Insulin Stimulates Adipogenesis through the Akt-TSC2-mTORC1 Pathway

BACKGROUND:The signaling pathways imposing hormonal control over adipocyte differentiation are poorly understood. While insulin and Akt signaling have been found previously to be essential for adipogenesis, the relative importance of their many downstream branches have not been defined. One direct substrate that is inhibited by Akt-mediated phosphorylation is the tuberous sclerosis complex 2 (TSC2) protein, which associates with TSC1 and acts as a critical negative regulator of the mammalian target of rapamycin (mTOR) complex 1 (mTORC1). Loss of function of the TSC1-TSC2 complex results in constitutive mTORC1 signaling and, through mTORC1-dependent feedback mechanisms and loss of mTORC2 activity, leads to a concomitant block of Akt signaling to its other downstream targets. METHODOLOGY/PRINCIPAL FINDINGS:We find that, despite severe insulin resistance and the absence of Akt signaling, TSC2-deficient mouse embryo fibroblasts and 3T3-L1 pre-adipocytes display enhanced adipocyte differentiation that is dependent on the elevated mTORC1 activity in these cells. Activation of mTORC1 causes a robust increase in the mRNA and protein expression of peroxisome proliferator-activated receptor gamma (PPARgamma), which is the master transcriptional regulator of adipocyte differentiation. In examining the requirements for different Akt-mediated phosphorylation sites on TSC2, we find that only TSC2 mutants lacking all five previously identified Akt sites fully block insulin-stimulated mTORC1 signaling in reconstituted Tsc2 null cells, and this mutant also inhibits adipogenesis. Finally, renal angiomyolipomas from patients with tuberous sclerosis complex contain both adipose and smooth muscle-like components with activated mTORC1 signaling and elevated PPARgamma expression. CONCLUSIONS/SIGNIFICANCE:This study demonstrates that activation of mTORC1 signaling is a critical step in adipocyte differentiation and identifies TSC2 as a primary target of Akt driving this process. Therefore, the TSC1-TSC2 complex regulates the differentiation of mesenchymal cell lineages, at least in part, through its control of mTORC1 activity and PPARgamma expression

Activation of a Metabolic Gene Regulatory Network Downstream of mTOR Complex 1

Aberrant activation of the mammalian target of rapamycin complex 1 (mTORC1) is a common molecular event in a variety of pathological settings, including genetic tumor syndromes, cancer, and obesity. However, the cell-intrinsic consequences of mTORC1 activation remain poorly defined. Through a combination of unbiased genomic, metabolomic, and bioinformatic approaches, we demonstrate that mTORC1 activation is sufficient to stimulate specific metabolic pathways, including glycolysis, the oxidative arm of the pentose phosphate pathway, and de novo lipid biosynthesis. This is achieved through the activation of a transcriptional program affecting metabolic gene targets of hypoxia-inducible factor (HIF1α) and sterol regulatory element-binding protein (SREBP1 and SREBP2). We find that SREBP1 and 2 promote proliferation downstream of mTORC1, and the activation of these transcription factors is mediated by S6K1. Therefore, in addition to promoting protein synthesis, mTORC1 activates specific bioenergetic and anabolic cellular processes that are likely to contribute to human physiology and disease

PP2A Phosphatase Activity Is Required for Stress and Tor Kinase Regulation of Yeast Stress Response Factor Msn2p

In response to stress and nutrient starvation, the Saccharomyces cerevisiae transcription factor Msn2p accumulates in the nucleus and activates expression of a broad array of genes. Here, we analyze the role of the Tor (target of rapamycin) signaling pathway in mediating these responses. Inactivation of the Tor pathway component Tap42p using tap42(Ts) alleles causes a sustained nuclear localization similar to that after the addition of the Tor kinase inhibitor rapamycin. Effects of Tap42p inactivation and rapamycin addition could be suppressed by deletion of TIP41, which encodes a Tap42p-interacting protein. These results support the notion that rapamycin affects Msn2p by inactivating Tap42p function. Tap42p interacts with the catalytic subunit of PP2A (protein phosphatase 2A) and PP2A-like phosphatases. Deletion of either the catalytic or regulatory subunit that forms the PP2A phosphatase complex prevents nuclear accumulation of Msn2p in the tap42(Ts) strain and in wild-type strains treated with rapamycin. These results suggest that Tap42p is an inhibitor of PP2A phosphatase, which in turn inhibits nuclear export of Msn2p. Interestingly, PP2A function is also required for nuclear accumulation of Msn2p in response to stresses, such as heat and osmotic shock, as well as nitrogen (but not glucose) starvation. Thus, PP2A and the Tor kinase pathway transduce stress and nitrogen starvation signals to Msn2p. Finally, Msn2p localization is unaffected by conditional loss of 14-3-3 protein function, ruling out the possibility that 14-3-3 proteins act as a scaffold to sequester Msn2p in the cytoplasm

Activation of a metabolic gene regulatory network downstream of mTOR complex 1

Aberrant activation of the mammalian target of rapamycin (mTOR) complex 1 (mTORC1) is a common molecular event in a large variety of pathological settings, including genetic tumor syndromes, cancer, and obesity. However, the cell intrinsic consequences of mTORC1 activation remain poorly defined. Through a combination of unbiased genomic, metabolomic, and bioinformatic approaches, we demonstrate that mTORC1 activation is sufficient to stimulate specific metabolic pathways, including glycolysis, the oxidative arm of the pentose phosphate pathway, and de novo lipid biosynthesis. This is achieved through the activation of a transcriptional program affecting metabolic gene targets of hypoxia-inducible factor (HIF1) and sterol regulatory element-binding protein (SREBP1 and SREBP2). We find that SREBP1 and 2 promote proliferation downstream of mTORC1, and the activation of these transcription factors is mediated by S6K1. Therefore, in addition to promoting protein synthesis, mTORC1 activates specific bioenergetic and anabolic cellular processes that are likely to contribute to human physiology and disease