The abundance and the subcellular distribution of GTP‐binding proteins was studied in membrane fractions (plasma membranes and low‐density microsomes) from 3T3‐L1 cells before and after differentiation to the insulin‐sensitive phenotype. After differentiation, the abundance of αi (α subunit of GTP‐binding protein Gi), αi (α subunit of GTP‐binding protein Gi), and of a 47‐kDa αs, (α subunit of GTP‐binding protein Gs) as detected by immunoblotting with specific antisera was reduced by 10–50% when normalized per membrane protein. In contrast, a 43‐kDa αs was increased about threefold after differentiation. Furthermore, cholera‐toxin‐catalyzed ADP‐ribosylation of both 43‐kDa and 47‐kDa αs was disproportionately increased ninefold and threefold, respectively, possibly reflecting the increased production of an ADP‐ribosylation factor in the differentiated cells. The small GTP‐binding protein Ha‐ras was reduced by 50%, whereas rab1 and other small GTP‐binding proteins tentatively identified as rab‐isoforms (ras‐homologous gene products from brain) were increased by 100% and 70%, respectively. Since the total protein content of 3T3‐L1 cells was increased threefold after differentiation, the observed increase of the 43‐kDa αs, rab1 and of the other rab isoforms was eightfold, sixfold and fivefold, respectively, when normalized/cell count. With the exception of the rab isoforms, all GTP‐binding proteins were predominantly, if not exclusively, located in the plasma membrane; comparable amounts of the rab isoforms were found in plasma membranes and low‐density microsomes. Insulin induced the characteristic redistribution of glucose transporters GLUT4 from low‐density microsomes to the plasma membranes, but failed to alter the subcellular distribution of any of the GTP‐binding proteins investigated. These data suggest that the increase in the abundance of the 43‐kDa αs subunit and of several rab isoforms might be related to specific functions of the adipocyte‐like phenotype, but that none of the investigated guanine‐nucleotide‐binding regulatory (G)‐proteins appears to be tightly associated with the GLUT4. Copyright © 1993, Wiley Blackwell. All rights reserved
CITATION STYLE
HUPPERTZ, C., SCHÜRMANN, A., & JOOST, H. ‐G. (1993). Abundance and subcellular distribution of GTP‐binding proteins in 3T3‐L1 cells before and after differentiation to the insulin‐sensitive phenotype. European Journal of Biochemistry, 215(3), 611–617. https://doi.org/10.1111/j.1432-1033.1993.tb18072.x
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