Uma. Motal et al., MAJOR HISTOCOMPATIBILITY COMPLEX CLASS I-BINDING PEPTIDES ARE RECYCLED TO THE CELL-SURFACE AFTER INTERNALIZATION, European Journal of Immunology, 23(12), 1993, pp. 3224-3229
Cytotoxic T lymphocytes (CTL) recognize target antigens as short, proc
essed peptides bound to major histocompatibility complex class I (MHC-
I) heavy and light chains (beta2-microglobulin; beta2-M).The heavy cha
in,which comprise the actual peptide binding alpha-1 and alpha-2 domai
ns, can exist at the cell surface in different forms, either free, bou
nd to beta2-M or as a ternary complex with beta2-m and peptides. MHC-1
chains are also known to internalize, and recycle to the cell surface
, and this has been suggested to be important in peptide presentation.
Whether MHC-1-bound peptides also can recycle is not known.We have in
vestigated this by using both peptide transporter mutant RMA-S cells a
nd EL4 cells loaded with D(b)-binding peptides, by two different appro
aches. First, peptides were covalently linked with galabiose (Galalpha
4Gal) at a position which did not interfere with D(b) binding or immun
ogenicity, and peptide recycling tested with Gal2-specific monoclonal
antibodies. By flow cytometry, a return of Gal2 epitopes to the cell s
urface was found, after cellular internalization and cell surface clea
rance by pronase treatment. This peptide recycling could be discrimina
ted from free fluid-phase uptake and was inhibited by methylamine, chl
oroquine and low temperature (18-degrees-C) but not by leupeptin. Seco
nd, specific CTL were reacted with peptide-loaded target cells after c
omplete removal of surface D(b) molecules by pronase, and after differ
ent times of incubation at 37-degrees-C to allow reexpression. By this
procedure, reappearance of target cell susceptibility was confirmed.
The results are in agreement with a model for optimizing peptide prese
ntation by recycling through an intracellular compartment similar to e
arly endosomes in certain antigen-presenting cells.