Rapid DNA replication origin licensing protects stem cell pluripotency.
Rapid DNA replication origin licensing protects stem cell pluripotency.
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DOI：
10.7554/elife.30473
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发表时间：
2017-11-17
期刊：
影响因子：
7.7
通讯作者：
Cook JG
中科院分区：
文献类型：
--
作者：
Matson JP;Dumitru R;Coryell P;Baxley RM;Chen W;Twaroski K;Webber BR;Tolar J;Bielinsky AK;Purvis JE;Cook JG
完整而稳健的人类基因组复制需要在许多 DNA 复制起点加载微型染色体维持 (MCM) 解旋酶复合物，这是一个称为起点许可的重要过程。许可仅限于细胞周期的 G1 期，但不同细胞类型的 G1 长度差异很大。通过定量单细胞分析，我们发现具有天然短 G1 期的多能干细胞加载 MCM 的速度比具有长 G1 期的同基因分化对应物快得多。在向所有谱系分化的最早阶段，MCM 负荷与 G1 延长同时减慢，揭示了 MCM 负荷的发育控制。相比之下，异位 Cyclin E 过量产生使短 G1 与快速 MCM 加载脱钩。干细胞的快速许可是由 MCM 负载蛋白 Cdt1 的积累引起的。过早减慢多能细胞中的 MCM 加载不仅会延长 G1 期，还会加速分化。因此，快速起源许可是干细胞的固有特征，有助于维持多能性。从红细胞到神经细胞，动物体内含有许多不同类型的特化细胞。这些细胞都以干细胞开始，具有分裂和产生更多干细胞或特化的潜力。所有分裂细胞必须首先解开它们的DNA，以便可以复制。为了实现这一目标，细胞在称为 G1 期的细胞周期部分将称为解旋酶的 DNA 解旋酶加载到其 DNA 上。细胞必须装载足够的解旋酶，以确保其 DNA 被完全、及时地复制。干细胞比它们特化的后代分裂得更快，并且 G1 期也短得多。然而这些细胞仍然设法加载足够的解旋酶来复制它们的 DNA。对于以不同速度分裂的细胞之间解旋酶加载的量、速率和时间如何变化知之甚少。现在马森等人。测量了解旋酶加载到单个人类细胞（包括干细胞和特化或“分化”细胞）DNA 上的速度。干细胞快速加载解旋酶，以弥补它们在 G1 期花费的短暂时间，而分化细胞加载酶的速度较慢。测量干细胞被触发特化时加载速率的变化表明，随着 G1 期变长，解旋酶加载速度减慢。马特森等人。发现解旋酶加载所需的关键蛋白质水平与加载速率相关。改变蛋白质的水平会改变酶加载的速度以及细胞的行为方式——例如，减慢解旋酶的加载使干细胞更快地分化。这些发现表明细胞分化和DNA复制过程密切相关。这项研究和未来的研究将帮助科学家了解早期动物发育过程中发生的情况，即当专业化首次发生时，以及癌细胞中出现了什么问题，癌细胞也分裂得很快。更好地了解这一过程也有助于再生医学——其中的挑战之一是制造足够的特化细胞来移植到组织损伤的患者体内，而这些细胞又不会癌变。
Complete and robust human genome duplication requires loading minichromosome maintenance (MCM) helicase complexes at many DNA replication origins, an essential process termed origin licensing. Licensing is restricted to G1 phase of the cell cycle, but G1 length varies widely among cell types. Using quantitative single-cell analyses, we found that pluripotent stem cells with naturally short G1 phases load MCM much faster than their isogenic differentiated counterparts with long G1 phases. During the earliest stages of differentiation toward all lineages, MCM loading slows concurrently with G1 lengthening, revealing developmental control of MCM loading. In contrast, ectopic Cyclin E overproduction uncouples short G1 from fast MCM loading. Rapid licensing in stem cells is caused by accumulation of the MCM loading protein, Cdt1. Prematurely slowing MCM loading in pluripotent cells not only lengthens G1 but also accelerates differentiation. Thus, rapid origin licensing is an intrinsic characteristic of stem cells that contributes to pluripotency maintenance. From red blood cells to nerve cells, animals’ bodies contain many different types of specialized cells. These all begin as stem cells, which have the potential to divide and make more stem cells or to specialize. All dividing cells must first unwind their DNA so that it can be copied. To achieve this, cells load DNA-unwinding enzymes called helicases onto their DNA during the part of the cell cycle known as G1 phase. Cells must load enough helicase enzymes to ensure that their DNA is copied completely and in time. Stem cells divide faster than their specialized descendants, and have a much shorter G1 phase too. Yet these cells still manage to load enough helicases to copy their DNA. Little is known about how the amount, rate and timing of helicase loading varies between cells that divide at different speeds. Now Matson et al. have measured how quickly helicase enzymes are loaded onto DNA in individual human cells, including stem cells and specialized or “differentiated” cells. Stem cells loaded helicases rapidly to make up for the short time they spent in G1 phase, while differentiated cells loaded the enzymes more slowly. Measuring how the loading rate changed when stem cells were triggered to specialize showed that helicase loading slowed as the G1 phase got longer. Matson et al. found that the levels of key proteins required for helicase loading correlated with the rates of loading. Altering the levels of the proteins changed how quickly the enzymes were loaded and how the cells behaved – for example, slowing down the loading of helicases made the stem cells specialize quicker. These findings show that the processes of cell differentiation and DNA replication are closely linked. This study and future ones will help scientists understand what is happening during early animal development, when specialization first takes place, as well as what has gone wrong in cancer cells, which also divide quickly. A better understanding of this process also helps in regenerative medicine – where one of the challenges is to make enough specialized cells to transplant into a patient with tissue damage without those cells becoming cancerous.