Transport of Peptide-MHC Class II Complexes in Developing Dendritic Cells
10.1126/science.288.5465.522
Crossref journal-article
American Association for the Advancement of Science (AAAS)
Science (221)
Abstract

Major histocompatibility complex class II (MHC II) molecules capture peptides within the endocytic pathway to generate T cell receptor (TCR) ligands. Immature dendritic cells (DCs) sequester intact antigens in lysosomes, processing and converting antigens into peptide–MHC II complexes upon induction of DC maturation. The complexes then accumulate in distinctive, nonlysosomal MHC II + vesicles that appear to migrate to the cell surface. Although the vesicles exclude soluble lysosomal contents and antigen-processing machinery, many contain MHC I and B7 costimulatory molecules. After arrival at the cell surface, the MHC and costimulatory molecules remain clustered. Thus, transport of peptide–MHC II complexes by DCs not only accomplishes transfer from late endocytic compartments to the plasma membrane, but does so in a manner that selectively concentrates TCR ligands and costimulatory molecules for T cell contact.

Bibliography

Turley, S. J., Inaba, K., Garrett, W. S., Ebersold, M., Unternaehrer, J., Steinman, R. M., & Mellman, I. (2000). Transport of Peptide-MHC Class II Complexes in Developing Dendritic Cells. Science, 288(5465), 522–527.

