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* Whitehead Institute for Biomedical Research, Cambridge, MA 02142;
Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore; and
Department of Biological Sciences, National University of Singapore, Singapore, Singapore
The accumulation of misfolded secreted IgM in the endoplasmic reticulum (ER) of X-box binding protein 1 (XBP-1)-deficient B cells has been held responsible for the inability of such cells to yield plasma cells, through the failure to mount a proper unfolded protein response. LPS-stimulated B cells incapable of secreting IgM still activate the XBP-1 axis normally, as follows: XBP-1 is turned on by cues that trigger differentiation and not in response to accumulation of unfolded IgM, but the impact of XBP-1 deficiency on glycoprotein folding and assembly has not been explored. The lack of XBP-1 compromised neither the formation of functional hen egg lysozyme-specific IgM nor the secretion of free 
-chains. Although XBP-1 deficiency affects the synthesis of some ER chaperones, including protein disulfide isomerase, their steady state levels do not drop below the threshold required for proper assembly and maturation of the Ig
/Igβ heterodimer and MHC molecules. Intracellular transport and surface display of integral membrane proteins are unaffected by XBP-1 deficiency. Given the fact that we failed to observe any defects in folding of a variety of glycoproteins, we looked for other means to explain the requirement for XBP-1 in plasma cell development. We observed significantly reduced levels of phosphatidylcholine, sphingomyelin, and phosphatidylinositol in total membranes of XBP-1-deficient B cells, and reduced ER content. Terminal N-linked glycosylation of IgM and class I MHC was altered in these cells. XBP-1 hence has important roles beyond folding proteins in the ER.
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
1 This work was supported by grants from the National Institutes of Health (to H.L.P.). A.M.M. was supported by a National Science Foundation graduate research fellowship. S.K.D. is supported by a Cancer Research Institute fellowship. E.J.K. was supported by a National Science Foundation East Asia and Pacific Summer Institutes fellowship grant and the Singapore National Research Foundation. National Research Foundation under Competitive Research Programme Award 2007-04, the Academic Research Fund (R-183-000-160-112), the Biomedical Research Council of Singapore (R-183-000-211-305), and the National Medical Research Council (R-183-000-224-213) to M.R.W. are gratefully acknowledged.
2 Address correspondence and reprint requests to Dr. Chih-Chi Andrew Hu and Dr. Hidde L. Ploegh, Whitehead Institute for Biomedical Research, Nine Cambridge Center, Cambridge, MA 02142. E-mail addresses: chih-chi{at}wi.mit.edu and ploegh{at}wi.mit.edu
3 Abbreviations used in this paper: mIgM, membrane form of IgM; ATF, activating transcription factor; Cer, ceramide; eIF2, eukaryotic initiation factor 2; ER, endoplasmic reticulum; HEL, hen egg lysozyme; KO, knockout; NEM, N-ethylmaleimide; PC, phosphatidylcholine; PDI, protein disulfide isomerase; PG, phosphatidylglycerol; PI, phosphatidylinositol; PS, phosphatidylserine; PTP-1B, protein-tyrosine phosphatase 1B; sIgM, secreted form of IgM; SM, sphingomyelin; µM, membrane µ; µS, secreted µ; UPR, unfolded protein response; WT, wild type; Endo H, endoglycosidase H; IRE-1, inositol-requiring enzyme 1; PERK, RNA-dependent protein kinase (PKR)-like ER kinase; PNGase F, peptide: N-glycosidase F; XBP-1, X-box binding protein 1; XBP-1s, spliced XBP-1 protein.
4 The online version of this article contains supplemental material.
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