Brown adipose tissue (BAT) is a therapeutic target for metabolic diseases; thus, understanding its metabolic circuitry is clinically important. Many studies of BAT compare rodents mildly cold to those Show more
Brown adipose tissue (BAT) is a therapeutic target for metabolic diseases; thus, understanding its metabolic circuitry is clinically important. Many studies of BAT compare rodents mildly cold to those severely cold. Here, we compared BAT remodeling between thermoneutral and mild-cold-adapted mice, conditions more relevant to humans. Although BAT is renowned for catabolic β-oxidative capacity, we find paradoxically that the anabolic de novo lipogenesis (DNL) genes encoding ACLY, ACSS2, ACC, and FASN were among the most upregulated by mild cold and that, in humans, DNL correlates with Ucp1 expression. The regulation and function of adipocyte DNL and its association with thermogenesis are not understood. We provide evidence suggesting that AKT2 drives DNL in adipocytes by stimulating ChREBPβ transcriptional activity and that cold induces the AKT2-ChREBP pathway in BAT to optimize fuel storage and thermogenesis. These data provide insight into adipocyte DNL regulation and function and illustrate the metabolic flexibility of thermogenesis. Show less
The voltage-gated Na(+) channels (Na(v)) form a family composed of 10 genes. The COOH termini of Na(v) contain a cluster of amino acids that are nearly identical among 7 of the 10 members. This COOH-t Show more
The voltage-gated Na(+) channels (Na(v)) form a family composed of 10 genes. The COOH termini of Na(v) contain a cluster of amino acids that are nearly identical among 7 of the 10 members. This COOH-terminal sequence, PPSYDSV, is a PY motif known to bind to WW domains of E3 protein-ubiquitin ligases of the Nedd4 family. We recently reported that cardiac Na(v)1.5 is regulated by Nedd4-2. In this study, we further investigated the molecular determinants of regulation of Na(v) proteins. When expressed in HEK-293 cells and studied using whole cell voltage clamping, the neuronal Na(v)1.2 and Na(v)1.3 were also downregulated by Nedd4-2. Pull-down experiments using fusion proteins bearing the PY motif of Na(v)1.2, Na(v)1.3, and Na(v)1.5 indicated that mouse brain Nedd4-2 binds to the Na(v) PY motif. Using intrinsic tryptophan fluorescence imaging of WW domains, we found that Na(v)1.5 PY motif binds preferentially to the fourth WW domain of Nedd4-2 with a K(d) of approximately 55 muM. We tested the binding properties and the ability to ubiquitinate and downregulate Na(v)1.5 of three Nedd4-like E3s: Nedd4-1, Nedd4-2, and WWP2. Despite the fact that along with Nedd4-2, Nedd4-1 and WWP2 bind to Na(v)1.5 PY motif, only Nedd4-2 robustly ubiquitinated and downregulated Na(v)1.5. Interestingly, coexpression of WWP2 competed with the effect of Nedd4-2. Finally, using brefeldin A, we found that Nedd4-2 accelerated internalization of Na(v)1.5 stably expressed in HEK-293 cells. This study shows that Nedd4-dependent ubiquitination of Na(v) channels may represent a general mechanism regulating the excitability of neurons and myocytes via modulation of channel density at the plasma membrane. Show less