The critical role of atypical protein kinase C in activating hepatic SREBP-1c and NFkB in obesity

Obesity is frequently associated with systemic insulin resistance, glucose intolerance, and hyperlipidemia. Impaired insulin action in muscle and paradoxical diet/ insulin-dependent overproduction of hepatic lipids are important components of obesity, but their pathogenesis and inter-relationships between muscle and liver are uncertain. We studied two murine obesity models, moderate high-fatfeeding and heterozygous muscle-specific PKC-l knockout, in both of which insulin activation of atypical protein kinase C (aPKC) is impaired in muscle, but conserved in liver. In both models, activation of hepatic sterol receptor element binding protein-1c (SREBP-1c) and NFkB (nuclear factorkappa B), major regulators of hepatic lipid synthesis and systemic insulin resistance, was chronically increased in the fed state. In support of a critical mediatory role of aPKC, in both models, inhibition of hepatic aPKC by adenovirally mediated expression of kinase-inactive aPKC markedly diminished diet/insulin-dependent activation of hepatic SREBP-1c and NFkB, and concomitantly improved hepatosteatosis, hypertriglyceridemia, hyperinsulinemia, and hyperglycemia. Moreover, in high-fat–fed mice, impaired insulin signaling to IRS-1–dependent phosphatidylinositol 3-kinase, PKB/Akt and aPKC in muscle and hyperinsulinemia were largely reversed. In obesity, conserved hepatic aPKC-dependent activation of SREBP-1c and NFkB contributes importantly to the development of hepatic lipogenesis, hyperlipidemia, and systemic insulin resistance. Accordingly, hepatic aPKC is a potential target for treating obesityassociated abnormalities.—Sajan, M. P., M. L. Standaert, S. Nimal, U. Varanasi, T. Pastoor, S. Mastorides, U. Braun, M. Leitges, and R. V. Farese. The critical role of atypical protein kinase C in activating hepatic SREBP-1c and NFkB in obesity. J. Lipid Res. 2009. 50: 1133–1145. Supplementary key words atypical protein kinase C • high fat feeding • hyperlipidemia • insulin • insulin resistance • IRS-1 • IRS-2 • liver • muscle • NFkB • obesity • phosphatidylinositol 3-kinase • SREBP-1c • type 2 diabetes Obesity, particularly when accompanied by systemic insulin resistance, glucose intolerance, and hyperlipidemia (i.e., a “metabolic syndrome”) is a global health problem and a frequent forerunner of type 2 diabetes mellitus. Whereas both exogenous/diet-induced and genetically determined obesity can produce insulin resistance and metabolic syndrome features, vice versa, systemic insulin resistance can produce obesity and metabolic syndrome features. However, mechanisms underlying lipid abnormalities and insulin resistance in these situations, and the critical interplay between muscle and liver, are poorly understood. The high-fat–fed (HFF) mouse model is useful for studying diet-induced obesity-related insulin resistance. In our experience, feeding mice a Western-type 20% milk highfat diet for 3–4 weeks leads to diminished insulin activation of phosphatidylinositol (PI) 3-kinase (PI3K) effectors, atypical protein kinase C (aPKC) and protein kinase B (PKB/Akt) in muscle (1, 2), with little or no effect on hepatic aPKC and PKB/Akt activation (1). In this HFF model, we have observed no increases in basal (unstimulated) activities of conventional (a,b2) or novel (y,y) PKCs in liver, despite observing increases in muscle (unpublished). Accordingly, our HFF model may partly differ from others wherein higher dietary fat was used, thereby activating hepatic novel PKCs. Insulin signaling to aPKC and PKB/Akt (particularly the PKBb/Akt2 isoform) is important, as these kinases control key metabolic processes, viz., glucose transport in muscle is controlled by both aPKC and PKB/Akt, enzymes regulating hepatic glucose output are mainly controlled by PKB/Akt (3, 