Control of peptide-chain initiation in rat skeletal muscle. Development of methods for preparation of native ribosomal subunits and analysis of the effect of insulin on formation of 40 S initiation complexes.
A method was developed for isolation of native ribosomal subunits from rat gastrocnemius muscle. Native 40 S subunits which were isolated by this method retained their associated nonribosomal proteins and consisted primarily of particles with equilibrium densities of 1.41 and 1.48 g/cm3. Based on the binding of radiolabeled Met-tRNAmeti, the 1.41 g/cm3 particle was identified as the 40 S initiation complex. Insulin deficiency in vivo resulting from either diabetes or fasting led to a 2-fold increase in 75 S monomers but had no effect on the numbers of native 40 and 60 S subunits or the relative distribution of the 1.41 and 1.48 g/cm3 particles. The rate of protein synthesis in perfused muscle preparations derived from insulin-deficient rats was reduced to about half the control value. Addition of insulin to the perfusate restored protein synthesis and 75 S monomers to control levels. The effect of insulin on protein synthesis was associated with a 1.5-fold increase in the amount of Met-tRNAmeti bound to the 1.41 g/cm3 particle. These findings identify formation of 40 S initiation complexes as a site of action of insulin on protein synthesis in skeletal muscle.
Kelly
F J
FJ
Jefferson
L S
LS
D000818:Animals; D002499:Centrifugation, Density Gradient; D003921:Diabetes Mellitus, Experimental / Q000378:metabolism; D007328:Insulin / Q000494:pharmacology*; D008297:Male; D008715:Methionine / Q000378:metabolism; D009132:Muscles / Q000378:metabolism*; D010442:Peptide Chain Initiation, Translational / Q000187:drug effects*; D012313:RNA / Q000032:analysis; D051381:Rats; D011919:Rats, Inbred Strains; D012270:Ribosomes / Q000378:metabolism*
D007328:Insulin; D012313:RNA; D008715:Methionine