Genistein enhances NAD+ biosynthesis by binding to Prohibitin 1 and upregulating nicotinamide phosphoribosyltransferase in adipocytes

Decreased NAD+ levels in adipocytes cause adipose-tissue dysfunction, leading to systemic glucose and lipid metabolism failure. Therefore, developing small molecules and nutraceuticals that can increase NAD+ levels in adipocytes is necessary. Genistein, a nutraceutical derived from soybeans, has various physiological activities and improves glucose and lipid metabolism. In this study, we aimed to unravel the effects of genistein on the intracellular NAD+ levels in adipocytes and the underlying molecular mechanisms. We showed that genistein enhanced NAD+ biosynthesis by increasing the expression of nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in NAD+ biosynthesis. A pull-down assay using genistein-immobilized beads identified prohibitin 1 (PHB1) as a target protein of genistein. The knockdown of PHB1 suppressed the genistein-induced increase in NAMPT expression and NAD+ levels in adipocytes. Genistein-bound PHB1 contributed to the stabilization of the transcription factor CCAAT/enhancer-binding protein β through activation of extracellular signal-regulated kinase, resulting in increased NAMPT expression at the transcriptional level. Genistein induced dephosphorylation of peroxisome proliferator-activated receptor at serine 273 and increased the insulin-sensitizing adipokine, adiponectin, in adipocytes, whereas the knockdown of NAMPT and PHB1 abolished these genistein-mediated effects. Our results proved the potential efficacy of nutraceuticals in promoting NAD+ levels and restoring metabolic function in adipocytes. Furthermore, we identified PHB1, localized to the plasma membrane, as a candidate target protein for increased expression of NAMPT in adipocytes. Overall, these findings will assist in developing NAD+ boosting strategies to alleviate the metabolic dysfunctions in adipose tissues. Significance Statement Increasing NAD+ levels is an important preventive strategy for maintaining metabolic function. Here, we showed that genistein, a nutraceutical, which increases NAD+ levels in adipocytes, increased NAD+ biosynthesis by upregulating nicotinamide phosphoribosyltransferase (NAMPT), a rate-limiting enzyme in the NAD+ biosynthesis pathway. Our findings also showed that genistein increased NAMPT expression by binding to prohibitin 1 in the plasma membrane. Genistein-induced increase in NAD+ levels promoted adiponectin expression, an insulin-sensitizing adipokine, in adipocytes. This study provides evidence that nutraceuticals, such as genistein, are effective in enhancing NAD+ biosynthesis in adipocytes and that PHB1 is a candidate target protein for increased expression of NAMPT to maintain metabolic functions in adipose tissues. ### Competing Interest Statement The authors have declared no competing interest.