Abnormal energy regulation may contribute to onset and progression of chronic metabolic conditions such as obesity, diabetes mellitus, cardiovascular disease, and cancer, which cause -60%of the world's mortality (rev. in (1). Our lab is interested in how long chain fatty acids (LCFAs) reach nuclei, bind Deroxisomal proliferator receptor a (PPARa), and initiate transcription for energy metabolism or storage. We Hypothesize that liver fatty acid binding protein (L-FABP) transfers and channels LCFAs to nuclei, binding to and initiating PPARa transcriptional activity. Using purified L-FABP and PPARa, L-FABP overexpressed L- cells, L-FABP null mice, and cultured hepatocytes from L-FABP null mice, we propose to:
Aim 1. Determine if the phenotype of L-FABP null mice is consistent with abnormal PPARa regulation. L-FABP null mice share many PPARa null mouse features (inhibition of LCFA oxidation, hypertriglycerid- emia, sex/age-dependent obesity). L-FABP null mice exhibit upregulation of SCP-2/SCP-x, the converse of upregulation of L-FABP in SCP-2/SCP-x null mice. We have bred SCP-x, SCP-2/SCP-x, and L-FABP/SCP- 2/SCP-x null mice to clarify the in vivo role of LCFA/LCFA-CoA binding proteins in PPARa regulation.
Aim 2. Examine the role of L-FABP in targeting LCFAs to the nucleus for interaction with PPARa. L-FABP enhances saturated LCFA targeting to nuclei. We will use novel fluorescent polyunsaturated (n-3, n-5) and branched-chain (phytanic acid) LCFAs, fluorescent-L-FABP (EYFP-, Cy3-, Cy5-) and immunogold EM, to show if: (i) L-FABP cotransports bound LCFAs into nuclei; (ii)LCFAs enhance L-FABP distribution into nuclei; (iii)nuclear targeting of LCFAs depend on relative binding affinities of L-FABP and PPARa.
Aim 3. Resolve molecular interactions of L-FABP with PPARa. Physical and immunological techniques show that L-FABP binds PPARa in vitro. We will examine ligand specificity, conformational responsiveness, and co-activator or co-repressor binding, using: (i) purified proteins in vitro; (ii) FRET between EYFP-L- FABP/ECFP-PPARa or Cy3-L-FABP/Cy5-PPARa in living cells; (iii)immunogold EM; and (iv) fluorescence correlation spectroscopy in living cells (L-cells, primary cultured hepatocytes).
Aim 4. Determine the mechanism of L-FABP-mediated LCFA transfer to PPARa. We will determine if LCFA transfers from L-FABP to PPARa by direct molecular interactions or by LCFA diffusion. These findings will contribute to our basic understanding of how different types of fatty acids may activate a nuclear receptor, and thereby induce transcription of genes directing their energy metabolism and storage. Differences in this nuclear receptor/fatty acid energy regulation could contribute to the pathogenesis of obesity, insulin resistance, type 2 diabetes mellitus, and hyperlipidemic conditions.
| Milligan, Sherrelle; Martin, Gregory G; Landrock, Danilo et al. (2018) Ablating both Fabp1 and Scp2/Scpx (TKO) induces hepatic phospholipid and cholesterol accumulation in high fat-fed mice. Biochim Biophys Acta Mol Cell Biol Lipids 1863:323-338 |
| Martin, Gregory G; Seeger, Drew R; McIntosh, Avery L et al. (2018) Scp-2/Scp-x ablation in Fabp1 null mice differentially impacts hepatic endocannabinoid level depending on dietary fat. Arch Biochem Biophys 650:93-102 |
| Martin, Gregory G; Landrock, Danilo; Chung, Sarah et al. (2017) Fabp1 gene ablation inhibits high-fat diet-induced increase in brain endocannabinoids. J Neurochem 140:294-306 |
| Storey, Stephen M; Huang, Huan; McIntosh, Avery L et al. (2017) Impact of Fabp1/Scp-2/Scp-x gene ablation (TKO) on hepatic phytol metabolism in mice. J Lipid Res 58:1153-1165 |
| McIntosh, Avery L; Storey, Stephen M; Huang, Huan et al. (2017) Sex-dependent impact of Scp-2/Scp-x gene ablation on hepatic phytol metabolism. Arch Biochem Biophys 635:17-26 |
| Landrock, Danilo; Milligan, Sherrelle; Martin, Gregory G et al. (2017) Effect of Fabp1/Scp-2/Scp-x Ablation on Whole Body and Hepatic Phenotype of Phytol-Fed Male Mice. Lipids 52:385-397 |
| Huang, Huan; McIntosh, Avery L; Martin, Gregory G et al. (2016) FABP1: A Novel Hepatic Endocannabinoid and Cannabinoid Binding Protein. Biochemistry 55:5243-55 |
| Huang, Huan; McIntosh, Avery L; Landrock, Kerstin K et al. (2015) Human FABP1 T94A variant enhances cholesterol uptake. Biochim Biophys Acta 1851:946-55 |
| Martin, Gregory G; Landrock, Danilo; Landrock, Kerstin K et al. (2015) Relative contributions of L-FABP, SCP-2/SCP-x, or both to hepatic biliary phenotype of female mice. Arch Biochem Biophys 588:25-32 |
| Klipsic, Devon; Landrock, Danilo; Martin, Gregory G et al. (2015) Impact of SCP-2/SCP-x gene ablation and dietary cholesterol on hepatic lipid accumulation. Am J Physiol Gastrointest Liver Physiol 309:G387-99 |
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