Author's School

Graduate School of Arts & Sciences

Author's Department/Program

Biology and Biomedical Sciences: Molecular Genetics and Genomics

Language

English (en)

Date of Award

8-15-2012

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Chair and Committee

Shin-ichiro Imai

Abstract

Nicotinamide phosphoribosyltransferase: NAMPT) is the rate-limiting enzyme in the NAD biosynthesis pathway in mammals. Interestingly, NAMPT has intra- and extracellular forms: iNAMPT and eNAMPT, respectively), and eNAMPT is secreted from adipose tissue as one of the adipokines. However, the mechanism underlying the regulation of eNAMPT secretion has not been fully understood yet. Here, we have demonstrated that deacetylation on NAMPT by SIRT1 controls its secretion from adipocytes. iNAMPT is acetylated in both brown and white adipose tissues, and the level of acetylation of iNAMPT is decreased during fasting when Sirtuins are activated. We also found that enhancement of eNAMPT secretion under low glucose culture condition in differentiated brown and white adipocytes is completely inhibited by nicotinamide: NAM), an inhibitor of SIRTs, suggesting that deacetylation by SIRTs is required to induce eNAMPT secretion by low glucose stimulus. Consistently, eNAMPT levels in the blood were enhanced after 48 hours fasting in WT but not in Sirt1 KO mice, and iNAMPT acetylation levels are decreased while eNAMPT secretion is increased in differentiated adipocytes by SIRT1 overexpression. In addition, iNAMPT physically interacts with SIRT1 in differentiated adipocytes, and this interaction is enhanced under a low-glucose culture condition, suggesting that SIRT1 deacetylates iNAMPT by direct interaction and increases eNAMPT secretion. We have found five acetylation sites on iNAMPT and only 4 out of 5 were deacetylated. SIRT1 is responsible for deacetylating lysine 53, which results in both increased enzymatic activity and secretion of eNAMPT. These results indicate a novel feedback loop regulating systemic NAD biosynthesis through SIRT1 and eNAMPT in adipose tissues. Next, in an effort to investigate the function of eNAMPT secreted from adipose tissue, we generated adipocyte-specific Nampt KO: ANKO) mice. We first confirmed that female ANKO mice show significant reductions in plasma eNAMPT levels. Interestingly, female ANKO show reduction of NAD levels not only in white and brown adipose tissue but also in the hypothalamus compared to control Namptflox/flox mice, suggesting that loss of Nampt only in adipose tissues could also influence hypothalamic function, including governing glucose homeostasis by lowering NAD levels in this tissue. Consistent with this notion, female ANKO exhibit severely impaired glucose tolerance and glucose-stimulated insulin secretion accompanied by hyperinsulinemia, and administration of NMN partially restores insulin sensitivity but almost completely ameliorates the responsiveness of pancreatic β cells to glucose and hyperinsulinemia in vivo. Taken together, our results suggest the new possibility that adipose tissues play an important role in maintaining glucose homeostasis by controlling eNAMPT secretion and eNAMPT-mediated NAD biosynthesis at a systemic level and provide important insight into therapeutic and preventive intervention for metabolic diseases such as type 2 diabetes.

Comments

Permanent URL: http://dx.doi.org/10.7936/K7WS8R8Z

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