Peptide binding to surface class II molecules is the major pathway of formation of immunogenic class II-peptide complexes for viable antigen presenting cells.
Although studies on fixed APCs have demonstrated that peptide can bind to cell surface class II molecules, the mechanisms by which peptide-class II complexes are formed in viable cells is largely unexplored. To explore the possibility that peptide loading of class II molecules was occurring after endocytosis of peptides as well as by surface binding, we utilized an immunogenic hemagglutinin peptide (HAP 128-145) from the influenza strain A/Japan/57, and studied the appearance of surface complexes of HAP 128-145 bound to HLA-DRw11 molecules on human B-lymphoblastoid cells (BLCLs). Detection of the bound peptide was made possible by a rabbit anti-serum (alpha HAP) raised against HAP 128-145, which recognizes both the free peptide as well as peptide bound to DRw11 on living APCs. Pretreatment of the BLCLs with a variety of inhibitors of protein synthesis and intracellular trafficking failed to decrease the levels of HAP 128-145/DRw11 surface complexes. However, significant inhibition in the appearance of these complexes was caused by a decrease in the temperature at which the cells were incubated with peptide. Temperature-specific inhibition was also observed for fixed DRw11-positive APCs and purified DRw11 molecules indicating that the effect of temperature was directly on the class II molecules. We conclude that surface binding of peptide by class II molecules on human B cells is a major pathway of formation of immunogenic class II-peptide complexes for at least some soluble antigenic peptides, and that endocytosis of soluble peptides with subsequent binding of peptide by intracellular class II molecules plays little if any role in the formation of such complexes. Moreover, class II molecules have evolved to stably bind peptide optimally at physiologic temperatures, independent of cell metabolism.
Nygard
N R
NR
Department of Medicine, Washington University School of Medicine, St. Louis, MO 63110.
Giacoletto
K S
KS
Bono
C
C
Gorka
J
J
Kompelli
S
S
Schwartz
B D
BD
D000595:Amino Acid Sequence; D000938:Antigen-Presenting Cells / Q000276:immunology*; D000956:Antigens, Viral / Q000737:chemistry / Q000276:immunology; D006684:HLA-DR Antigens / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D009980:Influenza A virus / Q000276:immunology; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000276:immunology*; D011485:Protein Binding; D012995:Solubility; D013696:Temperature
D000956:Antigens, Viral; D006684:HLA-DR Antigens; D010455:Peptides