Program in Molecular Medicine; Department of Cell Biology
3T3 Cells; Adaptor Protein Complex alpha Subunits; Adaptor Protein Complex gamma Subunits; Adaptor Proteins, Vesicular Transport; Adipocytes; Animals; Cell Compartmentation; *Cell Differentiation; *Clathrin; Coated Pits, Cell-Membrane; Cytosol; Fibroblasts; Fluorescent Antibody Technique; Golgi Apparatus; Intracellular Membranes; Mice; Proteins; Subcellular Fractions
Cell Biology | Molecular Biology
Mechanisms for intracellular retention of proteins are induced during adipocytic differentiation of 3T3-L1 cells. To investigate the potential role of clathrin lattices in these retention processes, we performed a morphological and biochemical analysis of coated vesicle components in 3T3-L1 cells. Optical sectioning and image restoration revealed a marked increase in the staining of clathrin and beta adaptins in the perinuclear region of cells with differentiation. In addition, predominance of beta (subunit of the AP-2, plasma membrane adaptor) over beta' (subunit of the AP-1, Golgi adaptor) adaptin was observed in immunoblots of clathrin-coated vesicles purified from nondifferentiated fibroblasts, and this ratio was reversed in coated vesicles purified from differentiated adipocytes. These results indicate that the relative abundance of TGN-derived clathrin lattices increases markedly during adipocytic differentiation. Subcellular fractionation indicated that cytosolic AP-1 and AP-2 adaptors comprised approximately 70% of the total cellular adaptor pool. Interestingly, neither the concentration nor the relative ratio of cytosolic AP-1 to AP-2 adaptors increased significantly during differentiation. These data suggest that the increase in TGN-derived lattices results from differentiation-induced mechanisms for enhanced assembly or stabilization of adaptors on Golgi membranes. Interestingly, double-immunofluorescence microscopy also revealed that whereas extensive colocalization between clathrin and beta adaptins occurred both in fibroblasts and adipocytes, structures stained only with anti-adaptin antibody could be detected. Taken together these results suggest that membranes coated with adaptors, but not clathrin, can exist in these cells.
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Citation: J Cell Biol. 1993 Oct;123(1):79-87.
The Journal of cell biology
Chakrabarti, Ranjan; Joly, Marguerite; and Corvera, Silvia, "Redistribution of clathrin-coated vesicle adaptor complexes during adipocytic differentiation of 3T3-L1 cells" (1993). Open Access Articles. 956.