PROCESSING OF MAJOR HISTOCOMPATIBILITY CLASS-I - RESTRICTED ANTIGENS IN THE ENDOPLASMIC-RETICULUM

We have introduced long precursor peptides directly into the endoplasm ic reticulum (ER) of a mutant cell line (T2-D-b) that lacks the abilit y to transport peptides from the cytosol to the ER in a transporter as sociated with antigen processing (TAP)-dependent way. This was done by expressing various influenza A-derived peptides containing the natura lly processed epitope ASNENMDAM (366-374) preceded by the influenza he magglutinin ER translocation sequence. Peptides derived from these min igenes that became associated with D-b were isolated and identified by combined reversed phase liquid chromatography and detection by cytoto xic T lymphocytes. Our results establish that NH2-terminal extensions of at least 40 residues can be trimmed from peptides entering the ER, but that proteolysis of larger proteins may be limited.