4), and hepatic sterol receptor element binding protein-1c (SREBP-1c), which trans-activates multiple genes involved in lipid synthesis, can be controlled by both aPKC (3, 4) and PKB/Akt (5, 6). Also, hepaticNFkB (nuclear This work was supported by funds from the Department of Veterans Affairs Merit Review Program and the National Institutes of Health DK-38079 (R. V. F.), and by the Deutsche Forschungsgemeinschaft Sta314/2-1 and KE246/7-2 (M. L.). Manuscript received 8 October 2008 and in revised form 16 January 2009. Published, JLR Papers in Press, February 6, 2009. DOI 10.1194/jlr.M800520-JLR200 Abbreviations: aPKC, atypical protein kinase C; SREBP-1c, hepatic sterol receptor element binding protein-1c; KO, knockout; NFkB, nuclear factor-kappa B. 1 To whom correspondence should be addressed. e-mail: [email protected] This article is available online at http://www.jlr.org Journal of Lipid Research Volume 50, 2009 1133 by gest, on N ovem er 7, 2017 w w w .j.org D ow nladed fom factor-kappa B), which is activated in HFF mice and linked to systemic insulin resistance (7, 8) and inflammation, can be activated by aPKC through (a) phosphorylation/ activation of IkB kinase (IKKb), which phosphorylates IkBb, thereby releasing NFkB for translocation to nuclear transcription sites (9); and (b) phosphorylation/activation of the p65/RelA subunit of NFkB (10). However, insulin effects on IKKb and NFkB are unknown. Differences in activation of aPKC and PKB/Akt inmuscle and liver in HFF mice most likely reflect tissue-specific variations in PI3K activation by insulin receptor substrates (IRS), IRS-1 and IRS-2. However, definitive information on PI3K activation in HFF mice is lacking, and findings in other HFF models are inconsistent. In rats fed 58% lard, activation of hepatic IRS-1– and IRS-2–dependent PI3K is oddly enough enhanced (11); in rats fed 21% milk fat, activation of muscle aPKC by insulin in vivo and PI-3,4,5-(PO4)3 (PIP3) in vitro is impaired, but activation of IRS-1– and IRS-2– dependent PI3K is intact (12); in rats fed 65% fat, aPKC and PKB activation in muscle is diminished (13). Germane to tissue-specific differences in aPKC/PKB activation, findings from knockout mice suggest that IRS-1 and IRS-2 activate separate PI3K pools that couple differently to these downstream effectors during insulin action, depending upon the tissue. Thus, in IRS-1 knockout mice, insulin activation of both PKB/Akt (14, 15) and aPKC (15) is impaired in muscle, whereas in liver, PKB/Akt activation is impaired (14, 15), but aPKC activation is conserved (15). In IRS-2-deficient hepatocytes, activation of aPKC, as well as PKB/Akt, is impaired (16). To summarize, IRS-1 controls both aPKC and PKB/Akt in muscle, whereas in liver, IRS-2 controls aPKC, and both IRS-1 and IRS-2 control PKB/Akt activation by insulin. In HFF mice, we found that inhibition of hepatic aPKC by adenovirally mediated expression of kinase-inactive (KI) aPKC largely reversed excessive increases in hepatic SREBP-1c and IKKb/NFkB activities, thereby improving downstream targets of SREBP-1c and NFkB, hyperlipidemia, insulin signaling in muscle, hyperinsulinemia, and hyperglycemia. Similarly, in a genetic obesity model, heterozygous muscle-specific knockout of PKC-l (Ml-Het-KO) mice [wherein (a) aPKC activation is selectively impaired in muscle, thereby compromising insulin-stimulated glucose transport, (b) insulin signaling and actions in liver and adipocytes are intact, and (c) feedingand insulindependent activation of hepatic SREBP-1c and IKKb/NFkB are excessive, thereby producing abdominal obesity, hepatosteatosis, hyperlipidemia, and glucose intolerance (17)], we observed comparable dependency of hepatic SREBP-1c and IKKb/NFkB on hepatic aPKC and, moreover, similar phenotypic improvements following administration of adenovirus expressing KI-aPKC. EXPERIMENTAL PROCEDURES