title: 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). This study was undertaken 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 2 copies of a carboxy-terminally Sv5-tagged HLA-B27 were generated. The ER stress-induced protein ER degradation-enhancing α-mannosidase-like protein 1 (EDEM1) was overexpressed by transfection, and dimer levels were monitored by immunoblotting. EDEM1, the UPR-associated transcription factor X-box binding protein 1 (XBP-1), the E3 ubiquitin ligase hydroxymethylglutaryl-coenzyme A reductase degradation 1 (HRD1), and the degradation-associated proteins derlin 1 and derlin 2 were inhibited using either short hairpin RNA or dominant-negative mutants. The UPR-associated ERAD of HLA-B27 was confirmed using ER stress-inducing pharamacologic agents in kinetic and pulse chase assays.
  
  
  RESULTS
  We demonstrated 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 major histocompatibility complex class I monomeric molecules bound to EDEM1 were detected, and overexpression of EDEM1 led to inhibition of HLA-B27 dimer formation. EDEM1 inhibition resulted in up-regulation of HLA-B27 dimers, while 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 derlins 1 and 2.
  
  
  CONCLUSIONS
  The present findings indicate that the UPR ERAD pathway can dispose of HLA-B27 dimers, thus presenting a potential novel therapeutic target for modulation of HLA-B27-associated inflammatory disease.
authors.affiliation: University College London and University of East London, London, UK.
authors.forename: David B
authors.initials: DB
authors.lastname: Guiliano
authors.forename: Helen
authors.initials: H
authors.lastname: Fussell
authors.forename: Izabela
authors.initials: I
authors.lastname: Lenart
authors.forename: Edward
authors.initials: E
authors.lastname: Tsao
authors.forename: Darren
authors.initials: D
authors.lastname: Nesbeth
authors.forename: Adam J
authors.initials: AJ
authors.lastname: Fletcher
authors.forename: Elaine C
authors.initials: EC
authors.lastname: Campbell
authors.forename: Nasim
authors.initials: N
authors.lastname: Yousaf
authors.forename: Sarah
authors.initials: S
authors.lastname: Williams
authors.forename: Susana
authors.initials: S
authors.lastname: Santos
authors.forename: Amy
authors.initials: A
authors.lastname: Cameron
authors.forename: Greg J
authors.initials: GJ
authors.lastname: Towers
authors.forename: Paul
authors.initials: P
authors.lastname: Kellam
authors.forename: Daniel N
authors.initials: DN
authors.lastname: Hebert
authors.forename: Keith G
authors.initials: KG
authors.lastname: Gould
authors.forename: Simon J
authors.initials: SJ
authors.lastname: Powis
authors.forename: Antony N
authors.initials: AN
authors.lastname: Antoniou
mesh_terms: D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000502:physiology; D004721:Endoplasmic Reticulum / Q000502:physiology*; D060746:Endoplasmic Reticulum-Associated Degradation / Q000502:physiology*; D015796:HLA-B27 Antigen / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000502:physiology*; D017510:Protein Folding; D034741:RNA, Small Interfering / Q000494:pharmacology; D000071701:Regulatory Factor X Transcription Factors; D015398:Signal Transduction / Q000502:physiology; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000502:physiology; D044767:Ubiquitin-Protein Ligases / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000502:physiology; D000071717:X-Box Binding Protein 1
chemical_list: C487709:DERL1 protein, human; C513305:DERL2 protein, human; D004268:DNA-Binding Proteins; C486654:EDEM1 protein, human; D015796:HLA-B27 Antigen; D008565:Membrane Proteins; D034741:RNA, Small Interfering; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors; D000071717:X-Box Binding Protein 1; C000605543:XBP1 protein, human; C482345:SYVN1 protein, human; D044767:Ubiquitin-Protein Ligases