A protein of molecular weight 31,000 became labeled with 32P within 5 min after addition of insulin to differentiated 3T3-L1 preadipocytes previously incubated for 55 min with 32Pi. The effect was mimicked by antisera directed against the insulin receptor and was eliminated by anti-insulin antiserum. Incorporation of 32P into this protein was more than 20-fold greater in insulin-treated cells than in cells not exposed to the hormone. At concentrations greater than required with insulin, epidermal growth factor and serum (1-5%) also stimulated phosphorylation whereas 1-isoproterenol, a beta-adrenergic agonist that increases intracellular accumulation of cyclic AMP, was without effect. The 31,000-dalton protein has been tentatively identified as ribosomal protein S6 by two-dimensional polyacrylamide gel electrophoresis. Incorporation of 32P into S6 could be detected within the same time period (5 min) and at the same insulin concentrations (0.1-1.0 nM) as are required to stimulate hexose uptake in both 3T3-L1 cells and mature mammalian adipocytes. The mechanism by which this phosphorylation either mediates or reflects the intracellular actions of insulin remains to be elucidated.

Smith, C. J., Wejksnora, P. J., Warner, J. R., Rubin, C. S., & Rosen, O. M. (1979). Insulin-stimulated protein phosphorylation in 3T3-L1 preadipocytes. Proceedings of the National Academy of Sciences, 76(6), 2725–2729.