Hong Yu Wang

The Rab GTPase activating protein (RabGAP) AS160 translocates from the cytosol into the nucleus acting as a transcriptional co-activator of Signal Transducer and Activator of Transcription 3 (STAT3) to regulate proliferation of muscle satellite cells (MuSCs). How this AS160–STAT3 complex is regulated remains largely unclear yet. Here, we show that TBC1D1, a RabGAP related to AS160, forms a super-complex with AS160 and STAT3 to retain the AS160–STAT3 complex in the cytosol. Phosphorylation of TBC1D1-Thr⁵⁹⁶ by protein kinase B dissociates TBC1D1 from AS160 thus releasing the cytosolic retention of the AS160–STAT3 complex. A non-phosphorylatable alanine substitution of Thr⁵⁹⁶ inhibits MuSC proliferation and impairs repair of injured muscle. In contrast, TBC1D1 deficiency, but not its GAP-inactive mutation, promotes MuSC proliferation and muscle regeneration. Thus, TBC1D1 is a negative regulator of MuSC proliferation through cytosolic retention of the AS160–STAT3 complex and might be a valuable therapeutic target for muscle regeneration.

Obesity is a major pathological factor that induces insulin resistance and consequent type 2 diabetes through multiple mechanisms. Inactivation of the insulin receptor (INSR) contributes to the development of insulin resistance, whose protein level is down-regulated in obesity through as yet-undefined mechanisms. Here we show that the E3-ligase TRAF6 is a critical regulator of INSR maturation, whose inactivation prevents palmitic acid- or high-fat diet-induced diminution of the INSR. Consequently, genetic inactivation of TRAF6 enhances insulin signaling that further increases muscle glucose uptake and inhibits hepatic gluconeogenesis. TRAF6 inactivation increases the proprotein convertase FURIN that controls the processing of pro-INSR to mature INSR. Mechanistically, TRAF6 associates with the Golgi apparatus, where it ubiquitinates the cytosolic tail of FURIN, leading to its lysosomal degradation. This TRAF6-FURIN axis also regulates cholesterol metabolism via PCSK9 processing in the circulation. Collectively, our results reveal a critical role of TRAF6 in regulating proprotein processing and have therapeutic implications for metabolic control.

Dysfunction of muscle satellite cells is linked to diabetic myopathy. The mechanisms vitiating muscle satellite cell proliferative activity in diabetes are not well understood. Here, we show that AS160, a key cytosolic Rab-GTPase activating protein (RabGAP) in insulin signaling, is a moonlighting protein regulating muscle satellite cell proliferation as a transcriptional co-factor. Deletion of AS160, but not its GAP-inactive mutation, impairs muscle satellite cell proliferation and consequent muscle regeneration, and exacerbates age-related sarcopenia. Mechanistically, Thr⁶⁴² phosphorylation of AS160 promotes its translocation into the nucleus where AS160 functions as a co-factor of Signal Transducer and Activator of Transcription 3 (STAT3). AS160 binds to STAT3 to enhance the transcription of myogenic cascades and consequent muscle satellite cell proliferation. Disruption of the AS160-STAT3 interaction, or inhibition of AS160-Thr⁶⁴² phosphorylation, inhibits muscle satellite cell proliferation and impairs muscle regeneration. Together, our findings reveal a moonlighting function of AS160 as a transcriptional co-factor in the nucleus, and have therapeutic implications for muscle regeneration.

Insulin-like growth factor 1 (IGF1) is a regulator of both cellular hypertrophy and lipogenesis, which are two key processes for pathogenesis of obesity. However, the in vivo role of IGF1 in the development of obesity remains unclear. Here, we show that IGF1 expression is increased in adipose tissue in obese human patients and animal models. Elevation of IGF1 is associated with increased lipogenic gene expression and decreased energy expenditure. Genetic down-regulation of IGF1 normalizes lipogenic gene expression, restores aberrant energy metabolism and alleviates obese phenotype of a genetic mouse model with IGF1-hypersecretion. Importantly, genetic down-regulation of IGF1 exerts similar effects on development of diet-induced obesity. Furthermore, berberine that is an AMP-activated protein kinase (AMPK) activator in medicinal herbs inhibits IGF1 secretion, decreases lipogenic gene expression and alleviates diet-induced adiposity. Collectively, our findings demonstrate that hypersecretion of IGF1 is a critical factor for the development of obesity and can be targeted using AMPK activators to alleviate obesity.

