Interaction between heat-shock protein 73 and HLA-DRB1 alleles associated or not with rheumatoid arthritis.

Abstract

HLA-DRB1 alleles whose third hypervariable region contains a QKRAA/QRRAA/RRRAA motif are associated with rheumatoid arthritis (RA) through unknown mechanisms. We previously demonstrated that the QKRAA motif was also expressed on the Escherichia coli 40-kd heat-shock protein (HSP) DnaJ. The QKRAA motif helps DnaJ bind its partner chaperone, the E coli 70-kd HSP DnaK. Furthermore, we observed that in lymphoblastoid cells, Hsp73, the constitutive 70-kd HSP, associates with HLA-DRB1*0401 (an allele with a QKRAA motif) and targets it to lysosomes. In this study, we sought to classify different HLA-DRB1 alleles according to their ability to bind Hsp73. To evaluate how well different HLA-DRB1 alleles could bind Hsp73, we developed a quantitative precipitation assay and a direct binding assay. Quantitative precipitation assay from total cellular proteins and from lysosomal extracts demonstrated that RA-associated HLA-DRB1 alleles bound Hsp73 better than did HLA-DRB1 alleles that were not associated with RA. HLA-DRB1*0401 was the best Hsp73 binder. These findings were confirmed by direct binding assay between purified proteins. HLA-DRB1*0401 was the best Hsp73 binder among the 8 different HLA-DRB1 alleles that were tested.