A critical role for tapasin in the assembly and function of multimeric MHC class I-TAP complexes

Science

Volumn 277, Issue 5330, 1997, Pages 1306-1309

  Ortmann, Bodo ^a,e   Copeman, James ^b,e   Lehner, Paul J ^e   Sadasivan, Bhanu ^c,e   Herberg, Jethro A ^f   Grandea, Andeas G ^g   Riddell, Stanley R ^g   Tampé, Robert ^h   Spies, Thomas ^g   Trowsdale, John ^d,f   Cresswell, Peter ^e  

^a UNIVERSITY OF COLOGNE   (Germany)

^b MOUNT SINAI HOSPITAL   (Canada)

^c BRIGHAM AND WOMEN S HOSPITAL   (United States)

^d Immunology Division   (United Kingdom)

^e YALE UNIVERSITY SCHOOL OF MEDICINE   (United States)

^f IMPERIAL CANCER RESEARCH FUND   (United Kingdom)

^g Fred Hutchinson Cancer Research Center   (United States)

^h MAX PLANCK INSTITUTE OF BIOCHEMISTRY   (Germany)

Author keywords

[No Author keywords available]

Indexed keywords

CALRETICULIN; CARRIER PROTEIN; CHAPERONE; IMMUNOGLOBULIN; MAJOR HISTOCOMPATIBILITY ANTIGEN CLASS 1; MEMBRANE PROTEIN;

ANTIGEN PRESENTATION; ANTIGEN RECOGNITION; ARTICLE; COMPLEX FORMATION; CYTOTOXIC T LYMPHOCYTE; ENDOPLASMIC RETICULUM; GENE EXPRESSION; GENETIC LINKAGE; MAJOR HISTOCOMPATIBILITY COMPLEX RESTRICTION; MOLECULAR CLONING; PRIORITY JOURNAL; PROTEIN ASSEMBLY; SIGNAL TRANSDUCTION;

AMINO ACID SEQUENCE; ANTIGEN PRESENTATION; ANTIPORTERS; ATP-BINDING CASSETTE TRANSPORTERS; CALCIUM-BINDING PROTEINS; CALRETICULIN; CELL LINE; CELL LINE, TRANSFORMED; CHROMOSOME MAPPING; CHROMOSOMES, HUMAN, PAIR 6; CLONING, MOLECULAR; DIMERIZATION; ENDOPLASMIC RETICULUM; HISTOCOMPATIBILITY ANTIGENS CLASS I; HLA ANTIGENS; HUMANS; IMMUNOGLOBULIN G; IMMUNOGLOBULINS; LINKAGE (GENETICS); MAJOR HISTOCOMPATIBILITY COMPLEX; MEMBRANE TRANSPORT PROTEINS; MOLECULAR SEQUENCE DATA; RIBONUCLEOPROTEINS; SEQUENCE HOMOLOGY, AMINO ACID; T-LYMPHOCYTES, CYTOTOXIC; TUMOR CELLS, CULTURED;

EID: 0030865333     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.277.5330.1306     Document Type: Article

References (44)

