Calnexin Improves the Folding Efficiency of Mutant Rhodopsin in the Presence of Pharmacological Chaperone 11-cis-Retinal

Abstract

The lectin chaperone calnexin (Cnx) is important for quality control of glycoproteins, and the chances of correct folding of a protein increase the longer the protein interacts with Cnx. Mutations in glycoproteins increase their association with Cnx, and these mutant proteins are retained in the endoplasmic reticulum. However, until now, the increased interaction with Cnx was not known to increase the folding of mutant glycoproteins. Because many human diseases result from glycoprotein misfolding, a Cnx-assisted folding of mutant glycoproteins could be beneficial. Mutations of rhodopsin, the glycoprotein pigment of rod photoreceptors, cause misfolding resulting in retinitis pigmentosa. Despite the critical role of Cnx in glycoprotein folding, surprisingly little is known about its interaction with rhodopsin or whether this interaction could be modulated to increase the folding of mutant rhodopsin. Here, we demonstrate that Cnx preferentially associates with misfolded mutant opsins associated with retinitis pigmentosa. Furthermore, the overexpression of Cnx leads to an increased accumulation of misfolded P23H opsin but not the correctly folded protein. Finally, we demonstrate that increased levels of Cnx in the presence of the pharmacological chaperone 11-cis-retinal increase the folding efficiency and result in an increase in correct folding of mutant rhodopsin. These results demonstrate that misfolded rather than correctly folded rhodopsin is a substrate for Cnx and that the interaction between Cnx and mutant, misfolded rhodopsin, can be targeted to increase the yield of folded mutant protein.

Accumulation of the Raf-1 Kinase Inhibitory Protein (Rkip) Is Associated with Cep290-mediated Photoreceptor Degeneration in Ciliopathies

“…In these disorders, the mutant protein undergoes misfolding and forms toxic aggregates in the cytosol (58,59). We provide evidence for the aberrant accumulation of a protein in photoreceptors that seems to function downstream of the mutated disease protein Cep290.…”

AAV Delivery of Wild-Type Rhodopsin Preserves Retinal Function in a Mouse Model of Autosomal Dominant Retinitis Pigmentosa

“…We found that an increase in rhodopsin expression was damaging to wild-type photoreceptors, but was protective to photoreceptors bearing the P23H transgene on the same background. However, the level of rhodopsin protein in the P23H transgenic mice is much lower than that in C57Bl/6 mice (Noorwez et al, 2009). As even wild-type opsin can misfold and aggregate, it is not surprising that a little extra is too much.…”

“…To determine if overexpression of rhodopsin from viral delivery damaged the retinas of wild-type mice, we injected AAV-RHO301 in C57BL/6J mice, which accumulate more rhodopsin than the P23H transgenic mice (Noorwez et al, 2009). Proteins were also extracted from wild-type C57BL/6J mice retinas 1 month after subretinal injection, and expression of rhodopsin was detected by anti-rhodopsin monoclonal antibody 1D4.…”

The co-chaperone and reductase ERdj5 facilitates rod opsin biogenesis and quality control

“…6C), as well as a global increase of the ∼28-, ∼36- and ∼45–60-kDa species of Rho(P23H), which correspond to an N-terminal truncated protein, the EndoH-sensitive glycoform and dimer, respectively (Fig. 6D) (17,29,47). These findings suggest that silencing of ERdj5 can affect the traffic of WT rod opsin and can increase accumulation of P23H in the ER, thus making it more prone to aggregation.…”