From Comparative Genomics to Comparative Genetics - What is Required for Life Without DNA Replication Origins?

所有细胞都含有生物体DNA的完整副本，并将其打包成染色体。在细胞分裂之前，它们的染色体必须复制。这被称为DNA复制，开始于染色体上称为起始点的特定位置。细菌只有一个复制起点，但拥有大染色体的生物，如人类，需要多个复制起点。我们发现，在一种情况下，起源是不必要的，没有起源的细胞可以比正常情况下生长得更快。我们对DNA复制的研究是在古菌成员Haloferax Volcanii中进行的。生命之树分为三类：真核生物、细菌和古生菌。古生菌是以生活在极端条件下而闻名的微生物。Haloferax Volcanii来自死海，我们之所以选择它，是因为在古生物中进行DNA复制的酶与真核生物中的酶相似，比如人类细胞。Haloferax cancanii使用三个起点来复制它的染色体，但当所有起点都被删除时，细胞生长得更快。在人体上做这些实验是不可能的。当真核生物或细菌的起源被删除时，DNA复制就会被阻止，细胞就会死亡。那么，盐生植物火山是如何存活的呢？没有起源的细胞使用一种称为重组的过程来启动DNA复制。重组是DNA修复的一种形式，用于修复染色体上的断裂。我们发现，重组在染色体上的随机位置开始DNA复制，而不是在特定的起始处。但是，如果这种使用重组的DNA复制替代模式更有效，那么为什么会有起源呢？在一种名为Haloferax Mediterranei的姐妹古菌物种中，起源是无法消除的。尝试此操作时，休眠的复制源将变为活动状态。这意味着地中海盐生植物需要起源，而火山海洋盐生植物可以利用重组来启动DNA复制。为什么这两种密切相关的微生物的行为会如此不同？我们认为，火山盐生盐藻具有地中海盐生盐藻缺失的关键基因，反之亦然。为了简化对这些关键基因的搜索，我们将研究这些物种和多达20个额外的盐生植物的基因组。我们在罗马尼亚的同事已经发现特兰西瓦尼亚的盐湖是Haloferax物种的丰富来源。我们将测试这些Haloferax物种，看看它们的起源是否可以消除(如Haloferax Volcanus II)，或者它们是否是必需的(如Haloferax Mediterranei)。然后我们将比较它们的基因组以定位负责的基因。同时，我们将检查使用重组启动DNA复制的后果。Haloferax Volcanii可以使用重组启动DNA复制，但这可能是危险的。我们将测试它是否会导致突变或染色体重组，以及是否有避免重组的替代方案。与起源不同，重组可以发生在染色体的任何地方，但这种情况发生的频率取决于DNA的长度。我们将测试这种可供选择的DNA复制模式是否存在最小染色体大小。我们的工作将通过增加我们对癌症的理解来促进人类健康。我们在Haloferax Volcanii中发现的东西与癌细胞有相似之处。Haloferax的染色体有许多副本，这被称为多倍体，当复制和细胞分裂不协调时，它可以帮助它存活下来。癌细胞通常在控制DNA复制的基因上发生突变，多倍体是癌症的常见特征。不协调复制的另一个后果是癌细胞比普通细胞生长得更快。这类似于我们观察到的无来源的Haloferax Volcanii更快的生长，后者使用另一种DNA复制模式。

All cells contain a complete copy of the organism's DNA, packaged into chromosomes. Before cells can divide, their chromosomes must be duplicated. This is called DNA replication and begins at specific sites on the chromosome called origins. Bacteria have a single replication origin but organisms with large chromosomes, such as humans, need many origins. We have found that in one case, origins are unnecessary and that cells without them can grow faster than normal.Our research on DNA replication was carried out in Haloferax volcanii, a member of the archaea. The tree of life is split into three groups: eukaryotes, bacteria and archaea. Archaea are microbes renowned for living in extreme conditions. Haloferax volcanii comes from the Dead Sea, we chose it because the enzymes that carry out DNA replication in archaea are similar to those in eukaryotes, such as human cells.Haloferax volcanii uses three origins to replicate its chromosome but when all origins are deleted, the cells grow faster. Doing these experiments in humans would be impossible. When origins are deleted from eukaryotes or bacteria, DNA replication is prevented and cells die. So how is Haloferax volcanii able to survive?Cells without origins use a process called recombination to start DNA replication. Recombination is a form of DNA repair that is used to mend breaks in the chromosome. We found that recombination starts DNA replication at random locations on the chromosome, instead of at specific origins. But if this alternative mode of DNA replication using recombination is more efficient, why have origins at all?In a sister species of archaea called Haloferax mediterranei, origins cannot be eliminated. When this is attempted, a dormant replication origin becomes active. This means that Haloferax mediterranei needs origins, while Haloferax volcanii can instead use recombination to start DNA replication. Why do these two closely-related microbes behave so differently?We propose that Haloferax volcanii has critical genes that are missing from Haloferax mediterranei, or vice versa. To simplify the search for these critical genes, we will study the genomes of these and up to 20 additional Haloferax species. Our colleagues in Romania have already discovered that salt lakes in Transylvania are a rich source of Haloferax species. We will test these Haloferax species to see if their origins can be eliminated (as in Haloferax volcanii) or if they are essential (as in Haloferax mediterranei). Then we will compare their genomes to locate the genes responsible.At the same time, we will examine the consequences of using recombination to start DNA replication.Haloferax volcanii can use recombination to start DNA replication but this may be hazardous. We will test whether it leads to mutations or chromosome rearrangements, and whether there are alternatives that avoid recombination.Unlike origins, recombination can take place anywhere on the chromosome, but how often this happens depends on the length of DNA. We will test if there is a minimum size of chromosome for this alternative mode of DNA replication.Our work will contribute to human health by increasing our understanding of cancer. What we have discovered in Haloferax volcanii has parallels with cancer cells. Haloferax has many copies of its chromosome, this is called polyploidy and helps it to survive when replication and cell division are not coordinated. Cancer cells often have mutations in the genes that control DNA replication and polyploidy is a common feature of cancer. Another consequence of uncoordinated replication is that cancer cells grow faster than ordinary cells. This is similar to the faster growth we observe with origin-less Haloferax volcanii, which use an alternative mode of DNA replication.