Abstract
Germinal center-associated DNA primase (GANP) associated with MCM3 of the DNA replication complex is up-regulated selectively in germinal center B cells. We studied promoter activity of the 5' region involved in the developmental stage-dependent expression in B lineage cells by luciferase reporter assay. Selective regulation of ganp expression was observed in the -737-bp promoter region in B and plasma cell lines but was significantly low in pre-B and T cell lines. The deletion constructs displayed a gap decrease after shortening the region from -134 to -108 bp. Further narrowing suggested the involvement of the PU.1 consensus sequence at -126 bp by electrophoretic mobility shift assay. The protein component PU.1 complex is not inhibited with mutated probes at the consensus site but is inhibited with the known PU.1 probe of CD72 and with anti-PU.1 antibody. Moreover, introduction of PU.1 cDNA enhanced the reporter gene activity in a dose-dependent manner in B cells, whereas the reporter construct with the mutated PU.1 site did not respond. Anti-CD40 stimulation induced the reporter activity with a 100% increase, which is not observed with the PU.1-mutated reporter construct. These results demonstrate that the germinal center-associated DNA primase expression is partly regulated by the transcription factor PU.1 expressed in B lineage cells.
Highlights
Immunization with T cell-dependent antigen induces proliferation of specific B cell clones in the germinal center (GC)1 region of secondary lymphoid follicles of the spleen and lymph nodes [1, 2]
These results demonstrate that the germinal center-associated DNA primase expression is partly regulated by the transcription factor PU.1 expressed in B lineage cells
During the differentiation of GC-B cells, various molecular events appear to be associated with an effective Ab production, some of which are probably closely associated with cell cycle progression that is inevitable for clonal expansion of antigenreactive B cell clones
Summary
Immunization with T cell-dependent antigen induces proliferation of specific B cell clones in the germinal center (GC)1 region of secondary lymphoid follicles of the spleen and lymph nodes [1, 2]. We studied promoter activity of the 5 region involved in the developmental stage-dependent expression in B lineage cells by luciferase reporter assay.
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