1. W. Suh et al., Science 264, 1322 (1994).
2. B. Ortmann, M. Androlewicz, P. Cresswell, Nature 368, 864 (1994).
3. E. Noessner and P. Parham, J. Exp. Med. 181, 327 (1995); W.-K. Suh et al., ibid. 184, 337 (1996).
4. E. Noessner and P. Parham, J. Exp. Med. 181, 327 (1995); W.-K. Suh et al., ibid. 184, 337 (1996).
5. B. Sadasivan, P. J. Lehner, B. Ortmann, T. Spies, P. Cresswell, Immunity 5, 103 (1996).
6. J. E. M. Van Leeuwen and K. P. Kearse, Proc. Natl. Acad. Sci. U.S.A. 93, 13997 (1996); J. C. Solheim, M. R. Harris, C. S. Kindle, T. H. Hansen, J. Immunol. 158, 2236 (1997).
7. J. E. M. Van Leeuwen and K. P. Kearse, Proc. Natl. Acad. Sci. U.S.A. 93, 13997 (1996); J. C. Solheim, M. R. Harris, C. S. Kindle, T. H. Hansen, J. Immunol. 158, 2236 (1997).
8. A. L. Goldberg and K. L. Rock, Nature 357, 375 (1992).
9. J. C. Howard, Curr. Opin. Immunol. 7, 69 (1995).
10. A. G. Grandea III, M. J. Androlewicz, R. S. Athwal, D. E. Geraghty, T. Spies, Science 270, 105 (1995).
11. R. Greenwood, Y. Shimizu, G. S. Sekhon, R. DeMars, J. Immunol. 153, 5525 (1994).
12. A. G. Grandea, P. J. Lehner, S. R. Riddell, P. Cresswell, T. Spies, Immunogenetics, in press.
13. 2-terminal sequencing was performed by the Keck Foundation Biotechnology Research Foundation, Yale University.
14. TAP complexes isolated from Swei cells were dialyzed extensively against 10 mM bicine, 150 mM NaCl, pH 8.2, and biotinylated by adding NHS-LC-biotin (Pierce) to a final concentration of 200 μM and incubating for 30 min at 4°C. Biotinylation was terminated by adding glycine to a final concentration of 10 mM. The biotinylated proteins were denatured by adding SDS (2%) and dithiothreitol (2 mM) and heating the preparation to 100°C for 5 min. For immunoprecipitation, the denatured proteins were diluted 10-fold with 1% Triton X-100 in TBS containing 10 mM iodoacetamide, incubated for 30 min at 25°C, and then precipitated with the appropriate antibodies and protein A-or protein G-Sepharose (Pharmacia) before being subjected to SDS-PAGE. Biotinylated proteins were detected with avidin-HRP and a chemiluminescent substrate (ECL, Amersham).
15. T. H. Meyer, P. M. Van Endert, S. Uebel, B. Ehring, R. Tampé, FEBS Lett. 351, 443 (1994).
16. M. J. Androlewicz, B. Ortmann, P. M. van Endert, T. Spies, P. Cresswell, Proc. Natl. Acad. Sci. U.S.A. 91, 12716 (1994).
17. B. Ortmann et al., data not shown.
18. 2-terminal amino acid sequencing, and from a sequence from a tryptic peptide that proved to be residues 77 through 83 (Fig. 2A), oligonucleotide primers were designed according to preferred human codon usage, but they were degenerate where necessary in the last six bases. RNA was extracted from Swei cells with TRIZOL (Gibco-BRL) and reverse transcriptase-PCR performed. Products were gel purified and cloned into pCR2.1 (Invitrogen). We generated the cycle sequencing template by PCR using resuspended colonies of clones as the template. Sequencing was performed by the Keck facility (Howard Hughes Medical Institute, Yale Medical School). The remaining sequence was obtained from a walking approach by vectorette PCR [P. M. Sharp et al., Nucleic Acids Res. 16, 8207 (1988)]. Polyadenylated RNA was purified with Oligotex (Qiagen), and the cDNA prepared (Gibco-BRL). Restriction digestions of 500 ng of cDNA were performed with Alu I, Hae III, Bsa AI, Hinc II, Msl I, Eco RV with Xmn I, and Pvu II with Stu I, and the vectorettes were ligated. Vectorette PCR was performed with a tapasin-and a vectorette-specific primer, with the above cDNA digestions as separate templates. PCR products were