Nutrient regulation of PKCepsilon is mediated by leucine, not insulin, in skeletal muscle.
Nutrients enhance signaling pathways involved in skeletal muscle growth through an increased rate of protein synthesis. These studies have led to an understanding of the potential role of the mammalian target of rapamycin (mTOR) in this process. However, activation of mTOR cannot account for all the stimulatory effects of nutrients. The purpose of these experiments was to examine the effect of nutrients on the cellular distribution and activation state of novel PKC isoforms (PKCepsilon and PKCdelta) in the gastrocnemius of rats by use of modification state-dependent phosphopeptide-specific antibodies. The phosphorylation of PKCepsilon on the catalytic domain autophosphorylation site (Ser(729)) was elevated during feeding and then returned to basal levels when the feeding period ended. Meal feeding augmented the phosphorylation of the downstream effectors of mTOR, namely S6K1 and 4E-BP1. In contrast, the phosphorylation of PKCdelta on either the catalytic domain autophosphorylation site (Ser(643)) or activation loop site (Thr(505)) was unaffected. Similar results were obtained when animals were given leucine either acutely via gavage or chronically by dietary supplementations. The effect of leucine was not mimicked by injecting animals with insulin but could be induced by gavage with norleucine, a structural analog of leucine that does not increase plasma insulin concentration. Thus rises in insulin secondary to meal intake or leucine gavage are probably not responsible for increased phosphorylation of PKCepsilon in response to meal feeding. Elevating the leucine concentration stimulated the phosphorylation of PKCepsilon in gastrocnemius from perfused hindlimb and caused a shift in the distribution of PKCepsilon from the membrane fraction to the cytosolic fraction. The results indicate that leucine leads to an activation (autophosphorylation) and subcellular redistribution of PKCepsilon, but not PKCdelta, in gastrocnemius both in vivo and in vitro. Furthermore, activation of the mTOR signaling pathway above basal conditions does not appear to be necessary to induce phosphorylation or translocation of PKCepsilon, suggesting that multiple signaling pathways become activated with leucine.
Vary
Thomas C
TC
Dept. of Cellular and Molecular Physiology, Pennsylvania State University College of Medicine, Hershey, PA 17033, USA. tvary@psu.edu
Goodman
Stacy
S
Kilpatrick
Laurie E
LE
Lynch
Christopher J
CJ
D000824:Animal Nutritional Physiological Phenomena; D000818:Animals; D004305:Dose-Response Relationship, Drug; D007328:Insulin / Q000378:metabolism* / Q000494:pharmacology*; D007930:Leucine / Q000008:administration & dosage* / Q000378:metabolism*; D008297:Male; D018482:Muscle, Skeletal / Q000187:drug effects / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism*; D051744:Protein Kinase C-epsilon; D011494:Protein Kinases / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D058570:TOR Serine-Threonine Kinases
D007328:Insulin; D011494:Protein Kinases; C497257:Prkce protein, rat; C546845:mTOR protein, rat; D058570:TOR Serine-Threonine Kinases; D011493:Protein Kinase C; D051744:Protein Kinase C-epsilon; D007930:Leucine