Aj. Yellenshaw et al., MURINE TRANSPORTER ASSOCIATED WITH ANTIGEN PRESENTATION (TAP) PREFERENCES INFLUENCE CLASS I-RESTRICTED T-CELL RESPONSES, The Journal of experimental medicine, 186(10), 1997, pp. 1655-1662
The transporter associated with antigen presentation (TAP) complex shu
ttles cytosolic peptides into the exocytic compartment for association
with nascent major histocompatibility complex class I molecules. Bioc
hemical studies of murine and human TAP have established that substrat
e length and COOH-terminal residue identity are strong determinants of
transport efficiency. However, the existence of these specificities i
n the intact cell and their influences on T cell responses have not be
en demonstrated. We have devised a method for studying TAP-mediated tr
ansport in intact cells, using T cell activation as a readout. The app
roach makes use of a panel of recombinant vaccinia viruses expressing
peptides containing the K-d-restricted nonamer influenza nucleoprotein
residues 147-155. The COOH terminus of each construct was appended wi
th a dipeptide composed of an internal threonine residue followed by a
varying amino acid. Synthetic peptide versions of these 11-mers exhib
it vastly different transport capabilities in streptolysin O-permeabil
ized cells, in accordance with thr predicted influence of the COOH-ter
minal residues. Presentation of the endogenously expressed version of
each construct requires TAP-mediated transport and cooexpression with
a vac-encoded exocytic COOH-terminal dipeptidase, angiotensin converti
ng enzyme, to allow liberation of the minimal epitope. Recognition by
epitope-specific CTLs therefore signifies TAP-mediated transport of a
complete 11-mer within the target cell. Under normal assay conditions
no influences of the COOH-terminal residue were revealed. However, whe
n T cell recognition was limited, either by blocking CD8 coreceptor in
teractions or by decreasing the amount of transport substrate synthesi
zed, significant COOH-terminal effects were revealed. Under such condi
tions, those peptides that transported poorly in biochemical assays we
re less efficiently presented. Therefore, TAP specificity operates in
tile intact cell, appears to reflect previously defined rules with reg
ard to the influence of thr COOH-terminal residue, and can strongly in
fluence T cell responses.