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 Salmonella. The peptide repertoire of HLA-B27 complexes was analyzed by two different methods: (i) high-pressure liquid chromatography (HPLC) profiles of newly synthesized peptides eluted from B27 following metabolic labeling with arginine and (ii) reactivities with two B27 monoclonal antibodies, Ye-2 and B27.M2, sensitive to peptide-induced conformational changes. LMP, MECL, and PA28 expression was analyzed by reverse transcription-PCR (RT-PCR) of mRNA and by Western blot analysis for LMP2. Invasion of HLA-B27-transfected HeLa cells by Salmonella typhimurium induced significant changes in the reactivities of HLA-B27 with these two antibodies, which was accompanied by significant quantitative and qualitative changes in the HPLC profile of peptides eluted from HLA-B27. We also observed increases in the RT-PCR values for the LMP2, LMP7, and MECL proteasome subunit genes, as well as the proteasomal activator PA28alpha 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 reactivities observed upon bacterial invasion. We observed similar changes in reactivity with the Ye-2 or the B27.M2 antibody of lymphoblastoid cells upon gamma interferon treatment, which significantly correlated with the increased RT-PCR values for the LMP2 gene. This was accompanied by consistent HPLC profile changes for eluted peptides. Thus, Salmonella invasion leads to serologically recognizable changes in the B27-bound peptide repertoire, which may include peptides of host origin potentially through modulation of proteasome LMP2 subunit expression and, as a consequence, proteasomal activities.
Maksymowych
W P
WP
Departments of Medicine and Medical Microbiology and Immunology, University of Alberta, Edmonton, Canada. maksymow@gpu.srv.ualberta.ca
Ikawa
T
T
Yamaguchi
A
A
Ikeda
M
M
McDonald
D
D
Laouar
L
L
Lahesmaa
R
R
Tamura
N
N
Khuong
A
A
Yu
D T
DT
Kane
K P
KP
D017951:Antigen Presentation; D003546:Cysteine Endopeptidases / Q000096:biosynthesis / Q000378:metabolism*; D003602:Cytotoxicity, Immunologic; D015971:Gene Expression Regulation, Enzymologic; D015796:HLA-B27 Antigen / Q000276:immunology*; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology; D009097:Multienzyme Complexes / Q000378:metabolism*; D009124:Muscle Proteins; D010455:Peptides / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D011485:Protein Binding; D014176:Protein Biosynthesis; D011487:Protein Conformation; D012486:Salmonella typhimurium / Q000276:immunology* / Q000472:pathogenicity; D014407:Tumor Cells, Cultured
D015796:HLA-B27 Antigen; D009097:Multienzyme Complexes; D009124:Muscle Proteins; C085047:PSME1 protein, human; D010455:Peptides; C071313:LMP-2 protein; D007371:Interferon-gamma; D003546:Cysteine Endopeptidases; C077436:LMP7 protein; C098948:PSMB10 protein, human; D046988:Proteasome Endopeptidase Complex