Myocardium / Q000201:enzymology*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D021122:Protein Subunits
D009097:Multienzyme Complexes; D021122:Protein Subunits; D000249:Adenosine Monophosphate; D000255:Adenosine Triphosphate; D006003:Glycogen; D017346:Protein Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases