Endoplasmic Reticulum Degradation–Enhancing α‐Mannosidase–like Protein 1 Targets Misfolded HLA–B27 Dimers for Endoplasmic Reticulum–Associated Degradation

Abstract

Objective HLA-B27 forms misfolded heavy chain dimers, which may predispose individuals to inflammatory arthritis by inducing endoplasmic reticulum (ER) stress and the unfolded protein response (UPR). We wanted to define the role of the UPR induced ER associated degradation (ERAD) pathway in the disposal of HLA-B27 dimeric conformers. Methods HeLa cell lines expressing only two copies of a carboxy terminally Sv5 tagged HLA-B27 were generated. The ER stress induced EDEM1 protein was over expressed by transfection and dimer levels monitored by immunoblotting. EDEM1, the UPR associated transcription factor XBP-1, the E3 ubiquitin ligase HRD1, the degradation associated derlin 1 and 2 proteins were inhibited by either short hairpin RNA or dominant negative mutants. The UPR associated ERAD of HLA-B27 was confirmed using ER stress inducing pharamacological agents in kinetic and pulse chase assays. Results We demonstrate that UPR induced machinery can target HLA-B27 dimers, and that dimer formation can be controlled by alterations to expression levels of components of the UPR induced ERAD pathway. HLA-B27 dimers and misfolded MHC class I monomeric molecules were detected bound to EDEM1, with overexpression of EDEM1 inhibiting HLA-B27 dimer formation. EDEM1 inhibition resulted in upregulation of HLA-B27 dimers, whilst UPR induced ERAD of dimers was prevented in the absence of EDEM1. HLA-B27 dimer formation was also enhanced in the absence of XBP-1, HRD1 and derlin1/2. Conclusion The UPR ERAD pathway as described here can dispose of HLA-B27 dimers and presents a potential novel therapeutic target for the modulation of HLA-B27 associated inflammatory disease.

The Role of Autophagy in the Degradation of Misfolded HLA–B27 Heavy Chains

“…Proteasome inhibition with bortezomib resulted in a 1.5-fold increase and a 1.7-fold increase in dimeric and oligomeric forms of HLA-B27, respectively. This confirms previous findings that misfolded HLA-B27 undergoes ERAD in human cells (4,21,22) and extends these findings to HLA-B27-transgenic rats.…”

“…It will be of interest to compare the relative contributions of ERAD and autophagy to ERQC of HLA-B27 in other cells derived from HLA-B2-transgenic rats. HLA-B27 dimers form immediately after heavy chains are synthesized in the ER (5), exhibit a half-life exceeding 8 hours (22), and can be found in many cells at steady state (5,18,22), which suggests that they are not efficiently eliminated and may be resistant to ERAD. This is in sharp contrast to many proteins that misfold in the ER and are efficiently degraded by ERAD, including major histocompatibility complex class I heavy chains produced in the absence of b 2 m or peptide (21,44).…”

Evolution of protein N-glycosylation process in Golgi apparatus which shapes diversity of protein N-glycan structures in plants, animals and fungi

“…Clade V is distant to any other clades, comprises genes in all three kingdoms (Supplementary Figure S6). In this clade, three human genes encode enzymes playing roles as ER degradation enhancers (EDEMs, ENSG00000134109, ENSG00000088298 and ENSG00000116406)484950. Also, the yeast and Arabidopsis genes are involved in ERAD4651.…”

The relation between ER stress and HLA-B27 misfolding

“…HLA Class I molecules especially HLA-B27 exhibit unique protein folding status that can conflict with ER quality control mechanism and have tendency to switch to misfolded state [23]. Other two unique features of HLA-B27, besides a tendency to misfolded state, such as peptide binding specificity and ability for forming heavy chain homodimers in cell surface recycling, three are also considered to diversify it from among all other MHC Class I molecules [24][25].…”

“…In all SpAs, MHC genes have been presented with 40-50% genetic tendency and especially approximately 30% of the tendency has been related with HLA-B27 [31][32][33]. Although, HLA-B27 expression ratio in AS patients is about 90-95%, the pathogenesis of AS with HLA-B27 association has not to be enlighten, yet [23]. The possible role of HLA-B27 in pathogenesis of SpAs has several difficulties to understand precisely, since genetic and environmental elements of autoimmune diseases presentation that are known as complex and multifactorial disorder, increase the heterogeneity [34].…”