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Transcription Factor Binding and Induced Transcription Alter Chromosomal c-myc Replicator Activity

机译：转录因子结合和诱导转录改变染色体c-myc复制子活性。

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The observation that transcriptionally active genes generally replicate early in S phase and observations of the interaction between transcription factors and replication proteins support the thesis that promoter elements may have a role in DNA replication. To test the relationship between transcription and replication we constructed HeLa cell lines in which inducible green fluorescent protein (GFP)-encoding genes replaced the proximal ～820-bp promoter region of the c-myc gene. Without the presence of an inducer, basal expression occurred from the GFP gene in either orientation and origin activity was restored to the mutant c-myc replicator. In contrast, replication initiation was repressed upon induction of transcription. When basal or induced transcription complexes were slowed by the presence of α-amanitin, origin activity depended on the orientation of the transcription unit. To test mechanistically whether basal transcription or transcription factor binding was sufficient for replication rescue by the uninduced GFP genes, a GAL4p binding cassette was used to replace all regulatory sequences within ～1,400 bp 5′ to the c-myc gene. In these cells, expression of a CREB-GAL4 fusion protein restored replication origin activity. These results suggest that transcription factor binding can enhance replication origin activity and that high levels of expression or the persistence of transcription complexes can repress it.
机译：转录活性基因通常在S期早期复制的观察结果以及转录因子与复制蛋白之间相互作用的观察结果支持了启动子元件可能在DNA复制中起作用的观点。为了测试转录与复制之间的关系，我们构建了HeLa细胞系，其中可诱导绿色荧光蛋白（GFP）编码基因取代了c- myc 基因的近端820 bp启动子区域。在没有诱导物的情况下，GFP基因在任一方向上都发生了基础表达，并且起源活性恢复到了突变的c- myc 复制子中。相反，在转录诱导后复制起始受到抑制。当α-amanitin的存在使基础或诱导的转录复合物减慢时，起源活性取决于转录单位的方向。为了机械地测试基础转录或转录因子结合是否足以通过未诱导的GFP基因进行复制挽救，使用了GAL4p结合盒来替换c- myc 约1400 bp 5'内的所有调控序列。基因。在这些细胞中，CREB-GAL4融合蛋白的表达恢复了复制起点的活性。这些结果表明，转录因子结合可以增强复制起点的活性，而高水平的表达或转录复合物的持久性可以抑制它。

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