Insulin Control of Glycogen Metabolism in Knockout Mice Lacking the Muscle-Specific Protein Phosphatase PP1G/R_GL

Abstract

The regulatory-targeting subunit (R_GL, also called G_M) of the muscle-specific glycogen-associated protein phosphatase PP1G targets the enzyme to glycogen where it modulates the activity of glycogen-metabolizing enzymes. PP1G/R_GL has been postulated to play a central role in epinephrine and insulin control of glycogen metabolism via phosphorylation of R_GL. To investigate the function of the phosphatase, R_GLknockout mice were generated. Animals lacking R_GL show no obvious defects. The R_GL protein is absent from the skeletal and cardiac muscle of null mutants and present at ∼50% of the wild-type level in heterozygotes. Both the level and activity of C1 protein are also decreased by ∼50% in the R_GL-deficient mice. In skeletal muscle, the glycogen synthase (GS) activity ratio in the absence and presence of glucose-6-phosphate is reduced from 0.3 in the wild type to 0.1 in the null mutant R_GL mice, whereas the phosphorylase activity ratio in the absence and presence of AMP is increased from 0.4 to 0.7. Glycogen accumulation is decreased by ∼90%. Despite impaired glycogen accumulation in muscle, the animals remain normoglycemic. Glucose tolerance and insulin responsiveness are identical in wild-type and knockout mice, as are basal and insulin-stimulated glucose uptakes in skeletal muscle. Most importantly, insulin activated GS in both wild-type and R_GLnull mutant mice and stimulated a GS-specific protein phosphatase in both groups. These results demonstrate that R_GL is genetically linked to glycogen metabolism, since its loss decreases PP1 and basal GS activities and glycogen accumulation. However, PP1G/R_GL is not required for insulin activation of GS in skeletal muscle, and rather another GS-specific phosphatase appears to be involved.

ACKNOWLEDGMENTS

This work was supported in part by National Institutes of Health Grant DK36569 and by a research grant from the American Diabetes Association.

We thank Suzanne Baugh and Paula Ladd for their technical assistance in some of the work presented. We are indebted to David A. Williams and Cheryl Bock for generating the R_GL chimeric mice. We are grateful to Alain Baron (Amylin San Diego, Calif.) and Mark Heiman (Eli Lilly Co.) for performing glucose clamping and determining body composition and metabolic rates, respectively. We thank Alan Saltiel (Pfizer Global Research and Development), Patricia Cohen and Philip Cohen (University of Dundee) for their generous gifts of PTG, R6, and phospho-R_GL antibodies, respectively. We are particularly grateful to Peter J. Roach and Robert A. Harris for discussion of the work and for criticism of the manuscript.