Involvement of a phosphorylation-mediated pathway to regulate the function of NSPL1 in exercise

Skeletal-type neuroendocrine-specific protein like 1 (sk-NSPL1) has been demonstrated to be physiologically important in regulating the membrane translocation of glucose transporter 4 (GLUT4) in skeletal muscles. We investigated the levels of phosphorylation in proteins that are thought to be involved in exercise in wild-type and sk-NSPL1-deficient muscles with specific antibodies and phosphate-metal affinity chromatography resin (p-resin). In both normal skeletal muscle and sk-NSPL1-deficient muscle, adenosine monophosphate (AMP)-dependent kinase (AMPK) and acetyl-CoA carboxylase (ACC) were phosphorylated and adsorbed onto p-resin at high levels after exercise. On the other hand, the effect of 5-aminoimidazole-4-carboxyamide ribonucleoside (AICAR), which is an activator of AMPK, in blood glucose was greatly diminished in mutant mice. P-resin adsorbed sk-NSPL1 in the membrane fraction from wild-type muscle after exercise and AICAR administration. Isolated sk-NSPL1 from wild-type also had increased adsorption onto p-resin after treatment with Ca(2+) and adenosine triphosphate (ATP). After long-term incubation of sk-NSPL1-containing membrane without ATP, sk-NSPL1 adsorption onto anion-exchange resin was drastically reduced. These results suggest that the function of sk-NSPL1 is regulated by a [Ca(2+)](i)- and AMPK-mediated pathway under exercise, and support the hypothesis that sk-NSPL1 is an important factor in the downstream of the exercise-dependent pathway in GLUT4 translocation.