Two-dimensional electrophoretic analyses of proteins synthesized during differentiation of 3T3-L1 preadipocytes

Abstract

The changes in protein composition and cell surface proteins that occur during the adipocyte conversion of 3T3-L1 preadipocytes were monitored by two-dimensional polyacrylamide gel electrophoresis folowing incubation of cells with [35S]methionine for periods of 3 and 24 h. Alterations in the biosynthesis of more than 30 cytoplasmic proteins, 9 non-histone, chromosome-associated proteins, and 24 membrane proteins, were detected. Although the methodological limitations of the electrophoretic systems employed result in an underestimate of the total number of differences, the alterations observed exceed the enzyme changes known to occur during differentiation of these cells. One major alteration occurring during differentiation is a decrease in the content of a protein whose position following two-dimensional electrophoresis tentatively identified it as actin. A fall in actin content accompanying adipocyte conversion was confirmed by direct analysis of the DNase 1 inhibitory activity in homogenates prepared from cells during the course of differentiation. Studies of cell surface proteins by lactoperoxidase-catalyzed iodination reveal a number of changes during differentiation including an increase in a polypeptide(s) in the molecular weight range of 16,500 to 18,500, a decrease in at least four proteins of molecular weights greater than 100,000, and in a protein of molecular weight 95,000.