Phosphorylation of adipose triglyceride lipase Ser⁴⁰⁴is not related to 5′-AMPK activation during moderate-intensity exercise in humans

Abstract

Mason RR, Meex RC, Lee-Young R, Canny BJ, Watt MJ. Phosphorylation of adipose triglyceride lipase Ser 404 is not related to 5=-AMPK activation during moderate-intensity exercise in humans. Am J Physiol Endocrinol Metab 303: E534 -E541, 2012. First published June 19, 2012 doi:10.1152/ajpendo.00082.2012.-Intramyocellular triacylglycerol provides fatty acid substrate for ATP generation in contracting muscle. The protein adipose triglyceride lipase (ATGL) is a key regulator of triacylglycerol lipolysis and whole body energy metabolism at rest and during exercise, and ATGL activity is reported to be enhanced by 5=-AMP-activated protein kinase (AMPK)-mediated phosphorylation at Ser 406 in mice. This is a curious observation, because AMPK activation reduces lipolysis in several cell types. We investigated whether the phosphorylation of ATGL Ser 404 (corresponding to murine Ser 406 ) was increased during exercise in human skeletal muscle and with pharmacological AMPK activation in myotubes in vitro. In human experiments, skeletal muscle and venous blood samples were obtained from recreationally active male subjects before and at 5 and 60 min during exercise. ATGL Ser 404 phosphorylation was not increased from rest during exercise, but ATGL Ser 404 phosphorylation correlated with myosin heavy chain 1 expression, suggesting a possible fiber type dependency. ATGL Ser 404 phosphorylation was not related to increases in AMPK activity, and immunoprecipitation experiments indicated no interaction between AMPK and ATGL. Rather, ATGL Ser 404 phosphorylation was associated with protein kinase A (PKA) signaling. ATGL Ser 406 phosphorylation in C2C12 myotubes was unaffected by 5-aminoimidazole-4-carboxaminde-1-␤-D-ribofuranoside, an AMPK activator, and the PKA activator forskolin. Our results demonstrate that ATGL Ser 404 phosphorylation is not increased in mixed skeletal muscle during moderateintensity exercise and that AMPK does not appear to be an activating kinase for ATGL Ser 404/406 in skeletal muscle.5=-adenosine monophosphate-activated protein kinase; skeletal muscle; lipolysis; fatty acid metabolism LIPOLYSIS IS A HIGHLY CONSERVED PROCESS that involves the sequential removal of fatty acids from triacylglycerol to supply substrate for ATP production. Free fatty acids (FFA) derived from adipose tissue provide the majority of fatty acid substrate in humans, while energy derived from muscle triacylglycerol lipolysis constitutes an important fuel source, particularly during prolonged moderate intensity exercise (37, 41). Lipolysis is controlled by the actions of several lipases. Adipose triglyceride lipase (ATGL) is the predominant triacylglycerol lipase and is critical for efficient triacylglycerol lipolysis in all tissues assessed to date (2,14,29,30,36), including skeletal muscle (5, 16). Whole body deletion of ATGL in mice impairs the FFA supply required to sustain normal substrate metabolism, resulting in increased carbohydrate metabolism and impaired exercise performance (14, 16). In humans, mutations in both alleles of ...

Dissecting adipose tissue lipolysis: molecular regulation and implications for metabolic disease

“…Additionally, in mouse AT, Ser 406 phosphorylation was increased with fasting, exercise, and ex vivo stimulation of the cAMP-dependent pathway in a PKA-dependent manner, thereby implicating this kinase in the phosphorylation of ATGL (Pagnon et al 2012). Notably, in human skeletal muscle (SM), Ser 404 phosphorylation is also associated with PKA signaling and not with AMPK signaling, indicating that PKA is indeed the upstream kinase for this site (Mason et al 2012). Whether PKA and/or AMPK are responsible for phosphorylation of Ser 430 is currently unknown, and so far no reports have been published on the functional role of this site.…”

Biochemistry and pathophysiology of intravascular and intracellular lipolysis

“…Phosphorylation of Ser 406 increases the catalytic activity of ATGL in hormone-stimulated lipolysis; however, the protein kinase responsible for enzyme phosphorylation is controversial. Watt and colleagues (Mason et al 2012;Pagnon et al 2012) reported that PKA phosphorylates Ser 406, but Ahmadian et al (2012) found that AMPkinase was responsible for this modification.…”

Adipose triglyceride lipase deletion from adipocytes, but not skeletal myocytes, impairs acute exercise performance in mice

“…Furthermore, the higher IMTG content in SMAKO mice would provide more substrate for HSL-mediated lipolysis, as suggested by the dramatic reduction in IMTG content in exercised SMAKO mice. Indeed, both ATGL and HSL are activated by similar pathways during muscle contraction (43). This would explain our observation of appropriate whole body fat oxidation and a tendency for greater relative increases in fat oxidation with skeletal myocyte-specific ATGL deletion.…”