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Post-translational Modification of the NKG2D Ligand RAET1G Leads to Cell Surface Expression of a Glycosylphosphatidylinositol-linked Isoform

Ohashi, Maki and Eagle, Robert A. and Trowsdale, John (2010) Post-translational Modification of the NKG2D Ligand RAET1G Leads to Cell Surface Expression of a Glycosylphosphatidylinositol-linked Isoform. Journal of Biological Chemistry, 285 (22). pp. 16408-16415. ISSN 0021-9258. PMCID PMC2878053. http://resolver.caltech.edu/CaltechAUTHORS:20100615-134555139

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Abstract

NKG2D is an important activating receptor on lymphocytes. In human, it interacts with two groups of ligands: the major histocompatibility complex class I chain-related A/B (MICA/B) family and the UL-16 binding protein (ULBP) family, also known as retinoic acid early transcript (RAET1). MIC proteins are membrane-anchored, but all of the ULBP/RAET1 proteins, except for RAET1E and RAET1G, are glycosylphosphatidylinositol (GPI)-anchored. To address the reason for these differences we studied the association of RAET1G with the membrane. Using epitope-tagged RAET1G protein in conjunction with antibodies to different parts of the molecule and in pulse-chase experiments, we showed that the C terminus of the protein was cleaved soon after protein synthesis. Endoglycosidase H and peptide N-glycosidase treatment and cell surface immunoprecipitation indicated that most of the protein stayed in the endoplasmic reticulum, but some of the cleaved form was modified in the Golgi and transported to the cell surface. We examined the possibility of GPI anchoring of the protein in three ways: (i) Phosphatidylinositol (PI)-specific phospholipase C released the PI-linked form of the protein. (ii) The surface expression pattern of RAET1G decreased in cells defective in GPI anchoring through mutant GPI-amidase. (iii) Site-directed mutagenesis, to disrupt residues predicted to facilitate GPI-anchoring, resulted in diminished surface expression of RAET1G. Thus, a form of RAET1G is GPI-anchored, in line with most other ULBP/RAET1 family proteins. The cytoplasmic tail and transmembrane domains appear to result from gene duplication and frameshift mutation. Together with our previous results, our data suggest that RAET1G is regulated post-translationally to produce a GPI-anchored isoform.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1074/jbc.M109.077636 DOIArticle
http://www.jbc.org/content/285/22/16408PublisherArticle
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2878053/PubMed CentralArticle
Additional Information:© 2010 American Society for Biochemistry and Molecular Biology. Received October 23, 2009. Revision received February 19, 2010. First Published on March 19, 2010. This work was supported by a grant from Cancer Research United Kingdom. We thank Dr. N. Kanzawa and Prof. T. Kinoshita for cell lines, Prof. P. Lehner for plasmid, Dr. A. P. Kelly for helpful advice, and Dr. L. H. Boyle for help with the pulse-chase experiment.
Funders:
Funding AgencyGrant Number
Cancer Research United KingdomUNSPECIFIED
Subject Keywords:Glycolipids/Glycosylphosphatidylinositol Anchors ; Immunology ; Immunology/Cellular Response ; Membrane/Proteins ; Protein/Post-translational Modification ; Protein/Processing ; Natural Killer (NK) Cell ; NKG2D
PubMed Central ID:PMC2878053
Record Number:CaltechAUTHORS:20100615-134555139
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:20100615-134555139
Official Citation:Ohashi, M., R. A. Eagle, et al. (2010). "Post-translational Modification of the NKG2D Ligand RAET1G Leads to Cell Surface Expression of a Glycosylphosphatidylinositol-linked Isoform." Journal of Biological Chemistry 285(22): 16408-16415. doi:10.1074/jbc.M109.077636
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:18691
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:15 Jul 2010 23:49
Last Modified:03 Nov 2016 22:34

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