Based on recent evidence that fatty acid synthase and endogenously produced fatty acid derivatives are required for adipogenesis in 3T3-L1 adipocytes, we conducted a siRNA-based screen to identify other fatty acid metabolizing enzymes that may mediate this effect. Of 24 enzymes screened, Stearoyl CoA Desaturase 2 (SCD2) was found to be uniquely and absolutely required for adipogenesis. Remarkably, SCD2 also controls the maintenance of adipocyte-specific gene expression in fully differentiated 3T3-L1 adipocytes, including the expression of SCD1. Despite the high sequence similarity between SCD2 and SCD1, silencing of SCD1 did not downregulate 3T3-L1 cell differentiation or gene expression. SCD2 mRNA expression was also uniquely elevated 44 fold in adipose tissue upon feeding mice a high fat diet, while SCD1 showed little response. The inhibition of adipogenesis caused by SCD2 depletion was associated with a decrease in PPAR mRNA and protein, while in mature adipocytes loss of SCD2 diminished PPAR protein levels, with little change in mRNA levels. In the latter case, SCD2 depletion did not change the degradation rate of PPAR protein, but decreased the metabolic labeling of PPAR protein using [35S] methionine/cysteine, indicating protein translation was decreased. This requirement of SCD2 for optimal protein synthesis in fully differentiated adipocytes was verified by polysome profile analysis, where a shift in the mRNA to monosomes was apparent in response to SCD2 silencing. These results reveal that SCD2 is required for the induction and maintenance of PPAR protein levels and adipogenesis in 3T3-L1 cells.