Hormone-sensitive lipase (HSL) is the predominant lipase effector of catecholamine-stimulated lipolysis in adipocytes. HSL-dependent lipolysis in response to catecholamines is mediated by protein kinase A (PKA)-dependent phosphorylation of perilipin A (Peri A), an essential lipid droplet (LD)-associated protein.It is believed that perilipin phosphorylation is essential for the translocation of HSL from the cytosol to the LD, a key event in stimulated lipolysis. Using adipocytes retrovirally engineered from murine embryonic fibroblasts of perilipin null mice (Peri؊/؊ MEF), we demonstrate by cell fractionation and confocal microscopy that up to 50% of cellular HSL is LD-associated in the basal state and that PKA-stimulated HSL translocation is fully supported by adenoviral expression of a mutant perilipin lacking all six PKA sites (Peri A⌬1-6). PKA-stimulated HSL translocation was confirmed in differentiated brown adipocytes from perilipin null mice expressing an adipose-specific Peri A⌬1-6 transgene. Thus, PKA-induced HSL translocation was independent of perilipin phosphorylation. However, Peri A⌬1-6 failed to enhance PKA-stimulated lipolysis in either MEF adipocytes or differentiated brown adipocytes. Thus, the lipolytic action(s) of HSL at the LD surface requires PKA-dependent perilipin phosphorylation. In Peri؊/؊ MEF adipocytes, PKA activation significantly enhanced the amount of HSL that could be cross-linked to and co-immunoprecipitated with ectopic Peri A. Notably, this enhanced cross-linking was blunted in Peri؊/؊ MEF adipocytes expressing Peri A⌬1-6. This suggests that PKA-dependent perilipin phosphorylation facilitates (either direct or indirect) perilipin interaction with LD-associated HSL. These results redefine and expand our understanding of how perilipin regulates HSL-mediated lipolysis in adipocytes.The enzymatic hydrolysis of stored neutral lipid in adipocytes is an exquisitely regulated process that maintains whole body energy homeostasis in response to physiological demands. In both white and brown adipose tissue (BAT) 4 basal (constitutive) rates of adipocyte lipolysis are rapidly and dramatically up-regulated by lipolytic hormones such as catecholamines (1, 2). In white adipose tissue, catecholamine-induced lipolysis provides fatty acids as fuel to peripheral tissues during times of energy need, such as fasting and exercise (3-5). In BAT of human newborns and rodents, catecholamine-stimulated lipolysis provides fatty acids for heat production (via ␤-oxidation and mitochondrial uncoupling) in response to hypothermia (i.e. adaptive thermogenesis) (2, 6). Fatty acids that are released during adipocyte lipolysis also function as modulators of glucose and insulin action and insulin production (7,8). Moreover, the dysregulated release of fatty acids from adipocytes that occurs in obesity is implicated in the etiology of obesity-related complications, including type 2 diabetes (8 -10). Thus, in addition to its role in whole body energy homeostasis, the regulation of adipocyte lipolysis is vital to me...

5'-Adenosine monophosphate-activated protein kinase (AMPK) is a master regulator of energy homeostasis in eukaryotes. Despite three decades of investigation, the biological roles of AMPK and its potential as a drug target remain incompletely understood, largely because of a lack of optimized pharmacological tools. We developed MK-8722, a potent, direct, allosteric activator of all 12 mammalian AMPK complexes. In rodents and rhesus monkeys, MK-8722-mediated AMPK activation in skeletal muscle induced robust, durable, insulin-independent glucose uptake and glycogen synthesis, with resultant improvements in glycemia and no evidence of hypoglycemia. These effects translated across species, including diabetic rhesus monkeys, but manifested with concomitant cardiac hypertrophy and increased cardiac glycogen without apparent functional sequelae.

Hormonally stimulated lipolysis occurs by activation of cyclic AMP-dependent protein kinase (PKA) which phosphorylates hormone-sensitive lipase (HSL) and increases adipocyte lipolysis. Evidence suggests that catecholamines not only can activate PKA, but also the mitogen-activated protein kinase pathway and extracellular signal-regulated kinase (ERK). We now demonstrate that two different inhibitors of MEK, the upstream activator of ERK, block catecholamine-and ␤ 3 -stimulated lipolysis by ϳ30%. Furthermore, treatment of adipocytes with dioctanoylglycerol, which activates ERK, increases lipolysis, although MEK inhibitors decrease dioctanoylglycerol-stimulated activation of lipolysis. Using a tamoxifen regulatable Raf system expressed in 3T3-L1 preadipocytes, exposure to tamoxifen causes a 14-fold activation of ERK within 15-30 min and results in ϳ2-fold increase in HSL activity. In addition, when differentiated 3T3-L1 cells expressing the regulatable Raf were exposed to tamoxifen, a 2-fold increase in lipolysis is observed. HSL is a substrate of activated ERK and site-directed mutagenesis of putative ERK consensus phosphorylation sites in HSL identified Ser 600 as the site phosphorylated by active ERK. When S600A HSL was expressed in 3T3-L1 cells expressing the regulatable Raf, tamoxifen treatment fails to increase its activity. Thus, activation of the ERK pathway appears to be able to regulate adipocyte lipolysis by phosphorylating HSL on Ser 600 and increasing the activity of HSL.