Initiation of DNA replication

噬菌体T4长期以来一直作为研究DNA复制、转录和重组的模型，其相对较小的尺寸和明确的生物化学使其成为将体外生物化学整合到染色体背景中的极好的模型生物。 作为表征T4感染期间染色体交易动态的第一步，我们采用了一组独特的宏阵列策略来鉴定病毒DNA合成的起源，并在真实的时间内监测新生DNA在整个基因组中的实际积累。 我们已经发现，T4 DNA合成起源于至少五个离散的基因座在一个单一的人口感染的细胞，附近的奥里亚，oriC，oriE，oriF，和oriG。这是第一个直接的证据，多个活跃的起源在一个单一的人口T4感染的细胞，这意味着T4将提供洞察到其他多起源系统，如人类。 从起源基因座产生的新生DNA在空间和时间上受到调节，导致多个短DNA在起源附近的积累，这些DNA后来延伸到全长基因组。 这种复制模式似乎对染色体功能至关重要，而改变复制动力学的突变也会改变整个基因组的转录模式，这表明这两个过程是相互依赖的。事实上，当转录在感染后期被阻止时，用于启动复制的起点发生变化。据推测，复制和转录之间的相互作用塑造了T4基因组，我们的工作为未来表征T4基因组功能的分子动力学提供了基础，并为多起源染色体的进化提供了见解。

The bacteriophage T4 has long served as a model for the study of DNA replication, transcription, and recombination, and its relatively small size and well defined biochemistry make it an excellent model organism in which to integrate in vitro biochemistry into a chromosomal context. As a first step in characterizing the dynamics of chromosomal transactions during T4 infection, we employed a unique set of macro array strategies to identify the origins of viral DNA synthesis and monitor the actual accumulation of nascent DNA across the genome in real time. We have found that T4 DNA synthesis originates from at least five discrete loci within a single population of infected cells, near oriA, oriC, oriE, oriF, and oriG. This is the first direct evidence of multiple active origins within a single population of T4-infected cells, and it implies that T4 will provide insights into other multi-origin systems, like humans. The nascent DNAs produced from origin loci are regulated spatially and temporally, leading to the accumulation of multiple, short DNAs near the origins, which are later extended to full length genomes. This pattern of replication appears pivotal to the chromosome function, and mutations that alter replication dynamics also alter transcription patterns across the genome, suggesting that the two processes are dependent on one another. Indeed, when transcription is prevented at later times during infection, the origins used to initiate replication change. Presumably this interplay between replication and transcription has shaped the T4 genome, and our work provides the foundation for the future characterization of the molecular dynamics that contribute to T4 genome function and provide insights into the evolution of multi origin chromosomes.