Characterization of AMP-activated protein kinase γ-subunit isoforms and their role in AMP binding

Abstract

The AMP-activated protein kinase (AMPK) cascade plays an important role in the regulation of energy homeostasis within the cell. AMPK is a heterotrimer composed of a catalytic subunit (alpha) and two regulatory subunits (beta and gamma). We have isolated and characterized two isoforms of the gamma subunit, termed gamma2 and gamma3. Both gamma2 (569 amino acids) and gamma3 (492 amino acids) have a long N-terminal domain which is not present in the previously characterized isoform, gamma1. As with gamma1, mRNA encoding gamma2 is widely expressed in human tissues, whereas significant expression of gamma3 mRNA was only detected in skeletal muscle. Using isoform-specific antibodies, we determined the AMPK activity associated with the different gamma isoforms in a number of rat tissues. In most tissues examined more than 80% of total AMPK activity was associated with the gamma1 isoform, with the remaining activity being accounted for mainly by the gamma2 isoform. Exceptions to this were testis and, more notably, brain where all three isoforms contributed approximately equally to activity. There was no evidence for any selective association between the alpha1 and alpha2isoforms and the various gamma isoforms. However, the AMP-dependence of the kinase complex is markedly affected by the identity of the gamma isoform present, with gamma2-containing complexes having the greatest AMP-dependence, gamma3 the lowest, and gamma1 having an intermediate effect. Labelling studies, using the reactive AMP analogue 8-azido-[(32)P]AMP, indicate that the gamma subunit may participate directly in the binding of AMP within the complex.

Lack of starvation-induced activation of AMP-activated protein kinase in the hypothalamus of the Lou/C rats resistant to obesity

“…The a2 AMPK activity was similar to that previously reported 8 but represented only 5% of the total activity of AMPK, in agreement with the repartition of a isoforms of AMPK reported for the whole brain. 24 Immunoblots with specific antibodies confirmed that the a1 isoform of AMPK was predominant in the whole hypothalamus. The liver, which contains about equal amounts of both AMPK isoforms, was taken as a control (Figure 1d).…”

AMP-activated protein kinase inhibits IL-6-stimulated inflammatory response in human liver cells by suppressing phosphorylation of signal transducer and activator of transcription 3 (STAT3)

“…Previously it has been demonstrated that in the rodent liver, the AMPKα1-and AMPKα2-containing complexes account for about half of total AMPK activity each [23]. In human liver, and in the human hepatocyte cell line HepG2, the abundance of the two isoforms of the catalytic subunit has not been previously described.…”

Role of 5′AMP-activated protein kinase in skeletal muscle

“…5 This increase in AMP levels leads to allosteric activation of AMPK through AMP binding to the g-subunit. 10 Recent data suggest that AMP binding also induces a conformational change in the kinase domain leading to protection of Thr172 from dephosphorylation. Thus, increases in AMP levels play dual roles in activation of AMPK.…”