K-b, K-d, and L-d molecules share common tapasin dependencies as determined using a novel epitope tag

The endoplasmic reticulum protein tapasin is considered to be a class I-ded icated chaperone because it facilitates peptide loading by proposed mechani sms such as peptide editing, endoplasmic reticulum retention of nonpeptide- bound molecules, and/or localizing class I near the peptide source. Nonethe less, the primary functions of tapasin remain controversial as do the relat ive dependencies of different class I molecules on tapasin for optimal pept ide loading and surface expression, Tapasin dependencies have been addresse d in previous studies by transfecting different class I alleles into tapasi n-deficient LCL721.220 cells and then monitoring surface expression and Ag presentation to T cells. Indeed, by these criteria, class I alleles have di sparate tapasin-dependencies. In this study, we report a novel and more dir ect method of comparing tapasin dependency by monitoring the ratio of folde d vs open forms of the different mouse class I heavy chains, L-d, K-d and K -b. Furthermore, we determine the amount of de novo heavy chain synthesis r equired to attain comparable expression in the presence vs absence of tapas in, Our findings show that tapasin dramatically improves peptide loading of all three of these mouse molecules.