Major Histocompatibility Complex Class I Viral Antigen Processing in the Secretory Pathway Defined by the trans-Golgi Network Protease Furin

Abstract

SummaryClassical antigen presentation by major histocompatibility complex class I molecules involves cytosolic processing of endogenously synthesized antigens by proteasomes and translocation of processed peptides into the endoplasmic reticulum (ER) by transporters associated with antigen presentation (TAP). Alternative pathways for processing of endogenous antigens, generally involving the ER, have been suggested but not fully proved. We analyzed the potential for class I presentation of proteolytic maturation of secretory antigens in the exocytic pathway. We found that hepatitis B (HB) virus secretory core protein HBe can efficiently deliver COOHterminally located antigenic peptides for endogenous class I loading in the absence of TAP. Antigen presentation to specific cytotoxic T lymphocytes correlates with protein maturation at the COOH terminus, since modification of maturation and transport of HBe through the secretory pathway alters antigen presentation. Both maturation and a necessary processing step occur in the Golgi or post-Golgi compartment. Antigen presentation is independent of proteasome activity, but inhibitors of the trans -Golgi network resident protease furin inhibit both HBe maturation and antigen presentation. These results define a new antigen processing pathway located in the secretory route, with a central role for proteolytic maturation mediated by the subtilisin protease family member furin as an efficient source for antigen presentation.

Peptide transporter TAP mediates between competing antigen sources generating distinct surface MHC class I peptide repertoires

“…However, processing by yet other proteolytic systems deliver peptides that are presented in a TAP-independent way. The above-mentioned SP and SPP proteases produce such TAPindependent peptides within the ER and proprotein convertases like PC7 and furin have been shown to facilitate TAP-independent presentation in the secretory route [29][30][31][32]. Interestingly, our preliminary data show that presentation of the Trh4 peptide is independent of these known enzyme systems, indicating that yet other pathways exist.…”

“…Peptides for which MHC-I binding and stability were analyzed were Trh4 379-387 (MCLRMTAVM), LCMV gp33 [33][34][35][36][37][38][39][40][41][42] [20][21][22][23][24][25][26][27][28] (TNLLNDRVL) and gp100 [25][26][27][28][29][30][31][32][33] (EGSRNQDWL). RMA-S cells were cultured for 2 days at 261C to accumulate peptide receptive MHC-I molecules on the cell surface [60].…”

“…Indeed, signal sequences are liberated by the combined action of signal peptidase (SP) and signal peptide peptidase (SPP) and are directly available for loading into MHC-I [28,29]. A second characterized processing pathway that bypasses TAP is active in the secretory route and is mediated by members of the proprotein convertase (PC) family, like furin [30][31][32]. This enzyme is located in the trans-Golgi network and mediates the proteolytic maturarion of many proproteins, e.g.…”

“…growth factors and matrix metalloproteinases [33]. Peptides located at the C-terminus of secreted proteins can be liberated by furin and subsequently gain access to MHC-I in a TAP-independent way [30,32].…”

“…growth factors and matrix metalloproteinases [33]. Peptides located at the C-terminus of secreted proteins can be liberated by furin and subsequently gain access to MHC-I in a TAP-independent way [30,32].Previously, we reported the TAP-independent presentation of a C-terminal peptide from the ceramide synthase Trh4, which is a multiple membrane-spanning protein in the ER [34][35][36]. This peptide was the first natural example of a C-terminal processing pathway of ER resident proteins, while previous studies suggested the existence of this route [37][38][39].…”