Recombination and the clearance of replicative blocks - to bypass or not to bypass?

重组和复制块的清除——绕过还是不绕过？

• 批准号: BB/J014826/1
• 负责人: Peter McGlynn
• 金额: $ 40.73万
• 依托单位: University of York
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2013
• 资助国家: 英国
• 起止时间: 2013 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FJ014826%2F1
• 关键词: Recombination; clearance; replicative; blocks; bypass; Recombination; clearance; replicative; blocks; bypass

The ability of a cell to form two new daughter cells allows organisms to grow and reproduce. This process requires the copying of vast amounts of DNA so that each daughter receives an accurate copy of all the genetic information required for survival. Because of the importance of generating accurate copies of the DNA, organisms have evolved very complex DNA replication machines that reduce the chances of mistakes being made. Unfortunately, we now know that many obstacles are encountered by these replication machines that, if not overcome, can cause mistakes in the copying process. Such mistakes can lead to errors in the genetic information that can have fatal consequences.Proteins that coat the DNA form frequent obstacles to the DNA copying machinery. These proteins are essential in all organisms for maintaining, reading and packing the genetic information, and so cannot be avoided. Although the DNA replication machinery can successfully push off most of these proteins from the DNA to be copied, the sheer number of proteins bound to the DNA mean that occasionally the copying process is stopped in its tracks. However, a specific class of enzymes can repair and restart broken down DNA replication machines. It is clear that these recombination enzymes can help to copy DNA that is bound by proteins but how they do so remains unclear. One proposal is that if a DNA replication machine becomes blocked by a protein then recombination enzymes can restart DNA replication on the other side of the protein block. This pathway would skip over the block, allowing copying to continue, but would also result in genes near the protein block not being copied. Consequently the involvement of recombination enzymes could be seen as harmful, resulting in failure to copy all the genetic information needed by the two daughter cells to survive. Alternatively, we have proposed that recombination enzymes might simply restart DNA replication near to where it initially came to a halt. This might give the DNA replication machinery a second chance to push the blocking protein off the DNA and continue to copy all of the genetic information. Such a process might therefore provide a mechanism to ensure accurate copying of DNA coated with proteins.We will use the model bacterium E. coli to determine the roles of recombination enzymes in copying DNA coated with proteins. We know a great deal about the basic mechanisms of both DNA replication and recombination in E. coli, allowing us to analyse how these two very complicated processes interact. We will determine the mechanisms by which recombination enzymes can help DNA replication machines to move through protein blocks. We will also establish what dictates the balance between accurate and inaccurate recombination mechanisms to understand when such processes might generate potentially very harmful changes to the genetic material.The conflict between the need to copy DNA and the need to have proteins bound to the DNA is one that all organisms must somehow resolve. This work will address therefore exactly what drives the accumulation of mutations within genes and what mechanisms help to minimise this accumulation. Acquisition of mutations in the genetic material is the driving force of evolution but such mutations are frequently harmful rather than beneficial and so must be kept in check. This is illustrated by the importance of mutations in the acquisition of human genetic disorders and the development of cancer. Understanding fundamental mechanisms of DNA replication and recombination in E. coli has greatly advanced our knowledge of genetic stability in more complex organisms such as ourselves. We are now in a position to use E. coli to address the interplay between these critical processes, an interplay that is central to understanding how genes are copied in as accurate a manner as possible.

细胞形成两个新子细胞的能力使生物可以生长和繁殖。此过程需要复制大量DNA，以便每个女儿都能获得生存所需的所有遗传信息的准确副本。由于生成DNA准确拷贝的重要性，生物已经进化出非常复杂的DNA复制机，以减少犯错的机会。不幸的是，我们现在知道这些复制机遇到了许多障碍，如果没有克服，这些障碍可能会在复制过程中造成错误。这样的错误可能会导致遗传信息的错误，这些信息可能会带来致命的后果。蛋白质覆盖DNA的蛋白质频繁地构成DNA复制机械的障碍物。这些蛋白质对于维持，读取和包装遗传信息至关重要，因此无法避免。尽管DNA复制机制可以成功地从DNA中取出大多数这些蛋白质，但与DNA结合的蛋白质数量的庞大数量意味着偶尔在其轨道中停止复制过程。但是，一类特定的酶可以修复并重新启动分解的DNA复制机。显然，这些重组酶可以帮助复制受蛋白质绑定的DNA，但它们的作用仍然不清楚。一个建议是，如果DNA复制机被蛋白质阻塞，那么重组酶可以在蛋白质块的另一侧重新启动DNA复制。该途径会跳过块，允许复制继续，但也会导致未复制蛋白质块附近的基因。因此，重组酶的参与可以看作是有害的，导致未能复制两个子细胞所需的所有遗传信息才能生存。另外，我们提出，重组酶可能只是重新启动DNA复制，靠近最初停止的位置。这可能会使DNA复制机械是第二次机会，将阻塞蛋白从DNA推出并继续复制所有遗传信息。因此，这样的过程可能会提供一种机制，以确保精确复制涂有蛋白质的DNA。我们将使用模型细菌大肠杆菌来确定重组酶在复制涂有蛋白质的DNA中的作用。我们对大肠杆菌中DNA复制和重组的基本机制了解很多，从而使我们能够分析这两个非常复杂的过程如何相互作用。我们将确定重组酶可以帮助DNA复制机通过蛋白质块移动的机制。我们还将确定什么决定了准确和不准确的重组机制之间的平衡，以了解何时可能会对遗传物质产生潜在的非常有害的变化。复制DNA的需求与将蛋白质绑定到DNA的需求之间的冲突是所有生物体必须以某种方式解决的所有生物体的冲突。因此，这项工作将准确地解决什么推动基因内突变积累的原因，以及哪些机制有助于最大程度地减少这种积累。遗传物质中突变的获取是进化的驱动力，但这种突变通常是有害的，而不是有益的，因此必须保留。突变在获得人类遗传疾病和癌症发展中的重要性来说明这一点。理解大肠杆菌中DNA复制和重组的基本机制已极大地提高了我们对更复杂的生物（例如我们自身）的遗传稳定性的了解。现在，我们可以使用大肠杆菌来解决这些关键过程之间的相互作用，这是了解如何以尽可能准确的方式复制基因的至关重要的相互作用。