INVASION BY SALMONELLA-TYPHIMURIUM INDUCES INCREASED EXPRESSION OF THE LMP, MECL, AND PA28 PROTEASOME GENES AND CHANGES IN THE PEPTIDE REPERTOIRE OF HLA-B27

We have analyzed proteasomal adaptation and associated changes in the B27-bound peptide repertoire in response to cellular invasion with Sal monella. The peptide repertoire of HLA-B27 complexes was analyzed by t wo different methods: (i) high-pressure liquid chromatography (HPLC) p rofiles of newly synthesized peptides eluted from B27 following metabo lic labeling with arginine and (ii) reactivities with two B27 monoclon al antibodies, Ye-2 and B27.M2, sensitive to peptide-induced conformat ional changes. LMP, MECL, and PA28 expression was analyzed by reverse transcription-PCR (RT-PCR) of mRNA and by Western blot analysis for LM P2, Invasion of HLA-B27-transfected HeLa cells by Salmonella typhimuri um induced significant changes in the reactivities of HLA-B27 with the se two antibodies, which was accompanied by significant quantitative a nd qualitative changes in the HPLC profile of peptides eluted from HLA -B27. We also observed increases in the RT-PCR values for the LMP2, LM P7, and MECL proteasome subunit genes, as well as the proteasomal acti vator PA28 alpha and -beta genes, and increased expression of the LMP2 protein by Western blotting. Upregulation of LMP2, but not LMP7, gene expression showed a close correlation with the changes in antibody re activities observed upon bacterial invasion. We observed similar chang es in reactivity with the Ye-2 or the B27.M2 antibody of lymphoblastoi d cells upon gamma interferon treatment, which significantly correlate d with the increased RT-PCR values for the LMP2 gene. This was accompa nied by consistent HPLC profile changes for eluted peptides, Thus, Sal monella invasion leads to serologically recognizable changes in the B2 7-bound peptide repertoire, which may include peptides of host origin potentially through modulation of proteasome LMP2 subunit expression a nd, as a consequence, proteasomal activities.