The processes controlling targeting of glucose transporters to apical and basolateral membranes of polarized cells are complex and not-well understood. We have engineered SGLT1 and GLUT4 constructs linked to fluorescent proteins to highlight the differences in transporter expression and trafficking, in real time, in different cell types. Activity was assessed in parallel using a FRET glucose sensor. In COS cells and HEK cells, SGLT1 was distributed between the plasma membrane and intracellular compartments, but there was little expression in CHO cells. Trafficking was investigated using the lysosome inhibitors NH4Cl (10 mmol/L) and chloroquine (150 μmol/L) and the proteasome inhibitors MG-262 (1 μmol/L) and lactacystin (5 μmol/L). Lysosome inhibitors caused SGLT1 accumulation into intracellular bodies, whereas proteasome inhibitors induced SGLT1 accumulation in the plasma membrane, even in CHO cells. Our data suggest that a fraction of SGLT1 is rapidly degraded by lysosomes and never reached the plasma membrane; another fraction reaches the membrane and is subsequently degraded by lysosomes following internalization. The latter process is regulated by the ubiquitin/proteasome pathway, acting at a late stage of the lysosomal pathway. Using the cholesterol inhibitor MβCD (3 mmol/L), a dominant negative dynamin (K44A) and caveolin, we showed that SGLT1 internalization is lipid raft-mediated, but caveolin-independent. In contrast, GLUT4 internalization is dynamin-dependent, but cholesterol-independent. The physiological relevance of these data is discussed in terms of differential membrane compartmentalization of the transporters and expression under stress conditions.

FXR agonists are used to treat non-alcoholic fatty liver disease (NAFLD), in part because they reduce hepatic lipids. Here, we show that FXR activation with the FXR agonist GSK2324 controls hepatic lipids via reduced absorption and selective decreases in fatty acid synthesis. Using comprehensive lipidomic analyses, we show that FXR activation in mice or humans specifically reduces hepatic levels of mono- and polyunsaturated fatty acids (MUFA and PUFA). Decreases in MUFA are due to FXR-dependent repression of Scd1, Dgat2, and Lpin1 expression, which is independent of SHP and SREBP1c. FXR-dependent decreases in PUFAs are mediated by decreases in lipid absorption. Replenishing bile acids in the diet prevented decreased lipid absorption in GSK2324-treated mice, suggesting that FXR reduces absorption via decreased bile acids. We used tissue-specific FXR KO mice to show that hepatic FXR controls lipogenic genes, whereas intestinal FXR controls lipid absorption. Together, our studies establish two distinct pathways by which FXR regulates hepatic lipids.

Dysregulation of lipid homeostasis is a precipitating event in the pathogenesis and progression of hepatosteatosis and metabolic syndrome. These conditions are highly prevalent in developed societies and currently have limited options for diagnostic and therapeutic intervention. Here, using a proteomic and lipidomic-wide systems genetic approach, we interrogated lipid regulatory networks in 107 genetically distinct mouse strains to reveal key insights into the control and network structure of mammalian lipid metabolism. These include the identification of plasma lipid signatures that predict pathological lipid abundance in the liver of mice and humans, defining subcellular localization and functionality of lipid-related proteins, and revealing functional protein and genetic variants that are predicted to modulate lipid abundance. Trans-omic analyses using these datasets facilitated the identification and validation of PSMD9 as a previously unknown lipid regulatory protein. Collectively, our study serves as a rich resource for probing mammalian lipid metabolism and provides opportunities for the discovery of therapeutic agents and biomarkers in the setting of hepatic lipotoxicity.