N-linked carbohydrates in tyrosinase are required for its recognition by human MHC class II-restricted CD4(+) T cells

Housseau, F Moorthy, A Langer, DA Robbins, PF Gonzales, MI Topalian, SL

F. Housseau et al., N-linked carbohydrates in tyrosinase are required for its recognition by human MHC class II-restricted CD4(+) T cells, EUR J IMMUN, 31(9), 2001, pp. 2690-2701

40

INGLESE

Article

Immunology

EUROPEAN JOURNAL OF IMMUNOLOGY

0014-2980 → ACNP

31

9

2001

2690 - 2701

ISI

0014-2980(200109)31:9<2690:NCITAR>2.0.ZU;2-9

Glycosylation of mammalian proteins is known to influence their intracellul ar trafficking, half life, and susceptibility to enzymatic degradation. Rar e instances of natural T cell epitopes dependent upon glycosylation for rec ognition have been described. We report here on human CD4(+) T lymphocyte. cultures and clones from two melanoma patients that recognize the melanoma- associated Ag tyrosinase in the context of HLA-DR4 and -DR8. These T cells recognize tyrosinase, normally a heavily glycosylated molecule, when expres sed constitutively in melanoma cells or in COS-7 transfectants pulsed as ly sates onto autologous APC. However, these T cells fail to recognize tyrosin ase expressed in bacteria, nor do they react with overlapping peptides cove ring full-length tyrosinase, suggesting a critical role for glycosylation i n the processing and/or composition of the stimulatory epitopes. The requir ement for glycosylation was demonstrated by the failure of tyrosinase-speci fic CD4(+) T cells to recognize tyrosinase synthesized in the presence of g lycosylation inhibitors, or deglycosylated enzymatically. Site-directed mut agenesis of each of seven potential N-glycosylation sites showed that four sites were required to generate forms of tyrosinase that could be recognize d by individual T cell clones. These data indicate that certain carbohydrat e moieties are required for processing the tyrosinase peptides recognized b y CD4(+) T cells. Posttranslational modifications of human tumor-associated proteins such as tyrosinase could be a critical factor for the development of antitumor immune responses.