Prominence of β₂-Microglobulin, Class I Heavy Chain Conformation, and Tapasin in the Interactions of Class I Heavy Chain with Calreticulin and the Transporter Associated with Antigen Processing

Newly synthesized class I heavy (H) chain/β₂m heterodimers awaiting peptides in the endoplasmic reticulum are associated with the transporter associated with Ag processing (TAP). We present evidence here that calreticulin, but not calnexin, displays steady state association with class I/TAP complexes. To separate the ability of β₂m to bind with TAP and calreticulin from that of H chain, we studied human cell lines that lack expression of β₂m or H chain. Little if any H chain was detected in association with TAP and calreticulin in the β₂m^- cell line Daudi. By contrast, high levels of β₂m are found with TAP and calreticulin in the H chain-deficient cell line LCL 721.221, even after preclearance of the trace amount of class IB protein expressed by this cell line. Thus, β₂m appears to bind TAP in the absence of H chain, providing an elegant mechanism to retain β₂m in the endoplasmic reticulum at the site of peptide loading. To investigate whether other molecules participate in the binding of β₂m and H chain to TAP and calreticulin, we analyzed the deletion mutant cell line LCL 721.220, which lacks tapasin. In 721.220, TAP and calreticulin are not associated with each other. Also, in these cells, H chain/β₂m are not associated with TAP, but H chain and a low level of β₂m are associated with calreticulin. These results suggest that tapasin is an obligatory mediator of the assemblage of calreticulin/H chain/β₂m with TAP.