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TAPBPR isoforms exhibit altered association with MHC class I.

Porter KM, Hermann C, Traherne JA, Boyle LH - Immunology (2014)

Bottom Line: TAPBPR transcripts lacking exon 5 result in loss of the membrane proximal IgC domain and loss of ability to bind to MHC class I.This longer TAPBPR protein interacted with MHC class I but was attenuated in its ability to down-regulate surface expression of MHC class I.The abundance of these alternative transcripts in peripheral blood mononuclear cells and dendritic cells suggests an important role of TAPBPR isoforms in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Cambridge Institute of Medical Research, University of Cambridge, Wellcome Trust, Cambridge, UK.

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Alternative TAPBPR transcripts. (a, b) TAPBPR was amplified from cDNA generated from the indicated cell lines using primers specific for exon 1 and exon 7 of TAPBPR. The PCR products were cloned into the pCR-blunt-II TOPO vector and colonies containing inserts were identified by PCR using primers Sp6 and T7. A selection of colonies was further screened by sequence analysis. PCR products identified as alternative TAPBPR isoforms are highlighted with a symbol on the PCR screen. (c) Exon structure of seven human TAPBPR transcripts. cDNA sequences were aligned against human TAPBPL gene on chromosome 12p13.3 (OMIM:607081) using spideymRNA genomic alignment software (http://www.ncbi.nlm.nih.gov/spidey/) using default settings. The transcript for TAPBPR α has previously been reported (NM_018009.4). β, γ, δ, ε, ζ, η mRNA products have not been described previously.
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fig01: Alternative TAPBPR transcripts. (a, b) TAPBPR was amplified from cDNA generated from the indicated cell lines using primers specific for exon 1 and exon 7 of TAPBPR. The PCR products were cloned into the pCR-blunt-II TOPO vector and colonies containing inserts were identified by PCR using primers Sp6 and T7. A selection of colonies was further screened by sequence analysis. PCR products identified as alternative TAPBPR isoforms are highlighted with a symbol on the PCR screen. (c) Exon structure of seven human TAPBPR transcripts. cDNA sequences were aligned against human TAPBPL gene on chromosome 12p13.3 (OMIM:607081) using spideymRNA genomic alignment software (http://www.ncbi.nlm.nih.gov/spidey/) using default settings. The transcript for TAPBPR α has previously been reported (NM_018009.4). β, γ, δ, ε, ζ, η mRNA products have not been described previously.

Mentions: During the initial process of cloning TAPBPR into expression constructs, it became apparent that a number of alternative TAPBPR mRNA products existed in addition to the major TAPBPR transcript.10 We investigated the alternative transcripts using PCR, followed by cloning individual TAPBPR transcripts into the pCR®-Blunt II TOPO vector and screening colonies by sequence analysis. Using primers specific for TAPBPR exon 1 and exon 7 we amplified the major TAPBPR coding sequence, the α transcript, which was found in all cell types tested (Fig.1 and Table1). In IFN-γ-treated KG-1 cells we identified a longer TAPBPR mRNA product that we named the TAPBPR β transcript (Fig.1a). Although the TAPBPL gene is thought to be composed of seven exons,11 the TAPBPR β transcript has an additional exon after exon 6, which we refer to as exon 7a.


TAPBPR isoforms exhibit altered association with MHC class I.

Porter KM, Hermann C, Traherne JA, Boyle LH - Immunology (2014)

Alternative TAPBPR transcripts. (a, b) TAPBPR was amplified from cDNA generated from the indicated cell lines using primers specific for exon 1 and exon 7 of TAPBPR. The PCR products were cloned into the pCR-blunt-II TOPO vector and colonies containing inserts were identified by PCR using primers Sp6 and T7. A selection of colonies was further screened by sequence analysis. PCR products identified as alternative TAPBPR isoforms are highlighted with a symbol on the PCR screen. (c) Exon structure of seven human TAPBPR transcripts. cDNA sequences were aligned against human TAPBPL gene on chromosome 12p13.3 (OMIM:607081) using spideymRNA genomic alignment software (http://www.ncbi.nlm.nih.gov/spidey/) using default settings. The transcript for TAPBPR α has previously been reported (NM_018009.4). β, γ, δ, ε, ζ, η mRNA products have not been described previously.
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fig01: Alternative TAPBPR transcripts. (a, b) TAPBPR was amplified from cDNA generated from the indicated cell lines using primers specific for exon 1 and exon 7 of TAPBPR. The PCR products were cloned into the pCR-blunt-II TOPO vector and colonies containing inserts were identified by PCR using primers Sp6 and T7. A selection of colonies was further screened by sequence analysis. PCR products identified as alternative TAPBPR isoforms are highlighted with a symbol on the PCR screen. (c) Exon structure of seven human TAPBPR transcripts. cDNA sequences were aligned against human TAPBPL gene on chromosome 12p13.3 (OMIM:607081) using spideymRNA genomic alignment software (http://www.ncbi.nlm.nih.gov/spidey/) using default settings. The transcript for TAPBPR α has previously been reported (NM_018009.4). β, γ, δ, ε, ζ, η mRNA products have not been described previously.
Mentions: During the initial process of cloning TAPBPR into expression constructs, it became apparent that a number of alternative TAPBPR mRNA products existed in addition to the major TAPBPR transcript.10 We investigated the alternative transcripts using PCR, followed by cloning individual TAPBPR transcripts into the pCR®-Blunt II TOPO vector and screening colonies by sequence analysis. Using primers specific for TAPBPR exon 1 and exon 7 we amplified the major TAPBPR coding sequence, the α transcript, which was found in all cell types tested (Fig.1 and Table1). In IFN-γ-treated KG-1 cells we identified a longer TAPBPR mRNA product that we named the TAPBPR β transcript (Fig.1a). Although the TAPBPL gene is thought to be composed of seven exons,11 the TAPBPR β transcript has an additional exon after exon 6, which we refer to as exon 7a.

Bottom Line: TAPBPR transcripts lacking exon 5 result in loss of the membrane proximal IgC domain and loss of ability to bind to MHC class I.This longer TAPBPR protein interacted with MHC class I but was attenuated in its ability to down-regulate surface expression of MHC class I.The abundance of these alternative transcripts in peripheral blood mononuclear cells and dendritic cells suggests an important role of TAPBPR isoforms in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Cambridge Institute of Medical Research, University of Cambridge, Wellcome Trust, Cambridge, UK.

Show MeSH