Adipocyte differentiation takes place via a complex series of steps. While PPAR gamma and C/EBP alpha are known to be master regulators, the events at the earliest stage of adipocyte differentiation are not yet known. In this study, we cloned the genes that are induced at the beginning of the differentiation of 3T3-L1 preadipocyte cells. Of 102 clones obtained, only several clones were already reported as genes that are expressed differentially during adipocyte development. The expression of TCL/TC10 beta L (TC10-like/TC10 beta Long) and RGS2 (regulators of G protein signaling 2) genes isolated here rapidly increased after the addition of inducers (insulin, dexemethasone, 3-isobutyl-1-methylxanthine, fetal bovine serum [FBS]). Further, the antisense TCL/TC10 beta L inhibited the adipogenesis of mouse 3T3-L1 preadipocyte cells, prevented cytoplasmic triglyceride accumulation, and decreased the expression of PPAR gamma and C/EBP alpha. Moreover, the constitutive overexpression of TCL/TC10 beta L or RGS2 in the mouse fibroblast cell line NIH-3T3 results in efficient adipocyte conversion when stimulated with 10% FBS, insulin, 3-isobutyl-1-methylxanthine, dexamethasone, and PPAR gamma ligand BRL49653. These results strongly suggest that TCL/TC10 beta L and RGS2 have crucial roles in the program of adipocyte differentiation, probably linked to the PPAR gamma pathway. Using a subtraction protocol, the genes specifically regulated by TCL/TC10 beta L were also isolated. The expression pattern of some was similar to TCL/TC10 beta L expression in adipogenesis, suggesting that these genes are regulated by TCL/TC10 beta L.