cloned and sequenced as above. For each tapasin-specific primer, a number of products from different cDNA digestions were sequenced in both directions. New tapasin-specific primers were designed to the sequence obtained. Alignment of the multiple overlapping sequences allowed the derivation of a consensus sequence. We verified this by amplifying the full tapasin coding region from a separate synthesis of intact Swei cDNA using an enzyme with proofreading capability (Pfu DNA polymerase; Stratagene). Products were cloned into pCR2.1 and the clones sequenced in both directions. These sequences corresponded to the consensus sequence from vectorette PCR. Details of PCR reactions are available upon request.
19. M. R. Jackson, T. Nilsson, P. A. Peterson, EMBO J. 9, 3153 (1990).
20. P. Cosson, S. P. Lankford, J. S. Bonifacino, R. D. Klausner, Nature 351, 414 (1991); N. M. Green, ibid., p. 349.
21. P. Cosson, S. P. Lankford, J. S. Bonifacino, R. D. Klausner, Nature 351, 414 (1991); N. M. Green, ibid., p. 349.
22. G. A. Adams and J. K. Rose, Cell 41, 1007 (1985).
23. J. F. Collins, A. F. W. Coulson, Methods Enzymol. 183, 474 (1990); J. F. Collins and A. F. W. Coulson, in Nucleic Acid and Protein Sequence Analysis: A Practical Approach, M. J. Bishop and C. J. Rawlings, Eds. (IRL, Oxford, 1987), pp. 323-358. The University of California, Los Angeles-Department of Energy structure prediction server is at http://www. mbi.ucla.edu.
24. J. F. Collins, A. F. W. Coulson, Methods Enzymol. 183, 474 (1990); J. F. Collins and A. F. W. Coulson, in Nucleic Acid and Protein Sequence Analysis: A Practical Approach, M. J. Bishop and C. J. Rawlings, Eds. (IRL, Oxford, 1987), pp. 323-358. The University of California, Los Angeles-Department of Energy structure prediction server is at http://www. mbi.ucla.edu.
25. A. F. Williams and A. N. Barclay, Annu. Rev. Immunol. 6, 381 (1988).
26. Probe DNA was biotin-labeled and hybridized with standard techniques [G. Senger, I. Ragoussis, J. Trowsdale, D. Sheer, Cytogenet. Cell Genet. 64, 49 (1993); T. Jones et al., Genomics 40, 189 (1997)]. Briefly, 400 ng of labeled probe was mixed with 5 μg of Cot-1 DNA (Gibco-BRL, Paisley, UK), precipitated, denatured, allowed to anneal, and then applied to a denatured slide and hybridized overnight. Slides were washed and the probe signal detected with fluoresceinated avidin. Slides were counterstained with 4′,6′-diamidino-2-phenylindole (DAPI) and then mounted in Citifluor (Chemlab, Canterbury, UK). Chromosome images were taken with a Zeiss Axioskop epifluorescence microscope equipped with a Photometries KAF 1400-500 charge-coupled device camera attached to an Apple Powermac 8100 computer. Separate images of probe signals and DAPI banding patterns were pseudocolored and merged with SmartCapture software (Vysis, Chicago, IL).
27. Probe DNA was biotin-labeled and hybridized with standard techniques [G. Senger, I. Ragoussis, J. Trowsdale, D. Sheer, Cytogenet. Cell Genet. 64, 49 (1993); T. Jones et al., Genomics 40, 189 (1997)]. Briefly, 400 ng of labeled probe was mixed with 5 μg of Cot-1 DNA (Gibco-BRL, Paisley, UK), precipitated, denatured, allowed to anneal, and then applied to a denatured slide and hybridized overnight. Slides were washed and the probe signal detected with fluoresceinated avidin. Slides were counterstained with 4′,6′-diamidino-2-phenylindole (DAPI) and then mounted in Citifluor (Chemlab, Canterbury, UK). Chromosome images were taken with a Zeiss Axioskop epifluorescence microscope equipped with a Photometries KAF 1400-500 charge-coupled device camera attached to an Apple Powermac 8100 computer. Separate images of probe signals and DAPI banding patterns were pseudocolored and merged with SmartCapture software (Vysis, Chicago, IL).
