Regulation of caspase-3 activity by insulin in skeletal muscle cells involves both PI3-kinase and MEK-1/2.
A hallmark of skeletal muscle atrophy is increased activities of several proteolytic systems, including caspase-3. We have previously shown that conditions involving insulin deficiency or insulin resistance increase both overall protein degradation and caspase-3-mediated actin cleavage. In the present experiments, we examined how insulin regulates caspase-3 activity in L6 myotubes. Reducing the serum concentration in the culture media from 2 to 0.5% overnight increased caspase-3 activity and actin cleavage. Addition of insulin to proteolytically active cells attenuated both responses within 4 h. Individually, inhibitors of either phosphatidylinositide 3-kinase (PI3K) or MEK1/2 partially blocked the insulin-induced reduction in caspase-3 activity; in combination, the inhibitors completely prevented insulin from attenuating caspase-3 activity. Insulin suppressed caspase-3 activity by a complex mechanism that included direct inhibition due to an increased interaction between caspase-3 and cellular inhibitor of apoptosis-1 and indirect inhibition via phosphorylation (i.e., inactivation) of the proapoptotic protein Bad, which participates in the intrinsic (i.e., mitochondrial) apoptosis activation cascade. Unlike other cell types, the phosphorylation of Bad Ser112 was mediated by the PI3K/Akt pathway rather than the MEK/ERK/ribosomal S6 protein kinase pathway. In summary, our findings indicate that insulin regulates caspase-3 activity by a multistep process that is unique to skeletal muscle, thus providing insights about the muscle-specific nature of the atrophy process.
Gao
Yongmei
Y
Renal Division, Rm. 338, Woodruff Memorial Research Bldg., 1639 Pierce Dr., Emory Univ., Atlanta, GA 30322, USA.
Ordas
Ronald
R
Klein
Janet D
JD
Price
S Russ
SR
D000199:Actins / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism*; D002478:Cells, Cultured; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D047468:Immunoprecipitation; D007328:Insulin / Q000494:pharmacology*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D018485:Muscle Fibers, Skeletal / Q000187:drug effects* / Q000201:enzymology*; D018482:Muscle, Skeletal / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology*; D051058:Oncogene Protein v-akt / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects; D051025:bcl-Associated Death Protein / Q000378:metabolism
D000199:Actins; D007328:Insulin; D051025:bcl-Associated Death Protein; D019869:Phosphatidylinositol 3-Kinases; D051058:Oncogene Protein v-akt; D048049:Extracellular Signal-Regulated MAP Kinases; D020928:Mitogen-Activated Protein Kinases; D053148:Caspase 3