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Lipid droplets, perilipins and cytokeratins--unravelled liaisons in epithelium-derived cells.

ABSTRACT: Lipid droplets (LDs) are spherical accumulations of apolar lipids and other hydrophobic substances and are generally surrounded by a thin cortical layer of specific amphiphilic proteins (APs). These APs segregate the LDs from the mostly polar components of the cytoplasm. We have studied LDs in epithelium-derived cell cultures and in particular characterized proteins from the perilipin (PLIN) gene family - in mammals consisting of the proteins Perilipin, Adipophilin, TIP47, S3-12 and MLDP/OXPAT (PLIN 1-5). Using a large number of newly generated and highly specific mono- and polyclonal antibodies specific for individual APs, and using improved LD isolation methods, we have enriched and characterized APs in greater detail and purity. The majority of lipid-AP complexes could be obtained in the top layer fractions of density gradient centrifugation separations of cultured cells, but APs could also be detected in other fractions within such separations. The differently sized LD complexes were analyzed using various biochemical methods and mass spectrometry as well as immunofluorescence and electron- in particular immunoelectron-microscopy. Moreover, by immunoprecipitation, protein-protein binding assays and by immunoelectron microscopy we identified a direct linkage between LD-binding proteins and the intermediate-sized filaments (IF) cytokeratins 8 and 18 (also designated as keratins K8 and K18). Specifically, in gradient fractions of higher density supposedly containing small LDs, we received as co-precipitations cytidylyl-, palmitoyl- and cholesterol transferases and other specific enzymes involved in lipid metabolism. So far, common proteomic studies have used LDs from top layer fractions only and did not report on these transferases and other enzymes. In addition to findings of short alternating hydrophobic/hydrophilic segments within the PLIN protein family, we propose and discuss a model for the interaction of LD-coating APs with IF proteins.

SUBMITTER: Heid H

PROVIDER: S-EPMC3660578 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Lipid droplets, perilipins and cytokeratins--unravelled liaisons in epithelium-derived cells.

Heid Hans H Rickelt Steffen S Zimbelmann Ralf R Winter Stefanie S Schumacher Heiderose H Dörflinger Yvette Y

PloS one 20130521 5

Lipid droplets (LDs) are spherical accumulations of apolar lipids and other hydrophobic substances and are generally surrounded by a thin cortical layer of specific amphiphilic proteins (APs). These APs segregate the LDs from the mostly polar components of the cytoplasm. We have studied LDs in epithelium-derived cell cultures and in particular characterized proteins from the perilipin (PLIN) gene family - in mammals consisting of the proteins Perilipin, Adipophilin, TIP47, S3-12 and MLDP/OXPAT ( ...[more]

PMID: 23704888

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Project description:ObjectivesPolyunsaturated fatty acids (PUFAs) are structural components of membrane phospholipids and precursors of oxygenated lipid mediators with diverse functions, including the control of cell growth, inflammation and tumourigenesis. However, the molecular pathways that control the availability of PUFAs for lipid mediator production are not well understood. Here, we investigated the crosstalk of three pathways in the provision of PUFAs for lipid mediator production: (i) secreted group X phospholipase A2 (GX sPLA2) and (ii) cytosolic group IVA PLA2 (cPLA2α), both mobilizing PUFAs from membrane phospholipids, and (iii) adipose triglyceride lipase (ATGL), which mediates the degradation of triacylglycerols (TAGs) stored in cytosolic lipid droplets (LDs).MethodsWe combined lipidomic and functional analyses in cancer cell line models to dissect the trafficking of PUFAs between membrane phospholipids and LDs and determine the role of these pathways in lipid mediator production, cancer cell proliferation and tumour growth in vivo.ResultsWe demonstrate that lipid mediator production strongly depends on TAG turnover. GX sPLA2 directs ω-3 and ω-6 PUFAs from membrane phospholipids into TAG stores, whereas ATGL is required for their entry into lipid mediator biosynthetic pathways. ATGL controls the release of PUFAs from LD stores and their conversion into cyclooxygenase- and lipoxygenase-derived lipid mediators under conditions of nutrient sufficiency and during serum starvation. In starving cells, ATGL also promotes the incorporation of LD-derived PUFAs into phospholipids, representing substrates for cPLA2α. Furthermore, we demonstrate that the built-up of TAG stores by acyl-CoA:diacylglycerol acyltransferase 1 (DGAT1) is required for the production of mitogenic lipid signals that promote cancer cell proliferation and tumour growth.ConclusionThis study shifts the paradigm of PLA2-driven lipid mediator signalling and identifies LDs as central lipid mediator production hubs. Targeting DGAT1-mediated LD biogenesis is a promising strategy to restrict lipid mediator production and tumour growth.

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Lipid droplets, perilipins and cytokeratins--unravelled liaisons in epithelium-derived cells.

Publications

Lipid droplets, perilipins and cytokeratins--unravelled liaisons in epithelium-derived cells.

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