Lipid transport in the intestine

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The proximal intestinal Fatty Acid Binding Proteins (FABPs), FABP1 (aka liver FABP) and FABP2 (aka intestinal FABP) have long been thought to function in dietary lipid transport and processing in the enterocyte. Our previous work at the biophysical, cellular, and animal levels has shown clearly that these two coexpressed proteins, both of which bind long chain FA, have distinct functions. While both are involved in intestinal lipid uptake and assimilation, albeit in different ways, they each appear to also function uniquely in regulating systemic energy homeostasis. In particular, whereas high fat-fed FABP1-/- mice are hyperphagic and become markedly obese, FABP2-/- mice remain lean. Since FABP1 is highly expressed in both intestine and liver, we engineered an intestine-specific FABP1 knockout. We find that the obese phenotype is recapitulated in the FABP1int-/- mouse, underscoring an important role for intestinal FABPs in whole body energy metabolism. Unexpectedly, single cell transcriptomics has revealed that both FABP1 and FABP2 are expressed in enteroendocrine cells (EEC), and indeed we find alterations in plasma GIP and GLP-1 levels in FABP1-/- and FABP2-/- mice. Moreover, we and others have shown that in addition to FA, FABP1 and FABP2 bind endocannabinoids (EC). Preliminary results also identify other bioactive compounds as novel ligands for these proteins. Thus, we will explore the hypothesis that intestinal FABPs, potentially via EECs, bind specific ligands derived from the diet or their metabolites, and thereby function as nutrient sensors to effect downstream modulation of systemic energy balance. We will use multiple approaches--biochemical, physiological, and structure-based in silico methods--to begin to address this hypothesis in these Specific Aims: 1) To define the complete lipidomic binding profiles of FABP1 and FABP2, and determine the effects of known and newly identified ligands on intestinal epithelial cell transcriptomics and hormonal secretion. Studies will use in vitro lipidomic biochemical profiling as well as intestinal organoid cultures from FABP2 null and FABP1int-/- mice incubated with known and newly identified ligands to identify pathways modulated by specific lipid-FABP interactions; 2) To identify the roles of FABP1 and FABP2 in enteroendocrine signaling, and how this may regulate systemic metabolic effects. These studies address the hypothesis that FABP1 and FABP2, acting through binding of EC and/or other lipids, modulate EEC signaling and GIP and GLP-1 release into the blood, affecting body weight gain. Studies will use mice with EEC-specific ablation of FABP1 and FABP2, and isolated EECs from WT and FABP null mice. 3) To determine whether specific FABP1 and FABP2 ligands tailor the protein surface for sensing and signaling. We will analyze protein tertiary structures derived from NMR spectroscopy using known substrates and newly identified ligands. We hypothesize that by interacting with specific ligands, FABPs will display uniquely altered surface properties which promote interactions with different downstream effectors, i.e. ligand-dependent differences in FABP-protein interactions.