Analysis of interaction partners for perilipin and ADRP on lipid droplets.
Despite the critical roles of intracellular lipid droplets (LDs) in lipid storage and metabolism, little is known about the molecular mechanisms of their functions. Several protein components associated with the surface of LDs have been identified. A major one is perilipin in adipocytes and steroidogenic cells, whereas ADRP in most other cell types. They are loosely grouped as a small protein family sharing a common N-terminal motif, called the PAT domain. Perilipin regulates the breakdown of triacylglycerol in LDs via its phosphorylation. ADRP is characterized as a fatty acid binding protein and involved in lipid uptake and LD formation. For examining the functions of perilipin and ADRP at the molecular level, we performed yeast two-hybrid screening in this study, to find their functional partners. We identified CGI-58, a product of the causal gene of Chanarin-Dorfman syndrome (CDS), as an interactor for both perilipin and ADRP. Specific interaction between CGI-58 and perilipin was confirmed in a GST-pulldown assay and supported by fluorescence microscopic analyses. We further demonstrated that CGI-58 is principally located at the surface of LDs in 3T3-L1 cells, together with perilipin, and its expression is upregulated upon stimulation for adipocyte differentiation. Other than CGI-58, we also identified in yeast two-hybrid screening HSP86 and D52 tumor proteins as binding partners of perilipin, and IRG-47 of ADRP. These factors might be cooperated with perilipin and ADRP, and hence involved in membrane dynamics of LDs as well as the regulation of lipolysis on the surface of LDs.
Graduate School of Life Science, Himeji Institute of Technology, University of Hyogo, Kamigori, Hyogo 678-1297, Japan.
Tomohiro
T
Yamaguchi
Naoto
N
Omatsu
Asami
A
Omukae
Takashi
T
Osumi
D051103:1-Acylglycerol-3-Phosphate O-Acyltransferase; D041721:3T3-L1 Cells; D017667:Adipocytes / Q000502:physiology; D000818:Animals; D002352:Carrier Proteins; D002454:Cell Differentiation; D004950:Esterases / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism; D050356:Lipid Metabolism; D008066:Lipolysis; D008565:Membrane Proteins / Q000096:biosynthesis / Q000378:metabolism*; D051379:Mice; D009363:Neoplasm Proteins / Q000378:metabolism; D000070778:Perilipin-1; D000070780:Perilipin-2; D010750:Phosphoproteins / Q000096:biosynthesis / Q000378:metabolism*; D051381:Rats; D020798:Two-Hybrid System Techniques; D015003:Yeasts / Q000235:genetics
D002352:Carrier Proteins; D018841:HSP90 Heat-Shock Proteins; C107835:Hspca protein, mouse; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D000070778:Perilipin-1; D000070780:Perilipin-2; D010750:Phosphoproteins; C000602983:Plin2 protein, mouse; C000602984:Plin2 protein, rat; D051103:1-Acylglycerol-3-Phosphate O-Acyltransferase; C492986:Abhd5 protein, mouse; D004950:Esterases