Obesity is a rapidly growing epidemic in the USA and worldwide. While the molecular and cellular mechanism of obesity is incompletely understood, studies have shown that excess adiposity may arise from increased adipogenesis (hyperplasia) and adipocyte size (hypertrophy) . Emerging evidence underscores autophagy as an important mediator of adipogenesis and adiposity. We are interested in the upstream regulator of adipocyte autophagy and how it impacts adipocyte biology.

Given that metabolic stress activates transcription factor FoxO1 in obesity, my dissertation project is designed to depict the role of FoxO1 in adipocyte autophagy and biology. We found that FoxO1 upregulation was concomitant with elevation of autophagy activity during adipogenesis. Inhibition of FoxO1 suppressed autophagy flux and almost completely prevented adipocyte differentiation. For the first time, we found that the kinetics of FoxO1 activation followed a series of sigmoid curves that showed multiple activation-inactivation transitions during adipogenesis. Our study provides critical evidence casting light on the controversy in the literature that either persistent inhibition or activation of FoxO1 suppresses adipogenesis. In addition, we identified two central pathways that FoxO1-mediated autophagy regulated adipocyte biology: (1) to control lipid droplet growth via fat specific protein 27 (FSP27) in adipocytes; and (2) to differentially regulate mitochondrial uncoupling proteins (UCP) that have been implicated in browning of white adipose tissue and redox homeostasis. Mechanistically, FoxO1 appears to induce autophagy through the transcription factor EB (Tfeb), which was previously shown to regulate both autophagosome and lysosome. Chromatin immunoprecipitation assay demonstrated that FoxO1 directly bound to the promoter of Tfeb, and inhibition of FoxO1 attenuated the binding, which resulted in reduced Tfeb expression.

To investigate the role of FoxO1 in vivo, we have developed mouse models to modulate FoxO1 in adipose tissue using an inducible Cre-loxP system. Tamoxifen is widely used to activate the inducible Cre recombinase that spatiotemporally control target gene expression in animal models, but it was unclear whether tamoxifen itself may affect adiposity and confounds phenotyping. Part of my dissertation work was to address this important question. We found that tamoxifen led to reduced fat mass independent of Cre, which lasted for 4-5 weeks. Mechanistically, Tamoxifen induced reactive oxygen species (ROS) and augmented apoptosis. Our data reveals a critical period of recovery following tamoxifen treatment in the study of inducible knockout mice.

Together, my dissertation work demonstrates FoxO1 as a critical regulator of adipocyte autophagy via Tfeb during adipogenesis. FoxO1-mediated autophagy controls FSP27, lipid droplet growth, and mitochondrial uncoupling proteins. Further study of FoxO1-autophagy axis in obese subjects is of physiological significance, and the investigation is under way.