Immunogenetics (1999) 49 : 931–941 Q Springer-Verlag 1999 ORIGINAL PAPER Mario Lobigs 7 Arno Müllbacher 7 Robert V. Blanden Günter J. Hämmerling 7 Frank Momburg Antigen presentation in Syrian hamster cells: substrate selectivity of TAP controlled by polymorphic residues in TAP1 and differential requirements for loading of H2 class I molecules Received: 15 February 1999 / Revised: 20 May 1999 M. Lobigs (Y) 7 A. Müllbacher 7 R.V. Blanden Division of Immunology and Cell Biology, John Curtin School of Medical Research, The Australian National University, Canberra, P O Box 334, A.C.T. 2601, Australia e-mail: Mario.Lobigs6anu.edu.au, Tel.: 61-6249-4048, Fax: 61-6249-2595 G.J. Hämmerling 7 F. Momburg Department of Molecular Immunology, German Cancer Research Center, Heidelberg, Germany Abstract Expression of mouse major histocompatibili- ty complex (MHC) class I molecules in different cell lines derived from Syrian hamsters has revealed antig- en presentation deficiencies of some H2 allelic products in two cell lines (BHK and NIL-2) which were over- come by transient expression of the rat transporter as- sociated with antigen processing (TAP; Lobigs et al. 1995). Here we show that in both cell lines the endoge- nous MHC class I cell surface expression was complete- ly down-regulated. Lymphokine treatment induced en- dogenous and recombinant mouse MHC class I cell sur- face expression to levels similar to that in other Syrian hamster cell lines competent for antigen presentation through transduced H2 molecules. Accordingly, consti- tutive downregulation of expression of accessory mole- cules of the MHC class I pathway can reveal differences between H2 class I alleles in antigen presentation not encountered when the expression levels are aug- mented. In addition to the differential expression of MHC class I pathway genes, two cell lines representing competent (FF) and defective (BHK) antigen presenta- tion phenotypes for mouse class I MHC restriction ele- ments demonstrated substantial sequence polymor- phism in Tap1 but not Tap2. Cytokine-treated FF or BHK cells and human TAP-deficient T2 cells trans- fected with FF or BHK TAP1 in combination with FF TAP2 differed in their preference for C-terminal pep- tide residues, as shown by an in vitro peptide transport assay. Thus, polymorphic residues in TAP1 can in- fluence the substrate selectivity of the Syrian hamster peptide transporter. Key words Syrian Hamster 7 MHC 7 Antigen presentation 7 Peptide transporters (TAP) Introduction Major histocompatibility complex (MHC) class I antig- ens are exceptionally polymorphic in many species (Bjorkman and Parham 1990; Klein and Figueroa 1981). The polymorphic amino acid residues are clus- tered in the region of the MHC class I molecule in- volved in peptide binding, which is the basis for their different peptide binding motifs (Bjorkman et al. 1987; Rammensee et al. 1993). This led to the view that the MHC class I polymorphism has evolved as a strategy to prevent pathogen escape from cytotoxic T (Tc) cell sur- veillance (Zinkernagel and Doherty 1975). This strate- gy requires that the additional components of the class I antigen processing and presentation pathway which are needed for peptide loading can accommodate the diverse peptide ligand specificities of the polymorphic MHC class I molecules. Peptide translocation from the cytoplasm into the lu- men of the endoplasmic reticulum (ER) for assembly with MHC class I molecules is accomplished by an ATP-driven peptide transporter, a heterodimer consist- ing of TAP1 and TAP2 proteins (Deverson et al. 1990; Kelly et al. 1992; Monaco et al. 1990; Spies et al. 1990, 1992; Trowsdale et al. 1990). From antigen presentation studies (Lobigs and Müllbacher 1993; Yewdell et al. 1993) and in vitro peptide translocation assays (Andro- lewicz and Cresswell 1994; Obst et al. 1995) it is appar- ent that TAP has a broad substrate specificity. It ap- pears that only the C-terminal peptide residue exerts a pronounced influence on in vitro peptide transport rates. Mouse TAP was found to selectively transport peptides with hydrophobic C-terminal residues while human TAP is somewhat non-selective (Momburg et al. 1994b; Schumacher et al. 1994a). The optimal pep- tide length for in vitro transport (8–15 amino acids; Heemels and Ploegh 1994; Momburg et al. 1994a) con-