Previous genetic studies in mice have shown that functional loss of activin receptor-like kinase 7 (ALK7), a type I transforming growth factor-β receptor, increases lipolysis to resist fat accumulation in adipocytes. Although growth/differentiation factor 3 (GDF3) has been suggested to function as a ligand of ALK7 under nutrient-excess conditions, it is unknown how GDF3 production is regulated. Here, we show that a physiologically low level of insulin converts CD11c adipose tissue macrophages (ATMs) into GDF3-producing CD11c macrophages ex vivo and directs ALK7-dependent accumulation of fat in vivo. Depletion of ATMs by clodronate upregulates adipose lipases and reduces fat mass in ALK7-intact obese mice, but not in their ALK7-deficient counterparts. Furthermore, depletion of ATMs or transplantation of GDF3-deficient bone marrow negates the in vivo effects of insulin on both lipolysis and fat accumulation in ALK7-intact mice. The GDF3-ALK7 axis between ATMs and adipocytes represents a previously unrecognized mechanism by which insulin regulates both fat metabolism and mass.

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 (Smad2cKO) mice did not exhibit significant effects on weight gain, irrespective of diet, whereas Smad3 conditional knockout (Smad3cKO) male mice displayed a trend of reduced body weight on high‐fat diet. On both diets, Smad3cKO 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.

Key Points• TALEs targeted to the g-globin gene promoters reactivated their mRNA expression more than 70-fold with a collateral reduction in b-globin mRNA.• At day 19 of CD34 erythroid differentiation, TALEs increased g-globin more than 40fold in mRNA level and up to 70% of the total globin protein.The fetal-to-adult hemoglobin switch has been a focus of a long-standing effort to potentially treat sickle cell disease and b thalassemia by induction of fetal hemoglobin. In a continuation of this effort, we designed specific transcriptional activator-like effectors (TALEs) to target both the G g and A g-globin promoters. We fused the TALEs to a LIM domain binding protein (Ldb1) dimerization domain, followed by a T2A green fluorescent protein (GFP) cassette, which were assembled into a lentiviral vector. To prevent deletions caused by the repeats of TALEs during the lentivirus packing process, we changed the TALE encoding DNA by codon optimization. Intriguingly, 5 of 14 TALEs showed forced reactivation of fetalglobin expression in human umbilical cord blood-derived erythroid progenitor (HUDEP-2) cells, with a significant increase in the g-globin mRNA level by more than 70-fold. We also observed a more than 50% reduction of b-globin mRNA. High-performance liquid chromatography analysis revealed more than 30% fetal globin in TALE-induced cells compared with the control of 2%. Among several promoters studied, the b-globin gene promoter with the locus control region (LCR) enhancer showed the highest TALE expression during CD34 erythroid differentiation. At day 19 of differentiation, 2 TALEs increased fetalglobin expression more than 40-fold in the mRNA level and up to 70% of the total globin protein. These TALEs have potential for clinical translation.