title: BiP prevents rod opsin aggregation.
abstract: Mutations in rod opsin-the light-sensitive protein of rod cells-cause retinitis pigmentosa. Many rod opsin mutations lead to protein misfolding, and therefore it is important to understand the role of molecular chaperones in rod opsin biogenesis. We show that BiP (HSPA5) prevents the aggregation of rod opsin. Cleavage of BiP with the subtilase cytotoxin SubAB results in endoplasmic reticulum (ER) retention and ubiquitylation of wild-type (WT) rod opsin (WT-green fluorescent protein [GFP]) at the ER. Fluorescence recovery after photobleaching reveals that WT-GFP is usually mobile in the ER. By contrast, depletion of BiP activity by treatment with SubAB or coexpression of a BiP ATPase mutant, BiP(T37G), decreases WT-GFP mobility to below that of the misfolding P23H mutant of rod opsin (P23H-GFP), which is retained in the ER and can form cytoplasmic ubiquitylated inclusions. SubAB treatment of P23H-GFP-expressing cells decreases the mobility of the mutant protein further and leads to ubiquitylation throughout the ER. Of interest, BiP overexpression increases the mobility of P23H-GFP, suggesting that it can reduce mutant rod opsin aggregation. Therefore inhibition of BiP function results in aggregation of rod opsin in the ER, which suggests that BiP is important for maintaining the solubility of rod opsin in the ER.
authors.affiliation: UCL Institute of Ophthalmology, London, United Kingdom.
authors.forename: Dimitra
authors.initials: D
authors.lastname: Athanasiou
authors.forename: Maria
authors.initials: M
authors.lastname: Kosmaoglou
authors.forename: Naheed
authors.initials: N
authors.lastname: Kanuga
authors.forename: Sergey S
authors.initials: SS
authors.lastname: Novoselov
authors.forename: Adrienne W
authors.initials: AW
authors.lastname: Paton
authors.forename: James C
authors.initials: JC
authors.lastname: Paton
authors.forename: J Paul
authors.initials: JP
authors.lastname: Chapple
authors.forename: Michael E
authors.initials: ME
authors.lastname: Cheetham
mesh_terms: D015153:Blotting, Western; D045744:Cell Line, Tumor; D003593:Cytoplasm / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D029968:Escherichia coli Proteins / Q000378:metabolism / Q000494:pharmacology; D036681:Fluorescence Recovery After Photobleaching; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D018613:Microscopy, Confocal; D009154:Mutation; D011485:Protein Binding / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D017299:Rod Opsins / Q000235:genetics / Q000378:metabolism*; D013381:Subtilisins / Q000378:metabolism / Q000494:pharmacology; D014162:Transfection; D054875:Ubiquitination / Q000187:drug effects; D056811:Unfolded Protein Response / Q000187:drug effects
chemical_list: D000091342:Endoplasmic Reticulum Chaperone BiP; D029968:Escherichia coli Proteins; C000717247:HSPA5 protein, human; D006360:Heat-Shock Proteins; D017299:Rod Opsins; D049452:Green Fluorescent Proteins; D013381:Subtilisins; C527234:subtilase cytotoxin, E coli