Authors 7
  1. Shannon J. Turley (first)
  2. Kayo Inaba (additional)
  3. Wendy S. Garrett (additional)
  4. Melanie Ebersold (additional)
  5. Julia Unternaehrer (additional)
  6. Ralph M. Steinman (additional)
  7. Ira Mellman (additional)
References 35 Referenced 385
  1. Watts C., Annu. Rev. Immunol. 15, 821 (1997). (10.1146/annurev.immunol.15.1.821) / Annu. Rev. Immunol. by Watts C. (1997)
  2. 10.1146/annurev.iy.12.040194.001355
  3. Chapman H., Curr. Opin. Immunol. 10, 93 (1998). (10.1016/S0952-7915(98)80038-1) / Curr. Opin. Immunol. by Chapman H. (1998)
  4. Kornfeld S., Mellman I., Annu. Rev. Cell Biol. 5, 483 (1989). (10.1146/annurev.cb.05.110189.002411) / Annu. Rev. Cell Biol. by Kornfeld S. (1989)
  5. Stinchcombe J., Griffiths G. M., J. Cell Biol. 147, 1 (1999); (10.1083/jcb.147.1.1) / J. Cell Biol. by Stinchcombe J. (1999)
  6. 10.1083/jcb.135.3.611
  7. Raposo G., et al., J. Exp. Med. 183, 1161 (1996). (10.1084/jem.183.3.1161) / J. Exp. Med. by Raposo G. (1996)
  8. 10.1038/369113a0
  9. Pierre P., et al., Nature 388, 787 (1997); (10.1038/42039) / Nature by Pierre P. (1997)
  10. Talmor M., et al., Eur. J. Immunol. 28, 811 (1998). (10.1002/(SICI)1521-4141(199803)28:03<811::AID-IMMU811>3.0.CO;2-S) / Eur. J. Immunol. by Talmor M. (1998)
  11. Inaba K., et al., J. Exp. Med. 191, 927 (2000). (10.1084/jem.191.6.927) / J. Exp. Med. by Inaba K. (2000)
  12. 10.1038/32588
  13. Mellman I., Turley S., Steinman R. M., Trends Cell Biol. 8, 231 (1998). (10.1016/S0962-8924(98)01276-8) / Trends Cell Biol. by Mellman I. (1998)
  14. Zhong G., Reis e Sousa C., Germain R. N., Proc. Natl. Acad. Sci. U.S.A. 94, 13856 (1997). (10.1073/pnas.94.25.13856) / Proc. Natl. Acad. Sci. U.S.A. by Zhong G. (1997)
  15. DC cultures were generated as described (5 22). Immature I-A k+ DCs grown from C3H/HeN C3H/HeJ and CBA/J (Jackson Labs Bar Harbor ME) marrow suspensions were pulsed with HEL (1 to 3 mg/ml) with or without LPS (1 to 10 ng/ml E. coli 0111.B4) for 1 to 3 hours washed and chased in HEL- and LPS-free media. Similar results were obtained with DCs from each strain. DC cultures contained supernatant from mGM-CSF–expressing J558L cells (from A. Lanzavecchia Basel Institute Basel Switzerland). LPS was removed from HEL (Sigma) with Kuttsuclean adsorbent (Maruha Corporation Ibaraki Japan). Maturation stimuli included LPS CD40L tumor necrosis factor–α or replating (22).
  16. DCs were fixed with 4% (wt/vol) paraformaldehyde and washed in phosphate-buffered saline. Cells were incubated with monoclonal antibodies (mAbs) on ice as described (23). The rat mAb C4H3 was provided by R. Germain (NIH Bethesda MD). C4H3 was purified on protein A–Sepharose and biotinylated. Cells were labeled with biotin-conjugated mAbs (Pharmingen) to I-E k (14-4-4S) or I-A k (10-2.16) and fluorescein isothiocyanate (FITC)–conjugated mAb to CD86 (GL1). Isotype controls were purchased from Pharmingen. Biotinylated mAbs were detected with streptavidin-phycoerythrin. Fluorescence analysis was done with a FACSCalibur and CellQuest software.
  17. For ICM DCs were adhered to Alcian blue–coated glass coverslips for 20 min at 37°C in serum-free media. Cell were fixed in 4% (wt/vol) PFA and permeabilized (for intracellular staining) in 0.05% saponin 10 mM Hepes (pH 7.4) 10 mM glycine (pH 8.0) and 10% goat serum in RPMI 1640. Antibody incubations were done as described (6). Purified C4H3 was used for microscopy. To detect HEL we used the mouse mAb 1B12 provided by P. Allen (Washington University St. Louis MO). Rabbit anti–H-2M (ULM) and rabbit anti–mouse I-A (RIV) have been described elsewhere (6). Hybridomas TIB126 (anti–MHC I) TIB99 (anti-Thy1.2) and TIB93 (anti–I-Ak) were obtained from American Type Culture Collection. Rabbit anti-CD18 was a gift from P. Blier (Boehringer Ingelheim CT). Several mouse anti-rat mAbs were the gift of J. P. Soullilou (University of Nantes Nantes France) including OX-18 (anti–MHC I) anti-Thy1.1 and OX-6 (anti–MHC II). Rat B7 was detected with a CTLA-4–human IgG fusion protein (a gift of P. Linsley Bristol Meyers Squibb) in combination with a FITC-conjugated horse anti–human IgG. Mouse B7-2 was detected with GL1. Rat lysosomes were stained with rabbit lgp120 antiserum (24). Irrelevant isotype-matched mAbs preimmune rabbit serum or normal human serum were used as negative controls in all experiments. Coverslips were mounted in Mowiol with DABCO (Calbiochem La Jolla CA) and fluorescence was analyzed with conventional and confocal (0.5- to 1.0-μm optical sections) Zeiss microscopes.
