The α2–5′AMP-Activated Protein Kinase Is a Site 2 Glycogen Synthase Kinase in Skeletal Muscle and Is Responsive to Glucose Loading

Abstract

The 5AMP-activated protein kinase (AMPK) is a potential antidiabetic drug target. Here we show that the pharmacological activation of AMPK by 5-aminoimidazole-1-␤-4-carboxamide ribofuranoside (AICAR) leads to inactivation of glycogen synthase (GS) and phosphorylation of GS at Ser 7 (site 2). In muscle of mice with targeted deletion of the ␣2-AMPK gene, phosphorylation of GS site 2 was decreased under basal conditions and unchanged by AICAR treatment. In contrast, in ␣1-AMPK knockout mice, the response to AICAR was normal. Fuel surplus (glucose loading) decreased AMPK activation by AICAR, but the phosphorylation of the downstream targets acetyl-CoA carboxylase-␤ and GS was normal. Fractionation studies suggest that this suppression of AMPK activation was not a direct consequence of AMPK association with membranes or glycogen, because AMPK was phosphorylated to a greater extent in response to AICAR in the membrane/glycogen fraction than in the cytosolic fraction. Thus, the downstream action of AMPK in response to AICAR was unaffected by glucose loading, whereas the action of the kinase upstream of AMPK, as judged by AMPK phosphorylation, was decreased. The fact that ␣2-AMPK is a GS kinase that inactivates GS while simultaneously activating glucose transport suggests that a balanced view on the suitability for AMPK as an antidiabetic drug target should be taken. Diabetes 53:3074 -3081, 2004 T he 5ЈAMP-activated protein kinase (AMPK) system is a sensor of cellular energy status that adjusts the supply of ATP to the demand for the nucleotide (1). Activation of ␣2-AMPK stimulates muscle glucose transport (2,3). Once glucose has been taken up and converted to glucose-6-phosphate (G6P), it can be stored as glycogen or metabolized by glycolysis to generate ATP. It has been reported that AMPK phosphorylates muscle glycogen synthase (GS) in cell-free assays at site 2 (Ser 7) (4). Thus, AMPK activation may under some conditions decrease the potential for glycogen synthesis. Recently, we and others showed that GS activity decreases in response to acute 5-aminoimidazole-1-␤-4-carboxamide ribofuranoside (AICAR) treatment of muscle-like cells in culture (5), isolated and perfused skeletal muscle (6 -8), and fast twitch, but not slow twitch, muscle in vivo (7). AICAR treatment leads to decreased gel mobility of GS in perfused muscle, which together with the decreased activity, is reversed by protein phosphatase treatment (6). These observations indicate that regulation of GS activity by AICAR involves phosphorylation of GS. Although it has been suggested that AICAR-induced GS deactivation is mediated by AMPK due to the negative correlation between ␣2-AMPK and GS activity (6), these data do not prove a causal relation. Thus, by studying muscle from ␣-AMPK knockout (KO) mice in the present study, we aimed to verify that AMPK is a muscle GS kinase in vivo.AMPK activity decreases when muscle is exposed to fuel surplus. For example, glucose loading and glycogen accumulation suppress muscle AMPK phosphorylation/ activation at basal co...

Dual Regulation of Muscle Glycogen Synthase during Exercise by Activation and Compartmentalization

“…2). This is in agreement with a previous study, which reported that GS phosphorylation at sites 3a and 2 is decreased in rested rat muscle with low glycogen levels (39,40). Altogether our results indicate that GS FIGURE 3.…”

Dysregulation of muscle glycogen synthase in recovery from exercise in type 2 diabetes

“…Glycogen is a proposed regulator of GS activity [30] and GS phosphorylation at sites 2+2a [28]. Consistent with this, the exercise-associated decrease in muscle glycogen content has been shown to be a factor involved in GS activation [30][31][32].…”

“…Glycogen synthase kinase 3 (GSK3) and AMP-activated protein kinase (AMPK) are major upstream regulators of GS [27,28]. Insulin stimulation inhibited GSK-3β activity by increasing Ser9 phosphorylation on both experimental days ( p<0.001).…”

Adenosine Monophosphate-Activated Protein Kinase: A Central Regulator of Metabolism with Roles in Diabetes, Cancer, and Viral Infection

“…The CBM in the AMPK-b subunit causes the complex to associate with glycogen particles in intact cells (Hudson et al 2003;Polekhina et al 2003), and AMPK also associates with the surface of glycogen particles in vivo (Bendayan et al 2009). The function of this association remains uncertain, although AMPK does phosphorylate two proteins that are targeted to glycogen, that is (1) muscle glycogen synthase (mGS), whose phosphorylation at site 2 causes inactivation Jorgensen et al 2004), and (2) R5/PTG, which targets the catalytic subunit of protein phosphatase-1 (PP1) to glycogen (Vernia et al 2009). Phosphorylation of R5 is proposed to promote its ubiquitylation and degradation, so that phosphorylation of mGS and R5 by AMPK would not only cause inactivation of mGS, but also dissociation from glycogen of the protein phosphatase (PP1) that reactivates it, the two effects cooperating to inhibit glycogen synthesis.…”