Need for Tripeptidyl-peptidase II in Major Histocompatibility Complex Class I Viral Antigen Processing when Proteasomes are Detrimental

Abstract

CD8؉ T lymphocytes recognize infected cells that display virus-derived antigenic peptides complexed with major histocompatibility complex class I molecules. Peptides are mainly byproducts of cellular protein turnover by cytosolic proteasomes. Cytosolic tripeptidyl-peptidase II (TPPII) also participates in protein degradation. Several peptidic epitopes unexpectedly do not require proteasomes, but it is unclear which proteases generate them. We studied antigen processing of influenza virus nucleoprotein epitope NP 147-155 , an archetype epitope that is even destroyed by a proteasome-mediated mechanism. TPPII, with the assistance of endoplasmic reticulum trimming metallo-aminopeptidases, probably ERAAP (endoplasmic reticulum aminopeptidase associated with antigen processing), was crucial for nucleoprotein epitope generation both in the presence of functional proteasomes and when blocked by lactacystin, as shown with specific chemical inhibitors and gene silencing. Different protein contexts and subcellular targeting all allowed epitope processing by TPPII as well as trimming. The results show the plasticity of the cell's assortment of proteases for providing ligands for recognition by antiviral CD8 ؉ T cells. Our observations identify for the first time a set of proteases competent for antigen processing of an epitope that is susceptible to destruction by proteasomes.

Differential presentation of tumor antigen‐derived epitopes by MHC‐class I and antigen‐positive tumor cells

“…39 Alternative cytosolic pathways have also been reported and the processing proteases in these cases can be cysteine proteases 40 or tripeptidyl peptidase II. 41,42 Translocation of peptides into the ER is provided by transporters associated with antigen processing (TAP), but in some cases may also be TAP-independent, carried out by the sec61 translocon or by undefined mechanisms. 43 The exact pathway, which is involved in processing the epitope SSX2 103-111 has not been elucidated.…”

“…We used several antigen presenting cells to demonstrate that the Fab-VIE1 could bind to the specific MHC-peptide complex not only in its recombinant soluble form, but also in the native form expressed on the cell surface. Soluble Fab-VIE1 reacted with SSX2 103-111 peptide loaded T2 cells and HLA-A*0201 1 EBV-transformed lymphoblastoid cell line, but not with cells loaded with one of 6 different HLA-A*0201-binding peptides, SSX2 [41][42][43][44][45][46][47][48][49] , NY-ESO-1 157-165 , influenza matrix protein 58-66 , gp100 209-217 , Melan-A [26][27][28][29][30][31][32][33][34][35] and Carboanhydrase-IV 254-262 , respectively. No binding of Fab-VIE1 to the HLA-A*0201 2 EBV-transformed B-lymphocytes was observed, neither when pulsed with the SSX2 103-111 peptide nor with control peptides (Fig.…”

“…Specificity of selected Fab-VIE1 was demonstrated. In ELISA Fab-VIE1 reacted specifically with the SSX2 103-111 containing HLA-A*0201 complex, but not with 4 control complexes, containing HLA-A*0201-restricted epitopes of Melan-A, CMV pp65, NY-ESO-1 and SSX2 [41][42][43][44][45][46][47][48][49] respectively. Importantly, cross-reactivity to SSX2 41-49 / HLA-A*0201 complex could be excluded (Fig.…”

Endoplasmic Reticulum Targeting Alters Regulation of Expression and Antigen Presentation of Proinsulin

“…Tunicamycin (5 mg/ml), brefeldin A (10 mg/ml), and cycloheximide (CHX; 20 mg/ml) were purchased from Sigma (St. Louis, MO). Butabindide oxalate (250 mM) was purchased from Tocris (Bristol, U.K.) and added each 90-120 min following published protocols (21). The inhibitor concentrations used are indicated in parentheses.…”

“…This was achieved by 24-h incubation with Doxy followed by a brief acid treatment to remove cell-surface K d -insulin B 15-23 complexes. The generation of new K d -insulin B [15][16][17][18][19][20][21][22][23] complexes from previously synthesized PPI, or from newly synthesized PI, in the presence of protease inhibitors was then monitored.…”