Calnexin and ERp57 facilitate the assembly of the neonatal Fc receptor for IgG with beta 2-microglobulin in the endoplasmic reticulum.
The neonatal FcR (FcRn) consists of an MHC class I-like H chain in noncovalent association with beta(2)-microglobulin (beta(2)m). The proper folding of FcRn in the endoplasmic reticulum is essential for FcRn function. Using a low stringency immunoprecipitation of human FcRn, we observed the coprecipitation of an 88-kDa band. Mass spectrometry analysis revealed that this band was identical with calnexin (CNX). This association was verified by Western blotting the CNX or FcRn immunoprecipitates with either an anti-FcRn or anti-CNX Ab. In the beta(2)m-null FO-1 cell transfected with FcRn H chain alone or both FcRn H chain and beta(2)m, CNX bound to the FcRn H chain before the FcRn H chain association with beta(2)m. However, calreticulin only bound to the FcRn H chain-beta(2)m complex. Furthermore, the thiol oxidoreductase ERp57 was detected in FcRn-CNX complexes, suggesting its role in disulfide bond formation of the FcRn H chain. Removal of the N-linked glycosylation site from the FcRn H chain resulted in a decreased association of the FcRn H chain for beta(2)m. However, the absence of CNX did not significantly affect FcRn assembly as defined by the ability of FcRn to bind IgG and exit to the cell surface. This suggests that other chaperones compensate for the function of CNX in FcRn assembly. In addition, we found that tapasin and TAP were not involved in FcRn assembly, as shown by coimmunoprecipitation in THP-1 cells and IgG-binding assays in 721.220 (tapasin-deficient) and 721.174 (TAP-deficient) cells transfected with FcRn. These findings show the importance of chaperones in FcRn assembly.
Laboratory of Immunology, Virginia-Maryland Regional College of Veterinary Medicine, University of Maryland, 8075 Greenmead Drive, College Park, MD 20742, USA. xzhu@umd.edu
Xiaoping
X
Zhu
Junmin
J
Peng
Daohong
D
Chen
Xindong
X
Liu
Lilin
L
Ye
Hideki
H
Iijima
Kumar
K
Kadavil
Wayne I
WI
Lencer
Richard S
RS
Blumberg
D000595:Amino Acid Sequence; D037281:Calnexin / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology*; D037282:Calreticulin / Q000378:metabolism; D002236:Carbohydrate Conformation; D045744:Cell Line, Tumor; D004220:Disulfides / Q000737:chemistry; D004721:Endoplasmic Reticulum / Q000276:immunology* / Q000378:metabolism*; D019073:HT29 Cells; D006367:HeLa Cells; D006360:Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D015395:Histocompatibility Antigens Class I / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism*; D007231:Infant, Newborn; D007535:Isomerases / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D008969:Molecular Sequence Data; D011485:Protein Binding / Q000276:immunology; D011487:Protein Conformation; D019704:Protein Disulfide-Isomerases; D011499:Protein Processing, Post-Translational / Q000276:immunology; D021122:Protein Subunits / Q000378:metabolism; D011961:Receptors, Fc / Q000302:isolation & purification / Q000378:metabolism*; D001613:beta 2-Microglobulin / Q000378:metabolism*
D037282:Calreticulin; D004220:Disulfides; D006360:Heat-Shock Proteins; D015395:Histocompatibility Antigens Class I; D007074:Immunoglobulin G; D021122:Protein Subunits; D011961:Receptors, Fc; D001613:beta 2-Microglobulin; D037281:Calnexin; D007535:Isomerases; D019704:Protein Disulfide-Isomerases; C085868:PDIA3 protein, human; C100688:Fc receptor, neonatal