Processing of a class I-restricted epitope from tyrosinase requires peptide N-glycanase and the cooperative action of endoplasmic reticulum aminopeptidase 1 and cytosolic proteases.
Although multiple components of the class I MHC processing pathway have been elucidated, the participation of nonproteasomal cytosolic enzymes has been largely unexplored. In this study, we provide evidence for multiple cytosolic mechanisms in the generation of an HLA-A*0201-associated epitope from tyrosinase. This epitope is presented in two isoforms containing either Asn or Asp, depending on the structure of the tyrosinase precursor. We show that deamidation of Asn to Asp is dependent on glycosylation in the endoplasmic reticulum (ER), and subsequent deglycosylation by peptide-N-glycanase in the cytosol. Epitope precursors with N-terminal extensions undergo a similar process. This is linked to an inability of ER aminopeptidase 1 to efficiently remove N-terminal residues, necessitating processing by nonproteasomal peptidases in the cytosol. Our work demonstrates that processing of this tyrosinase epitope involves recycling between the ER and cytosol, and an obligatory interplay between enzymes involved in proteolysis and glycosylation/deglycosylation located in both compartments.
Altrich-VanLith
Michelle L
ML
Carter Immunology Center and Department of Microbiology, University of Virginia, Charlottesville, VA 22908-1386, USA.
Ostankovitch
Marina
M
Polefrone
Joy M
JM
Mosse
Claudio A
CA
Shabanowitz
Jeffrey
J
Hunt
Donald F
DF
Engelhard
Victor H
VH
D000626:Aminopeptidases / Q000378:metabolism*; D017951:Antigen Presentation; D003600:Cytosol / Q000201:enzymology*; D004721:Endoplasmic Reticulum / Q000201:enzymology / Q000378:metabolism*; D000939:Epitopes / Q000276:immunology*; D006031:Glycosylation; D015234:HLA-A Antigens / Q000276:immunology; D015789:HLA-A2 Antigen; D015395:Histocompatibility Antigens Class I / Q000276:immunology*; D006801:Humans; D015778:Minor Histocompatibility Antigens; D014442:Monophenol Monooxygenase / Q000276:immunology*; D010447:Peptide Hydrolases / Q000378:metabolism*; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase / Q000378:metabolism*
D000939:Epitopes; D015234:HLA-A Antigens; C435939:HLA-A*02:01 antigen; D015789:HLA-A2 Antigen; D015395:Histocompatibility Antigens Class I; D015778:Minor Histocompatibility Antigens; D014442:Monophenol Monooxygenase; D010447:Peptide Hydrolases; D000626:Aminopeptidases; C509944:ERAP1 protein, mouse; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase