Absence of functional relevance of human transporter associated with antigen processing polymorphism for peptide selection.
The polymorphic TAP (TAP1 and TAP2) genes code for a heteromeric complex that translocates cytosolic peptides across the membrane of the endoplasmic reticulum where they associate with MHC class I molecules. In the rat, extensive TAP2 polymorphism modifies the spectrum of peptides presented by MHC class I molecules on the cell surface. Although polymorphism of human TAP proteins is more limited, it has been proposed to be associated with susceptibility to autoimmune diseases. To analyze the peptide selectivity and transport efficiency of human TAP alleles, we have generated the most frequent TAP1 and TAP2 alleles using site-directed mutagenesis. Alleles were cloned in the baculovirus expression system and six TAP1/2 allelic combinations were overexpressed in Sf9 insect cells. Microsomes containing TAP1/2 dimers were isolated, and peptide binding assays were performed using random sequence peptide libraries of variable length (7-18 mer) and a 9-mer polyalanine peptide substituted with various amino acids in positions 1, 2, 3, 7, and 9. All studied allelic combinations selected peptides with identical substitutions and length. Furthermore, transport assays showed similar efficiency and kinetics of peptide translocation by the different TAP alleles. These data suggest that the major human TAP alleles transport identical peptide sets and are therefore unlikely to contribute to predisposition to autoimmune diseases.
Daniel
S
S
INSERM Unit 25, Hôpital Necker, Paris, France.
Caillat-Zucman
S
S
Hammer
J
J
Bach
J F
JF
van Endert
P M
PM
D000071181:ATP Binding Cassette Transporter, Subfamily B, Member 2; D000071450:ATP Binding Cassette Transporter, Subfamily B, Member 3; D018528:ATP-Binding Cassette Transporters / Q000235:genetics* / Q000502:physiology; D000483:Alleles; D000818:Animals; D017951:Antigen Presentation; D001327:Autoimmune Diseases / Q000235:genetics / Q000276:immunology; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004198:Disease Susceptibility; D006801:Humans; D008861:Microsomes; D016297:Mutagenesis, Site-Directed; D017924:Nucleopolyhedroviruses / Q000235:genetics; D010446:Peptide Fragments / Q000276:immunology; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism; D018411:Spodoptera / Q000166:cytology; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D000071181:ATP Binding Cassette Transporter, Subfamily B, Member 2; D000071450:ATP Binding Cassette Transporter, Subfamily B, Member 3; D018528:ATP-Binding Cassette Transporters; D010446:Peptide Fragments; D011994:Recombinant Proteins; C073119:TAP1 protein, human; C486172:Tap1 protein, rat; C489514:Tap2 protein, rat; C071692:TAP2 protein, human