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Membrane attachment and structure models of lipid storage droplet protein 1.

ABSTRACT: Neutral lipid triglycerides, a main reserve for fat and energy, are stored in organelles called lipid droplets. The storage and release of triglycerides are actively regulated by several proteins specific to the droplet surface, one of which in insects is PLIN1. PLIN1 plays a key role in the activation of triglyceride hydrolysis upon phosphorylation. However, the structure of PLIN1 and its relation to functions remain elusive due to its insolubility and crystallization difficulty. Here we report the first solid-state NMR study on the Drosophila melanogaster PLIN1 in combination with molecular dynamics simulation to show the structural basis for its lipid droplet attachment. NMR spin diffusion experiments were consistent with the predicted membrane attachment motif of PLIN1. The data indicated that PLIN1 has close contact with the terminal methyl groups of the phospholipid acyl chains. Structure models for the membrane attachment motif were generated based on hydrophobicity analysis and NMR membrane insertion depth information. Simulated NMR spectra from a trans-model agreed with experimental spectra. In this model, lipids from the bottom leaflet were very close to the surface in the region enclosed by membrane attachment motif. This may imply that in real lipid droplet, triglyceride molecules might be brought close to the surface by the same mechanism, ready to leave the droplet in the event of lipolysis. Juxtaposition of triglyceride lipase structure to the trans-model suggested a possible interaction of a conserved segment with the lipase by electrostatic interactions, opening the lipase lid to expose the catalytic center.

SUBMITTER: Lin P

PROVIDER: S-EPMC3918908 | biostudies-literature | 2014 Mar

REPOSITORIES: biostudies-literature

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Membrane attachment and structure models of lipid storage droplet protein 1.

Lin Penghui P Chen Xiao X Moktan Hem H Arrese Estela L EL Duan Lian L Wang Liying L Soulages Jose L JL Zhou Donghua H DH

Biochimica et biophysica acta 20131212 3

Neutral lipid triglycerides, a main reserve for fat and energy, are stored in organelles called lipid droplets. The storage and release of triglycerides are actively regulated by several proteins specific to the droplet surface, one of which in insects is PLIN1. PLIN1 plays a key role in the activation of triglyceride hydrolysis upon phosphorylation. However, the structure of PLIN1 and its relation to functions remain elusive due to its insolubility and crystallization difficulty. Here we report ...[more]

PMID: 24333382

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Project description:ObjectiveStorage of triglycerides in lipid droplets is governed by a set of lipid droplet-associated proteins. One of these lipid droplet-associated proteins, hypoxia-inducible lipid droplet-associated (HILPDA), was found to impair lipid droplet breakdown in macrophages and cancer cells by inhibiting adipose triglyceride lipase. Here, we aimed to better characterize the role and mechanism of action of HILPDA in hepatocytes.MethodsWe performed studies in HILPDA-deficient and HILPDA-overexpressing liver cells, liver slices, and mice. The functional role and physical interactions of HILPDA were investigated using a variety of biochemical and microscopic techniques, including real-time fluorescence live-cell imaging and Förster resonance energy transfer-fluorescence lifetime imaging microscopy (FRET-FLIM).ResultsLevels of HILPDA were markedly induced by fatty acids in several hepatoma cell lines. Hepatocyte-specific deficiency of HILPDA in mice modestly but significantly reduced hepatic triglycerides in mice with non-alcoholic steatohepatitis. Similarly, deficiency of HILPDA in mouse liver slices and primary hepatocytes reduced lipid storage and accumulation of fluorescently-labeled fatty acids in lipid droplets, respectively, which was independent of adipose triglyceride lipase. Fluorescence microscopy showed that HILPDA partly colocalizes with lipid droplets and with the endoplasmic reticulum, is especially abundant in perinuclear areas, and mainly associates with newly added fatty acids. Real-time fluorescence live-cell imaging further revealed that HILPDA preferentially localizes to lipid droplets that are being remodeled. Overexpression of HILPDA in liver cells increased the activity of diacylglycerol acyltransferases (DGAT) and DGAT1 protein levels, concurrent with increased lipid storage. Confocal microscopy coupled to FRET-FLIM analysis demonstrated that HILPDA physically interacts with DGAT1 in living liver cells. The stimulatory effect of HILPDA on lipid storage via DGAT1 was corroborated in adipocytes.ConclusionsOur data indicate that HILPDA physically interacts with DGAT1 and increases DGAT activity. Our findings suggest a novel regulatory mechanism by which fatty acids promote triglyceride synthesis and storage.

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Membrane attachment and structure models of lipid storage droplet protein 1.

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Membrane attachment and structure models of lipid storage droplet protein 1.

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