Protein kinases in hormonal regulation of adipocyte metabolism.

Abstract Along with liver and muscle tissue, adipose tissue helps maintain normal levels of glucose and lipids in the blood and has a very important role when it comes to storing lipids that can provide whole-body energy. After a meal is ingested, adipocytes take up glucose from the circulation and use it as a substrate for synthesis of new fatty acids (FAs) in a process known as de novo fatty acid synthesis, as well as for synthesis of glycerol. Adipocytes also take up fatty acids from the circulation and incorporate both newly synthesized and imported FAs into triacylglycerides (TAGs), in a process known as lipogenesis. TAGs are stored in large lipid droplets in the cytosol, and during fasting, or in response to physical exercise, they are hydrolysed in a process known as lipolysis, in which FAs are released into the bloodstream for use as energy substrates in other tissues. These cycles of lipogenesis and lipolysis are controlled by the concerted actions of insulin, a hormone that is secreted by the pancreas and catecholamines, hormones that are secreted by the adrenal glands, or derive from the nervous system. Both glucose- and fatty acid uptake, as well as lipid storage and mobilization, are regulated by cellular signaling, and kinases are central enzymatic players in hormone-induced cellular signaling. A dysfunctional adipose tissue can contribute to insulin resistance in many obese individuals. Therefore it is important to elucidate the cellular mechanisms that govern metabolic processes in adipocytes. Insulin is the hormone that promotes glucose uptake and lipogenesis in adipocytes, and when it induces glucose uptake, insulin exerts it actions through protein kinase B (PKB). Although PKB is known to mediate many effects of insulin, its role in lipogenesis in adipocytes is less clear. We show that PKB is important for the effects of insulin on lipogenesis (de novo and total). We also reveal that PKB can regulate Amp-activated protein kinase (AMPK) in adipocytes by a mechanism previously only seen in heart muscle cells. AMPK is a sensor of cellular energy status and known to inhibit lipogenesis. We speculate that insulin possibly mediates its lipogenic effects via a decrease in AMPK activity accomplished by PKB-phosphorylation of S485 on AMPK. Furthermore, we find that salt-inducible kinase 3 (SIK3), a kinase that belongs to the AMPK-related family of kinases, and displays structural similarities to AMPK, can be regulated by catecholamines in adipocytes. Catecholamines are hormones that bind to β-adrenergic receptors and act by increasing cellular levels of cAMP, which in turn activates protein kinase A (PKA). We find that in response to such β-adrenergic stimuli, SIK3 is phosphorylated on multiple serine and threonine residues. This 10 regulation coincides with an increase in binding of SIK3 to 14-3-3 molecules. 14-3-3 proteins are cellular scaffolding proteins that can result in cellular re-localization of their binding partners or in their binding to other proteins or lipids. We find that when SIK3 is phosphorylated in response to β-adrenergic stimuli, the kinase does not re-localize, but is partially de-activated. We speculate that SIK3 could potentially have a role in adipocyte metabolism, as it is regulated by catecholamines in this tissue. Finally, we address the current understanding of the role for AMPK in modulation of the effects of insulin and catecholamines on glucose uptake and lipid metabolism. To this date, it has been suggested that AMPK reduces insulin-induced glucose uptake and lipogenesis, as well as inhibits catecholamine-induced lipolysis in adipocytes. These findings are mainly based on studies performed with AMPK activating agents that act on AMPK in an indirect manner. We have used the allosteric activator A769662, that binds directly to AMPK, and find that AMPK does not appear to modulate hormonally induced glucose uptake, lipolysis or total lipogenesis. However, when we specifically measured the synthesis of new FAs, using acetate as a lipogenic substrate (as opposed to using glucose as a substrate, a molecule which can participate in both FA and glycerol synthesis), we observe that AMPK does indeed reduce insulin-induced de novo fatty acid synthesis. Collectively, we add novel findings to the available knowledge on key kinases and cellular signaling in adipocyte metabolism. Our findings contribute to the understanding of insulin- and catecholamine-mediated control of lipid storage in adipose tissue, a biological function that, when dysfunctional, is strongly linked to insulin resistance and type 2 diabetes (T2D)