CGI-58 plays an important role in lipid homeostasis with poorly defined mechanisms. Here we show that the protein functions as a novel lysophosphatidylglcyerol acyltransferase and promotes autophagy. The acyltransferase activity of CGI-58 requires a unique 17-amino acid sequence present in perilipin B, which also serves as the binding site for a CGI-58 substrate.

A peptide corresponding to this sequence, when used as a ligand to CGI-58, inhibits its acyltransferase activity and promotes dispersion of CGI-58 from lipid droplets. We identified the site of interaction by a QuikChange(r) site-directed mutagenesis kit (Stratagene) with forward (5′-CCTGATTTCAAGCGGAAGTACGACTCTCTTTGAAGATGACACG-3′) and reverse (5′-CGTGTCATCTTCAAACATAGACTCGTACTTCCGCTTGCAGG-3′) mutagenic primers. A co-immunoprecipitation experiment with a human perilipin B peptide revealed that the CGI-58 binding interface with perilipin is located within a region encompassing amino acids 382 to 405 of the protein. Find out bosterbio.com

CGI-58 Antibody: Applications and Research Benefits

The removal of the sequence surrounding the binding site reduced the affinity of CGI-58 for perilipin but not for the substrate oligosaccharide. This suggests that the residues bind to an intermediate conformation of perilipin B or that the intact 48-amino acid sequence is required to fold into a binding pocket that exposes a smaller binding surface.

CGI-58 also regulates LC3-mediated mitochondrial autophagy in part by promoting apoptotic vesicle formation. In adenovirus-infected C2C12 cells stably expressing an LC3-GFP fusion protein, overexpression of CGI-58 resulted in increased mitophagosome accumulation in response to bafilomycin A1 treatment. Moreover, a CGI-58-GFP chimera demonstrated diffuse distribution in undifferentiated 3T3-L1 preadipocytes, but clearly localized to the surfaces of lipid droplets in differentiated adipocytes.