Reduced glycogen availability is associated with increased AMPKα2 activity, nuclear AMPKα2 protein abundance, and GLUT4 mRNA expression in contracting human skeletal muscle

Glycogen availability can influence glucose transporter 4 (GLUT4) expression in skeletal muscle through unknown mechanisms. The multisubstrate enzyme AMP-activated protein kinase (AMPK) has also been shown to play an important role in the regulation of GLUT4 expression in skeletal muscle. During contraction, AMPK [alpha]2 translocates to the nucleus and the activity of this AMPK isoform is enhanced when skeletal muscle glycogen is low. In this study, we investigated if decreased pre-exercise muscle glycogen levels and increased AMPK [alpha]2 activity reduced the association of AMPK with glycogen and increased AMPK [alpha]2 translocation to the nucleus and GLUT4 mRNA expression following exercise. Seven males performed 60 min of exercise at ~70% [VO.sub.2] peak on 2 occasions: either with normal (control) or low (LG) carbohydrate pre-exercise muscle glycogen content. Muscle samples were obtained by needle biopsy before and after exercise. Low muscle glycogen was associated with elevated AMPK [alpha]2 activity and acetyl-CoA carboxylase [beta] phosphorylation, increased translocation of AMPK [alpha]2 to the nucleus, and increased GLUT4 mRNA. Transfection of primary human myotubes with a constitutively active AMPK adenovirus also stimulated GLUT4 mRNA, providing direct evidence of a role of AMPK in regulating GLUT4 expression. We suggest that increased activation of AMPK [alpha]2 under conditions of low muscle glycogen enhances AMPK [alpha]2 nuclear translocation and increases GLUT4 mRNA expression in response to exercise in human skeletal muscle.

Comparative structural analyses of purified glycogen particles from rat liver, human skeletal muscle and commercial preparations

Glycogen is a cellular energy store that is crucial for whole body energy metabolism, metabolic regulation and exercise performance. To understand glycogen structure we have purified glycogen particles from rat liver and human skeletal muscle tissues and compared their biophysical properties with those found in commercial glycogen preparations. Ultrastructural analysis of commercial liver glycogens fails to reveal the classical α-rosette structure but small irregularly shaped particles. In contrast, commercial slipper limpet glycogen consists of β-particles with similar branching and chain lengths to purified rat liver glycogen together with a tendency to form small α-particles, and suggest it should be used as a source of glycogen for all future studies requiring a substitute for mammalian liver glycogen.

Exercise-induced histone modifications in human skeletal muscle

Skeletal muscle adaptations to exercise confer many of the health benefits of physical activity and occur partly through alterations in skeletal muscle gene expression. The exact mechanisms mediating altered skeletal muscle gene expression in response to exercise are unknown. However, in recent years, chromatin remodelling through epigenetic histone modifications has emerged as a key regulatory mechanism controlling gene expression in general. The purpose of this study was to examine the effect of exercise on global histone modifications that mediate chromatin remodelling and transcriptional activation in human skeletal muscle in response to exercise. In addition, we sought to examine the signalling mechanisms regulating these processes. Following 60 min of cycling, global histone 3 acetylation at lysine 9 and 14, a modification associated with transcriptional initiation, was unchanged from basal levels, but was increased at lysine 36, a site associated with transcriptional elongation. We examined the regulation of the class IIa histone deacetylases (HDACs), which are enzymes that suppress histone acetylation and have been implicated in the adaptations to exercise. While we found no evidence of proteasomal degradation of the class IIa HDACs, we found that HDAC4 and 5 were exported from the nucleus during exercise, thereby removing their transcriptional repressive function. We also observed activation of the AMP-activated protein kinase (AMPK) and the calcium–calmodulin-dependent protein kinase II (CaMKII) in response to exercise, which are two kinases that induce phosphorylation-dependent class IIa HDAC nuclear export. These data delineate a signalling pathway that might mediate skeletal muscle adaptations in response to exercise.

AMPK-mediated transcription in skeletal muscle

Skeletal muscle phenotype plays a critical role in human performance and health, and skeletal muscle oxidative capacity is a key determinant of exercise tolerance. More recently, defective muscle oxidative metabolism has been implicated in a number of conditions associated with the metabolic syndrome, cardiovascular disease and muscle-wasting disorders. AMPK (AMP-activated protein kinase) is a critical regulator of cellular and organismal energy balance. AMPK has also emerged as a key regulator of skeletal muscle oxidative function, including metabolic enzyme expression, mitochondrial biogenesis and angiogenesis. AMPK mediates these processes primarily through alterations in gene expression. The present review examines the role of AMPK in skeletal muscle transcription and provides an overview of the known transcriptional substrates mediating the effects of AMPK on skeletal muscle phenotype.

