Phosphorylation state of the glucose transporter from 3T3-L1 adipocytes: effect of insulin and phorbol ester
Polyclonal antibodies against the purified human erythrocyte glucose transporter (GT) were used to study the phosphorylation state of GT in (/sup 32/P)orthophosphate-labeled 3T3-L1 adipocytes that were exposed to insulin or phorbol ester. Conditions were established in which the recovery of GT (identified as a polypeptide of M/sub r/ 51,000) after immunoprecipitation from detergent-solubilized adipocytes was about 50% of total cellular transporter, as quantitated by immunoblot analysis. Exposure of adipocytes to insulin (100 nM) for 10 min after prelabeling in /sup 32/P for 90 min, followed by the addition of phorbol myristate acetate (PMA; 1 ..mu..M) for 20 min elicited a marked phosphorylation of GT. Addition of excess purified human erythrocyte GT completely abolished the immunoprecipitation of the 51 K phosphoprotein; this finding validates the conclusion that this phosphoprotein is GT. Treatment with PMA alone resulted in only 30% of the incorporation of /sup 32/P into the 51 K region of the gel compared to that seen with the combination of PMA and insulin. Insulin alone gave only about 20% /sup 32/P incorporation into this region compared to the combination treatment. It remains to be determined if the phosphorylation into the 51 K region of the gel seen after treatment with either of the two agonists alone is into GT. The authors tentative hypothesis is that GT is not phosphorylated in basal cells, and that insulin causes little or no increase in the phosphorylation state. On the other hand, PMA elicits some phosphorylation of GT that can be increased about 3-fold by prior treatment with insulin. Presumably, this increase is due to the translocation of GT to the plasma membrane where it is a better substrate for activated protein kinase C.
- Research Organization:
- Dartmouth Medical School, Hanover, NH
- OSTI ID:
- 5230538
- Report Number(s):
- CONF-8606151-; TRN: 86-031629
- Journal Information:
- Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Vol. 45:6; Conference: 76. annual meeting of the Federation of American Society for Experimental Biology, Washington, DC, USA, 8 Jun 1986
- Country of Publication:
- United States
- Language:
- English
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59 BASIC BIOLOGICAL SCIENCES
GLUCOSE
MEMBRANE TRANSPORT
INSULIN
BIOLOGICAL EFFECTS
PHORBOL ESTERS
POLYPEPTIDES
PHOSPHORYLATION
AUTORADIOGRAPHY
CELL MEMBRANES
ERYTHROCYTES
FIBROBLASTS
MAN
PHOSPHORUS 32
PHOSPHOTRANSFERASES
SYNERGISM
ALDEHYDES
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BETA-MINUS DECAY RADIOISOTOPES
BIOLOGICAL MATERIALS
BLOOD
BLOOD CELLS
BODY FLUIDS
CARBOHYDRATES
CARCINOGENS
CELL CONSTITUENTS
CHEMICAL REACTIONS
CONNECTIVE TISSUE CELLS
DAYS LIVING RADIOISOTOPES
ENZYMES
ESTERS
HEXOSES
HORMONES
ISOTOPES
LIGHT NUCLEI
MAMMALS
MATERIALS
MEMBRANES
MONOSACCHARIDES
NUCLEI
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
PEPTIDE HORMONES
PEPTIDES
PHOSPHORUS ISOTOPES
PHOSPHORUS-GROUP TRANSFERASES
PRIMATES
PROTEINS
RADIOISOTOPES
SACCHARIDES
SOMATIC CELLS
TRANSFERASES
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550201 - Biochemistry- Tracer Techniques