Opposite Transcriptional Regulation in Skeletal Muscle of AMP-activated Protein Kinase γ3 R225Q Transgenic Versus Knock-out Mice

Abstract

AMP-activated protein kinase (AMPK) is an evolutionarily conserved heterotrimer important for metabolic sensing in all eukaryotes. The muscle-specific isoform of the regulatory ␥-subunit of the kinase, AMPK ␥3, has an important role in glucose uptake, glycogen synthesis, and fat oxidation in white skeletal muscle, as previously demonstrated by physiological characterization of AMPK ␥3 mutant (R225Q) transgenic (TgPrkag3 225Q ) and ␥3 knock-out (Prkag3 ؊/؊ ) mice. We determined AMPK ␥3-dependent regulation of gene expression by analyzing global transcription profiles in glycolytic skeletal muscle from ␥3 mutant transgenic and knock-out mice using oligonucleotide microarray technology. Evidence is provided for coordinated and reciprocal regulation of multiple key components in glucose and fat metabolism, as well as skeletal muscle ergogenics in TgPrkag3 225Q and Prkag3 ؊/؊ mice. The differential gene expression profile was consistent with the physiological differences between the models, providing a molecular mechanism for the observed phenotype. The striking pattern of opposing transcriptional changes between TgPrkag3 225Q and Prkag3 ؊/؊ mice identifies differentially expressed targets being truly regulated by AMPK and is consistent with the view that R225Q is an activating mutation, in terms of its downstream effects. Additionally, we identified a wide array of novel targets and regulatory pathways for AMPK in skeletal muscle.AMP-activated protein kinase (AMPK) 2 is a critical regulator of carbohydrate and fat metabolism in eukaryotic cells (reviewed in Refs. 1 and 2). AMPK is a heterotrimer that consists of ␣-, ␤-, and ␥-subunits, all of which are required for its activity. The catalytic ␣-subunit contains a conventional serine/threonine protein kinase domain, and phosphorylation of Thr-172 residue within the activation loop of the ␣-subunit by upstream kinases is essential for the activity of the heterotrimer (3-6). Once phosphorylated at Thr-172, AMPK can be further activated by allosteric binding of AMP to the evolutionary conserved cystathionine ␤-synthase domains in the regulatory ␥-subunit (7). The AMPK ␤-subunit acts as a scaffold for binding of the ␣-and ␥-subunits (8). The ␤-subunit also contains a glycogen-binding domain, and recent findings provide evidence that this motif is involved in targeting the AMPK complex to cellular glycogen stores (9, 10). The mammalian genome contains seven AMPK genes encoding for two ␣-, two ␤-, and three ␥-isoforms. Thus, there are 12 possible combinations of heterotrimeric AMPK, and the physiological function of the AMPK holoenzyme depends on the particular isoforms present in the complex.We have provided evidence that AMPK ␥3 is the predominant ␥-isoform expressed in glycolytic (white, fast-twitch type II) skeletal muscle (11). In contrast, it is expressed at low levels in oxidative (red, slowtwitch type I) skeletal muscle and is undetectable in brain, liver, heart, or white adipose tissue (11). Thus, the AMPK ␥3-subunit is the only isoform exhibiting tissue-specific ...

Gender difference of androgen actions on skeletal muscle transcriptome

“…In particular, the present study showed the higher expression of protein kinase AMP-activated g 3 non-catalytic subunit (Prkag3) and tripartite motifcontaining 63 (Trim63) in female mice. The Prkag3 increases fatty acid oxidation, glucose uptake, glycolysis, and glycogen synthesis (Nilsson et al 2006), most of which were concordant with the gene expression profile of energy metabolism in female mice seen in the present study. The Trim63 is known to induce muscle wasting (Glass 2005).…”

Role of AMP-activated protein kinase gamma 3 genetic variability in glucose and lipid metabolism in non-diabetic whites

“…SREBP1 positively regulates expression of the genes encoding HMG-CoA reductase and HMG-CoA synthetase; it also induces transcription of the LDL receptor gene in many different tissues [35]. Interestingly, SREBP1 (also known as sterol regulatory element binding protein-1 [SREBF1]) transcription is significantly upregulated in Prkag3 −/− knockout mice and downregulated in PRKAG3 R225Q mice [36]. On the other hand, PRKAG3-associated activity in skeletal muscle may also contribute to changes in LDLcholesterol serum levels.…”

“…On the other hand, PRKAG3-associated activity in skeletal muscle may also contribute to changes in LDLcholesterol serum levels. Beyond SREBP1 transcription [36], PRKAG3 regulates gene expression of various glucose-and lipid-metabolic key enzymes in white skeletal muscle [37] and therefore may have an effect on endogenous skeletal muscle cholesterol synthesis and LDL receptor expression. Such changes in cholesterol synthesis or LDL receptor expression of myocytes could alter their LDL-cholesterol ingestion, subsequently also altering serum LDL-cholesterol levels, as skeletal muscle tissue accounts for ∼5% of total peripheral LDL-cholesterol uptake [38,39].…”

Fatty acid-inducible ANGPTL4 governs lipid metabolic response to exercise

“…In vivo overexpression of an activating mutant of the muscle-specific isoform of the AMPKγ subunit supported the suppressive effect of AMPK on Angptl4 gene expression ( Fig. 5L) (22). Conversely, in vivo overexpression of a dominantnegative mutant of the AMPKα2 subunit led to a significant induction of Angptl4 mRNA (Fig.…”