SEQUENTIAL ACTIVATION OF 2 MITOGEN-ACTIVATED PROTEIN (MAP) KINASE ISOFORMS IN RAT SKELETAL-MUSCLE FOLLOWING INSULIN INJECTION

Skeletal muscle is a major target of insulin action. The possible role of MAP kinase activation in insulin receptor signalling in muscle was examined. After a 48-hr fast, rats were injected intravenously with insulin or saline, muscles were excised after 3-20 min, homogenized, and MAP kinases were partially purified by ammonium sulfate precipitation and Mono Q chromatography. Activity was assayed as 32P-incorporation into myelin basic protein. Two activity peaks were identified; peak I eluted with ∼0.1 M NaCl and peak II with ∼0.2 M NaCl. Three min after insulin injection the activity of peak II increased >2-fold, peak I was unchanged. After 10 min, the activity of peak II returned toward baseline, while peak I was activated ∼3-fold. Immunoblots confirmed the presence of MAP kinases eluting with activity peaks I and II; the former as a ∼41 kDa protein and the latter as a doublet of ∼42 and ∼44 kDa. The data suggest sequential activation of two MAP kinases in muscle; the isoform which activates/deactivates rapidly may represent ERK-1, while the more slowly responding isoform may be ERK-2. © 1993 Academic Press. All rights reserved.