RESUMO
The MHC class II-like molecule HLA-DM (DM) (H-2M in mice) catalyzes the exchange of CLIP for antigenic peptides in the endosomes of APCs. HLA-DO (DO) (H-2O in mice) is another class II-like molecule that is expressed in B cells, but not in other APCs. Studies have shown that DO impairs or modifies the peptide exchange activity of DM. To further evaluate the role of DO in Ag processing and presentation, we generated transgenic mice that expressed the human HLA-DOA and HLA-DOB genes under the control of a dendritic cell (DC)-specific promoter. Our analyses of DCs from these mice showed that as DO levels increased, cell surface levels of A(b)-CLIP also increased while class II-peptide levels decreased. The presentation of some, but not all, exogenous Ags to T cells or T hybridomas was significantly inhibited by DO. Surprisingly, H-2M accumulated in DO-expressing DCs and B cells, suggesting that H-2O/DO prolongs the half-life of H-2M. Overall, our studies showed that DO expression impaired H-2M function, resulting in Ag-specific down-modulation of class II Ag processing and presentation.
Assuntos
Apresentação de Antígeno/imunologia , Células Dendríticas/imunologia , Antígenos HLA-D/fisiologia , Actinas/imunologia , Animais , Apresentação de Antígeno/genética , Antígenos/imunologia , Antígenos de Diferenciação de Linfócitos B/imunologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Antígeno CD11c/genética , Células Cultivadas , Células Dendríticas/metabolismo , Genes Sintéticos , Antígenos H-2/imunologia , Antígenos H-2/metabolismo , Antígenos HLA-D/biossíntese , Antígenos HLA-D/genética , Antígenos de Histocompatibilidade Classe II/imunologia , Antígenos de Histocompatibilidade Classe II/metabolismo , Humanos , Hibridomas/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Muramidase/imunologia , Ovalbumina/imunologia , Fragmentos de Peptídeos/imunologia , Regiões Promotoras Genéticas/genética , Proteínas Recombinantes de Fusão/fisiologia , Microglobulina beta-2/imunologiaRESUMO
The class II transactivator (CIITA) regulates expression of the classical and non-classical MHC class II genes, HLA-DR, -DP, -DQ and -DM, but not the B cell-specific HLA-DO (DO). Here we show that only HLA-DR expression is completely dependent on CIITA, since residual expression of HLA-DM, -DP and the beta chain of DQ was observed in CIITA-deficient RJ2.2.5 cells. Although DO shows a unique expression pattern compared to other MHC class II genes, prolonged IFN-gamma treatment of HeLa cells induced DOB expression. Similar to all MHC class II promoters, the DOB promoter contains the highly conserved W, X1, and Y boxes in addition to a putative OCT box. Mutational analysis of the DOB promoter demonstrated that the X1, Y and OCT boxes are necessary for maximum promoter activity.Furthermore, our results demonstrate that CREB-1, RFXANK and Oct-2 occupy the DOB promoter in vivo, However, CIITA and Bob-1 were only minimally recruited. Finally, fusion of Bjab, a DOB-negative B cell line, with.174 B cells that lack the complete MHC class II region (including the DO genes), lead to DO expression. These data indicate that the expression of DO is regulated by an unidentified factor in B cells.
Assuntos
Regulação da Expressão Gênica , Antígenos HLA-D/genética , Antígenos de Histocompatibilidade Classe II/genética , Proteínas Nucleares , Transcrição Gênica , Adjuvantes Imunológicos/farmacologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico , Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Antígenos HLA-D/efeitos dos fármacos , Células HeLa , Antígenos de Histocompatibilidade Classe II/efeitos dos fármacos , Humanos , Interferon gama/farmacologia , Fator 2 de Transcrição de Octâmero , Regiões Promotoras Genéticas , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica/efeitos dos fármacosRESUMO
Peptide acquisition by MHC class II molecules is catalyzed by HLA-DM (DM). In B cells, HLA-DO (DO) inhibits or modifies the peptide exchange activity of DM. We show here that DO protein levels are modulated during B cell differentiation. Remarkably, germinal center (GC) B cells, which have low levels of DO relative to naive and memory B cells, are shown to have enhanced antigen presentation capabilities. DM protein levels also were somewhat reduced in GC B cells; however, the ratio of DM to DO in GC B cells was substantially increased, resulting in more free DM in GC B cells. We conclude that modulation of DM and DO in distinct stages of B cell differentiation represents a mechanism by which B cells regulate their capacity to function as antigen-presenting cells. Efficient antigen presentation in GC B cells would promote GC B cell-T cell interactions that are essential for B cells to survive positive selection in the GC.