The obese heart undergoes metabolic remodeling and exhibits impaired calcium (Ca²⁺) homeostasis, which are two critical assaults leading to cardiac dysfunction. The molecular mechanisms underlying these alterations in obese heart are not well understood. Here, we show that the Rab-GTPase activating protein AS160 is a lipid-responsive regulator of Ca²⁺ homeostasis through governing lysophosphatidylinositol metabolism and signaling. Palmitic acid/high fat diet inhibits AS160 activity through phosphorylation by NEK6, which consequently activates its downstream target Rab8a. Inactivation of AS160 in cardiomyocytes elevates cytosolic Ca²⁺ that subsequently impairs cardiac contractility. Mechanistically, Rab8a downstream of AS160 interacts with DDHD1 to increase lysophosphatidylinositol metabolism and signaling that leads to Ca²⁺ release from sarcoplasmic reticulum. Inactivation of NEK6 prevents inhibition of AS160 by palmitic acid/high fat diet, and alleviates cardiac dysfunction in high fat diet-fed mice. Together, our findings reveal a regulatory mechanism governing metabolic remodeling and Ca²⁺ homeostasis in obese heart, and have therapeutic implications to combat obesity cardiomyopathy.

Energy status is linked to the production of reactive oxygen species (ROS) in macrophages, which is elevated in obesity. However, it is unclear how ROS production is upregulated in macrophages in response to energy overload for mediating the development of obesity. Here, we show that the Rab-GTPase activating protein (RabGAP) TBC1D1, a substrate of the energy sensor AMP-activated protein kinase (AMPK), is a critical regulator of macrophage ROS production and consequent adipose inflammation for obesity development. TBC1D1 deletion decreases, whereas an energy overload-mimetic non-phosphorylatable TBC1D1^S231A mutation increases, ROS production and M1-like polarization in macrophages. Mechanistically, TBC1D1 and its downstream target Rab8a form an energy-responsive complex with NOX2 for ROS generation. Transplantation of TBC1D1^S231A bone marrow aggravates diet-induced obesity whereas treatment with an ultra-stable TtSOD for removal of ROS selectively in macrophages alleviates both TBC1D1^S231A mutation- and diet-induced obesity. Our findings therefore have implications for drug discovery to combat obesity.

Diabetic cardiomyopathy (DCM) is currently a progressive and nonstoppable complication in type 2 diabetic patients. Metabolic insults and insulin resistance are involved in its pathogenesis; however, the underlying mechanisms are still not clearly understood. Here we show that calcium dysregulation can be both a cause and a consequence of cardiac insulin resistance that leads to DCM. A western diet induces the development of DCM through at least three phases in mice, among which an early phase depends on impaired Thr⁴⁸⁴-phosphorylation of sarcoplasmic/endoplasmic reticulum calcium ATPase 2a (SERCA2a) elicited by insulin resistance. Mutation of SERCA2a-Thr⁴⁸⁴ to a nonphosphorylatable alanine delays calcium re-uptake into the sarcoplasmic reticulum in the cardiomyocytes and decreases cardiac function at the baseline. Importantly, this mutation blunts the early phase of DCM, but has no effect on disease progression in the following phases. Interestingly, impairment of sarcoplasmic reticulum calcium re-uptake caused by the SERCA2a-Thr⁴⁸⁴ mutation inhibited processing of insulin receptor precursor through FURIN convertase, resulting in cardiac insulin resistance. Collectively, these data reveal a bidirectional relationship between insulin resistance and impairment of calcium homeostasis, which may underlie the early pathogenesis of DCM. Our findings have therapeutic implications for early intervention of DCM.

Sarcoplasmic/endoplasmic reticulum calcium ATPase SERCA2 mediates calcium re-uptake from the cytosol into sarcoplasmic reticulum, and its dysfunction is a hallmark of heart failure. Multiple factors have been identified to modulate SERCA2 activity, however, its regulation is still not fully understood. Here we identify a Ral-GTPase activating protein RalGAPα1 as a critical regulator of SERCA2 in cardiomyocytes through its downstream target RalA. RalGAPα1 is induced by pressure overload, and its deficiency causes cardiac dysfunction and exacerbates pressure overload-induced heart failure. Mechanistically, RalGAPα1 regulates SERCA2 through direct interaction and its target RalA. Deletion of RalGAPα1 decreases SERCA2 activity and prolongs calcium re-uptake into sarcoplasmic reticulum. GDP-bound RalA, but not GTP-bound RalA, binds to SERCA2 and activates the pump for sarcoplasmic reticulum calcium re-uptake. Overexpression of a GDP-bound RalA^S28N mutant in the heart preserves cardiac function in a mouse model of heart failure. Our findings have therapeutic implications for treatment of heart failure.