title: Multiple functions encoded by the N-terminal PAT domain of adipophilin.
abstract: Adipophilin (ADPH), a member of the perilipin family of cytoplasmic lipid droplet (CLD)-binding proteins, is crucially dependent on triglyceride synthesis for stability. We have used cell lines expressing full-length or N-terminally modified forms of ADPH to investigate the role of the N-terminus in regulating ADPH stability and interactions with CLD. Full-length ADPH was unstable and could not be detected on CLDs unless cultures were incubated with oleic acid (OA) to stimulate triglyceride synthesis, or were treated with MG132 to block proteasomal degradation. By contrast, ADPH lacking amino acids 1-89 (Delta 2,3 ADPH), or N-terminally GFP-tagged full-length ADPH, was stable in the absence of OA or MG132, as was the closely related protein TIP47. However, none of these proteins localized to CLDs unless OA was added to the culture medium. Furthermore, immunofluorescence analysis showed that TIP47 localization to CLDs was prevented by full-length ADPH, but not by Delta 2,3 ADPH. These results suggest that the N-terminal region of ADPH mediates proteasomal degradation and access of TIP47 to the CLD surface and possibly contributes to CLD stability. Chimeras of ADPH and TIP47, generated by swapping their N- and C-terminal halves, showed that these properties are specific to ADPH.
authors.affiliation: Department of Pathology, University of Colorado Denver, Anschutz Medical Campus, Aurora, CO 80045, USA.
authors.forename: David J
authors.initials: DJ
authors.lastname: Orlicky
authors.forename: Greg
authors.initials: G
authors.lastname: Degala
authors.forename: Carrie
authors.initials: C
authors.lastname: Greenwood
authors.forename: Elise S
authors.initials: ES
authors.lastname: Bales
authors.forename: Tanya D
authors.initials: TD
authors.lastname: Russell
authors.forename: James L
authors.initials: JL
authors.lastname: McManaman
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism; D002460:Cell Line; D006801:Humans; D050356:Lipid Metabolism / Q000187:drug effects; D008565:Membrane Proteins; D051379:Mice; D050505:Mutant Proteins / Q000378:metabolism; D019301:Oleic Acid / Q000494:pharmacology; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism*; D000070780:Perilipin-2; D000070796:Perilipin-3; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013329:Structure-Activity Relationship; D013816:Thermodynamics; D014162:Transfection
chemical_list: D002352:Carrier Proteins; D008565:Membrane Proteins; D050505:Mutant Proteins; C000602982:PLIN2 protein, human; D010455:Peptides; D000070780:Perilipin-2; D000070796:Perilipin-3; C000602983:Plin2 protein, mouse; C523440:Plin3 protein, mouse; D011993:Recombinant Fusion Proteins; D019301:Oleic Acid; D046988:Proteasome Endopeptidase Complex