Histone modifications and skeletal muscle metabolic gene expression

1.      Skeletal muscle oxidative function and metabolic gene expression are co-ordinately downregulated in metabolic diseases such as insulin resistance, obesity and Type 2 diabetes. Altering skeletal muscle metabolic gene expression to favour enhanced energy expenditure is considered a potential therapy to combat these diseases.

2.      Histone deacetylases (HDACs) are chromatin-remodelling enzymes that repress gene expression. It has been shown that HDAC4 and 5 co-operatively regulate a number of genes involved in various aspects of metabolism. Understanding how HDACs are regulated provides insights into the mechanisms regulating skeletal muscle metabolic gene expression.

3.      Multiple kinases control phosphorylation-dependent nuclear export of HDACs, rendering them unable to repress transcription. We have found a major role for the AMP-activated protein kinase (AMPK) in response to energetic stress, yet metabolic gene expression is maintained in the absence of AMPK activity. Preliminary evidence suggests a potential role for protein kinase D, also a Class IIa HDAC kinase, in this response.

4.      The HDACs are also regulated by ubiquitin-mediated proteasomal degradation, although the exact mediators of this process have not been identified.

5.      Because HDACs appear to be critical regulators of skeletal muscle metabolic gene expression, HDAC inhibition could be an effective therapy to treat metabolic diseases.

6.      Together, these data show that HDAC4 and 5 are critical regulators of metabolic gene expression and that understanding their regulation could provide a number of points of intervention for therapies designed to treat metabolic diseases, such as insulin resistance, obesity and Type 2 diabetes.

Skeletal muscle GLUT-4 gene and protein regulation

Twenty two, young, healthy individuals participated in three studies aiming to assess the effect of various types of physical activity - acute exercise of moderate intensity and duration, varying intensity, short-term training - on skeletal muscle GLUT-4 gene and protein expression as well as on a range of genes encoding the proteins involved in carbohydrate metabolism in skeletal muscle.

Differential gene expression in the skeletal muscle of Psammomys obesis

Focuses on discovering and investigating altered gene expression in the skeletal muscle of Psammomys obesus which is a unique model of obesity and Type II diabetes in which its development is similar to that of the human population. Defects in the skeletal muscle are pivotal to the development of Type II diabetes. Using the latest techniques in molecular biology the regulation of a number of genes was confirmed to be altered in obese or diabetic animals compared to lean. This indicates that changes to gene expression contribute to the metabolic disturbances associated with obesity and Type II diabetes.

Effect of exercise and protein on skeletal muscle inflammatory mediators

Intense exercise results in muscular inflammation. Molecular techniques were used to identify novel inflammatory proteins in human muscle. Males and females displayed different levels of exercise-induced inflammatory proteins. Interestingly, dairy protein supplements reduced these inflammatory proteins post-exercise. Increased dietary red meat consumption, with training, had no impact on muscle inflammation, although strength gain was improved.

Creatine transporter expression in skeletal muscle

Creatine is an important molecule involved in providing energy to the body. Its major stores are in skeletal muscle. The creatine transporter protein (CreaT) mRNA is believed to be responsible for the uptake of the majority of creatine in skeletal muscle. This thesis examined factors that might have affected the expression of the creatine transporter in skeletal muscle.

Regulation of fat oxidative genes in human skeletal muscle

The ability of skeletal muscle to adapt fat oxidation rates is important for human health. Lipid metabolism requires the involvement of many proteins encoded by their corresponding genes. This thesis demonstrates that manipulating plasma free fatty acid levels alters the expression of selected genes involved in regulating fatty acid metabolism.

PKC and CaMK in human skeletal muscle : effects of exercise

The effects of exercise on novel signalling enzymes in skeletal muscle of humans was investigated. It was shown that exercise increased the activity of a calcium and calmodulin activated kinase. High-intensity, but submaximal, exercise increased the activity of some but not all isoforms of protein kinase C, a lipid-activated kinase family. These findings suggest that these enzymes may be part of the signalling process leading to beneficial adaptation to repeated exercise as well as the control of function within skeletal muscle during exercise.

Factors regulating skeletal muscle fatty acid oxidative genes

Skeletal muscle is the most significant site for whole body fat utilisation. The ability to regulate fat use has a significant impact on the development of obesity and Type II diabetes. The studies conducted during this PhD provided significant insight into the complex molecular regulation of skeletal muscle fat utilisation.

Regulation of GLUT4 expression in human skeletal muscle

Increasing the number of glucose transporters in muscle ameliorates many of the symptoms associated with type 2 diabetes. This thesis identifies mechanisms regulating glucose transporter gene expression, and therefore glucose transporter number, in human skeletal muscle and provides potential targets for the treatment and management of type 2 diabetes.