Abstract Adipose tissue is the primary site of energy storage, playing important roles in health. While adipose research largely focuses on obesity, fat also has other critical functions, producing adipocytokines and contributing to normal nutrient metabolism, which in turn play important roles in satiety and total energy homeostasis. SMAD2/3 proteins are downstream mediators of activin signaling, which regulate critical preadipocyte and mature adipocyte functions. Smad2 global knockout mice exhibit embryonic lethality, whereas global loss of Smad3 protects mice against diet‐induced obesity. The direct contributions of Smad2 and Smad3 in adipose tissues, however, are unknown. Here, we sought to determine the primary effects of adipocyte‐selective reduction of Smad2 or Smad3 on diet‐induced adiposity using Smad2 or Smad3 “floxed” mice intercrossed with Adiponectin ‐ Cre mice. Additionally, we examined visceral and subcutaneous preadipocyte differentiation efficiency in vitro . Almost all wild type subcutaneous preadipocytes differentiated into mature adipocytes. In contrast, visceral preadipocytes differentiated poorly. Exogenous activin A suppressed differentiation of preadipocytes from both depots. Smad2 conditional knockout ( Smad2 cKO) mice did not exhibit significant effects on weight gain, irrespective of diet, whereas Smad3 conditional knockout ( Smad3 cKO) male mice displayed a trend of reduced body weight on high‐fat diet. On both diets, Smad3 cKO mice displayed an adipose depot‐selective phenotype, with a significant reduction in subcutaneous fat mass but not visceral fat mass. Our data suggest that Smad3 is an important contributor to the maintenance of subcutaneous white adipose tissue in a sex‐selective fashion. These findings have implications for understanding SMAD‐mediated, depot selective regulation of adipocyte growth and differentiation.