  18. DCs were treated with nocodazole (30 μM Sigma) or Latrunculin B (0.4 to 10 μg/ml; Calbiochem San Diego CA) after the HEL pulse to allow internalization and immunogenic complex formation to occur.
  19. DCs harvested at various times during the HEL pulse-chase assay were fixed immediately with 4% PFA and 0.4% glutaraldehyde in 200 mM phosphate buffer. Cells were then processed for cryosectioning by embedding in 10% gelatin on ice. Blocks were saturated in 2.3 M sucrose overnight at 4°C. Ultrathin cryosections were retrieved with methylcellulose-sucrose and double-immunolabeled as described (25) with 5- and 10-nm gold particles. To identify MIICs and CIIVs we used rat mAbs LAMP-1 LAMP-2 GL-1 and C4H3 and the mouse mAb 10-2.16 against I-A k .
  20. Harding C. V., Geuze H. J., J. Cell Biol. 119, 531 (1992); (10.1083/jcb.119.3.531) / J. Cell Biol. by Harding C. V. (1992)
  21. Kleijmeer M. J., Oorschot V. M., Geuze H. J., J. Invest. Dermatol. 103, 516 (1994); (10.1111/1523-1747.ep12395666) / J. Invest. Dermatol. by Kleijmeer M. J. (1994)
  22. Kleijmeer M. J., Morkowski S., Griffith J. M., Rudensky A. Y., Geuze H. J., J. Cell Biol. 139, 639 (1997). (10.1083/jcb.139.3.639) / J. Cell Biol. by Kleijmeer M. J. (1997)
  23. 10.1038/42408
  24. W. S. Garrett and I. Mellman in preparation.
  25. J. Unternaehrer and I. Mellman in preparation.
  26. Pierre P., Mellman I., Cell 93, 1135 (1998); (10.1016/S0092-8674(00)81458-0) / Cell by Pierre P. (1998)
  27. Cella M., et al., Nature 388, 782 (1997); (10.1038/42030) / Nature by Cella M. (1997)
  28. Rescigno M., et al., Proc. Natl. Acad. Sci. U.S.A. 95, 5229 (1998). (10.1073/pnas.95.9.5229) / Proc. Natl. Acad. Sci. U.S.A. by Rescigno M. (1998)
  29. 10.1126/science.285.5425.221
  30. 10.1038/25764
  31. 10.1084/jem.176.6.1693
  32. Inaba K., et al., J. Exp. Med. 188, 2163 (1998). (10.1084/jem.188.11.2163) / J. Exp. Med. by Inaba K. (1998)
  33. Lewis V., et al., J. Cell Biol. 100, 1839 (1985). (10.1083/jcb.100.6.1839) / J. Cell Biol. by Lewis V. (1985)
  34. Slot J. W., Geuze H. J., Eur. J. Cell Biol. 38, 87 (1985). / Eur. J. Cell Biol. by Slot J. W. (1985)
  35. We thank members of the Mellman Steinman and Inaba laboratories for valuable advice. Supported by grants from NIH (AI-34098 to I.M.) the National Institute of Allergy and Infectious Diseases (AI-13013 and AI-39672 to R.M.S.) and the Ministry of Education Science and Culture of Japan (grants 08282104 10153226 and 10044268 to K.I.). I.M. is a member of the Ludwig Institute for Cancer Research.
Dates
Type When
Created 23 years, 1 month ago (July 27, 2002, 5:40 a.m.)
Deposited 1 year, 7 months ago (Jan. 13, 2024, 5:01 a.m.)
Indexed 3 months, 3 weeks ago (May 10, 2025, 12:03 p.m.)
Issued 25 years, 4 months ago (April 21, 2000)
Published 25 years, 4 months ago (April 21, 2000)
Published Print 25 years, 4 months ago (April 21, 2000)
Funders 0

None

@article{Turley_2000, title={Transport of Peptide-MHC Class II Complexes in Developing Dendritic Cells}, volume={288}, ISSN={1095-9203}, url={http://dx.doi.org/10.1126/science.288.5465.522}, DOI={10.1126/science.288.5465.522}, number={5465}, journal={Science}, publisher={American Association for the Advancement of Science (AAAS)}, author={Turley, Shannon J. and Inaba, Kayo and Garrett, Wendy S. and Ebersold, Melanie and Unternaehrer, Julia and Steinman, Ralph M. and Mellman, Ira}, year={2000}, month=apr, pages={522–527} }