28. P. van Endert et al., J. Exp. Med. 182, 1883 (1995).
29. A. Neisig, R. Wubbolts, X. Zang, C. Melief, J. Neefjes, J. Immunol. 156, 3196 (1996).
30. R. A. Henderson et al., Science 255, 1264 (1992); M. L. Wei and P. Cresswell, Nature 356, 443 (1992).
31. R. A. Henderson et al., Science 255, 1264 (1992); M. L. Wei and P. Cresswell, Nature 356, 443 (1992).
32. L. K. Denzin, C. Hammond, P. Cresswell, J. Exp. Med. 184, 2153 (1996).
33. R. Busch, I. Cloutier, R.-P. Sékaly, G. J. Hämmerling, EMBO J. 15, 412 (1996).
34. C. Hammond and A. Helenius, J. Cell. Biol. 126, 41 (1994).
35. P. M. van Endert et al., Immunity 1, 491 (1994).
36. P. M. Lutz and P. Cresswell, Immunogenetics 25, 228 (1987).
37. Single-letter abbreviations for the amino acid residues are as follows: A, Ala; C, Cys; D, Asp; E, Glu; F, Phe; G, Gly; H, His; I, Ile; K, Lys; L, Leu; M, Met; N, Asn; P, Pro; Q, Gln; R, Arg; S, Ser; T, Thr; V, Val; W, Trp; and Y, Tyr.
38. K. W. Willison et al., EMBO J. 6, 1967 (1987).
39. Tapasin cDNA was amplified by PCR with the Pfu DNA polymerase (Stratagene) under standard PCR conditions. The forward primer was 5′-GGGGTACCGCCGCCATGAAGTCCCTGTCTCTGCTCCT-3′ and the reverse primer was 5′-CCATCGATTCACTCTGCTTTCTTCTTTGAATCCTTGCA-3′. A consensus Kozak sequence was introduced at the 5′ end to ensure optimal expression. The 1367-bp amplification product was cloned into the polylinker of the pCR2.1 cloning vector (Invitrogen), sequenced in both directions, and subcloned into the pMCFR-PAC vector by using the 5′ Kpn and 3′ Cla sites. Stable tapasin-transfected 220-B8 clones were generated by electroporation at 210 volts, 960 mF, and selected in medium containing puromycin (500 ng/ml).
40. We transfected 220-A1 cells with the tapasin cDNA in pREP8D by electroporation and selected transfectants with L-histidinol as described [D. Arnold et al., Nature 360, 171 (1992)]. Positive isolates expressing increased surface amounts of HLA-A1 were identified by indirect immunofluorescence with mAb GS142.1 [K. A. Nelson, J. G. Bodmer, A. Martin, C. Navarette, D. M. Strong, in Immunobiology of HLA, B. Dupont, Ed. (Springer-Vertag, New York, 1989), pp. 292-301].
41. We transfected 220-A1 cells with the tapasin cDNA in pREP8D by electroporation and selected transfectants with L-histidinol as described [D. Arnold et al., Nature 360, 171 (1992)]. Positive isolates expressing increased surface amounts of HLA-A1 were identified by indirect immunofluorescence with mAb GS142.1 [K. A. Nelson, J. G. Bodmer, A. Martin, C. Navarette, D. M. Strong, in Immunobiology of HLA, B. Dupont, Ed. (Springer-Vertag, New York, 1989), pp. 292-301].
42. P. J. Lehner, J. T. Karttunen, G. W. G. Wilkinson, P. Cresswell, Proc. Natl. Acad. Sci. U.S.A. 94, 6904 (1997).
43. S. R. Riddell et al., Nature Med. 2, 216 (1996).
44. We thank P. Freemont for help with the homology analysis, members of D. Sheer's laboratory for FISH analysis, J. Yewdell for supplying influenza virus, T. Novak for the pMCFR-PAC vector, K. Stone and the Keck Foundation Biotechnology Resource Laboratory at Yale University for their contributions, and N. Dometios for preparation of the manuscript. B.O. was supported by an AIDS-Stipendium of the German Cancer Research Center and P.J.L. by the Wellcome Trust. The work described was supported by the Howard Hughes Medical Institute and by a grant (AI30581) from the NIH (T.S.).