Rare quadruple-helix DNA found in living human cells with glowing probes

发光探针在活人细胞中发现罕见的四重螺旋DNA

by Hayley Dunning, Imperial College London

伦敦帝国理工学院海莉·邓宁（Hayley Dunning）

Quadruple-helix DNA. Credit: Imperial College London

四重螺旋DNA。图片来源：伦敦帝国学院

New probes allow scientists to see four-stranded DNA interacting with molecules inside living human cells, unraveling its role in cellular processes.

新的探针使科学家能够看到四链DNA与活人细胞内的分子相互作用，从而揭示其在细胞过程中的作用。

DNA usually forms the classic double helix shape of two strands wound around each other. While DNA can form some more exotic shapes in test tubes, few are seen in real living cells.

DNA通常形成缠绕在一起的两条链的经典双螺旋形状。尽管DNA可以在试管中形成更多奇特的形状，但在实际的活细胞中却很少见到。

However, four-stranded DNA, known as G-quadruplex, has recently been seen forming naturally in human cells. Now, in new research published today in Nature Communications, a team led by Imperial College London scientists have created new probes that can see how G-quadruplexes are interacting with other molecules inside living cells.

但是，最近发现四链DNA（称为G-四链体）在人细胞中自然形成。现在，在《自然通讯》今天发表的一项新研究中，由伦敦帝国理工学院的科学家领导的一个团队创造了新的探针，可以观察到G四联体如何与活细胞内的其他分子相互作用。

G-quadruplexes are found in higher concentrations in cancer cells, so are thought to play a role in the disease. The probes reveal how G-quadruplexes are 'unwound' by certain proteins, and can also help identify molecules that bind to G-quadruplexes, leading to potential new drug targets that can disrupt their activity.

G-四链体在癌细胞中的浓度较高，因此被认为在该疾病中起作用。这些探针揭示了某些蛋白质如何解开G-四链体，还可以帮助鉴定与G-四链体结合的分子，从而导致可能破坏其活性的新药物靶标。

Needle in a haystack

大海捞针

One of the lead authors, Ben Lewis, from the Department of Chemistry at Imperial, said: "A different DNA shape will have an enormous impact on all processes involving it—such as reading, copying, or expressing genetic information.

帝国理工学院化学系的主要作者之一本·刘易斯说：“不同的DNA形状会对涉及它的所有过程产生巨大影响，例如读取，复制或表达遗传信息。

"Evidence has been mounting that G-quadruplexes play an important role in a wide variety of processes vital for life, and in a range of diseases, but the missing link has been imaging this structure directly in living cells."

“越来越多的证据表明，G-四链体在对生命至关重要的各种过程以及各种疾病中都起着重要作用，但缺失的环节一直是直接在活细胞中对这种结构进行成像。”

G-quadruplexes are rare inside cells, meaning standard techniques for detecting such molecules have difficulty detecting them specifically. Ben Lewis describes the problem as "like finding a needle in a haystack, but the needle is also made of hay."

G-四链体在细胞内很少见，这意味着用于检测此类分子的标准技术很难对其进行特异性检测。本·刘易斯（Ben Lewis）将问题描述为“就像在大海捞针中找到针头，但针头也是由干草制成”。

To solve the problem, researchers from the Vilar and Kuimova groups in the Department of Chemistry at Imperial teamed up with the Vannier group from the Medical Research Council's London Institute of Medical Sciences.

为了解决该问题，帝国理工大学化学系的Vilar和Kuimova小组的研究人员与医学研究理事会伦敦医学科学研究所的Vannier小组合作。

Fluorescence lifetime imaging microscopy map of nuclear DNA in live cells stained with the new probe. Colours represent fluorescence lifetimes between 9 (red) and 13 (blue) nanoseconds.Credit: Imperial College London

用新探针染色的活细胞中核DNA的荧光寿命成像显微图。颜色表示荧光寿命在9（红色）和13（蓝色）纳秒之间。图片来源：伦敦帝国学院

They used a chemical probe called DAOTA-M2, which fluoresces (lights up) in the presence of G- quadruplexes, but instead of monitoring the brightness of fluorescence, they monitored how long this fluorescence lasts. This signal does not depend on the concentration of the probe or of G-quadruplexes, meaning it can be used to unequivocally visualize these rare molecules.

他们使用了一种名为DAOTA-M2的化学探针，该探针在存在G-四链体的情况下发出荧光（亮起），但他们没有监测荧光的亮度，而是监测了这种荧光的持续时间。该信号不依赖于探针或G-四链体的浓度，这意味着它可以用于清晰地观察这些稀有分子。

Dr. Marina Kuimova, from the Department of Chemistry at Imperial, said: "By applying this more sophisticated approach we can remove the difficulties which have prevented the development of reliable probes for this DNA structure."

帝国理工大学化学系的Marina Kuimova博士说：“通过应用这种更复杂的方法，我们可以消除阻碍这种DNA结构的可靠探针研发的困难。”

Looking directly in live cells

直接在活细胞中看

The team used their probes to study the interaction of G-quadruplexes with two helicase proteins—molecules that 'unwind' DNA structures. They showed that if these helicase proteins were removed, more G-quadruplexes were present, showing that the helicases play a role in unwinding and thus breaking down G-quadruplexes.

该团队使用他们的探针来研究G-四链体与两种解旋酶蛋白（“解开” DNA结构的分子）之间的相互作用。他们表明，如果去除了这些解旋酶蛋白，则会存在更多的G-四链体，表明解旋酶在解链并因此破坏G-四链体中起作用。

Dr. Jean-Baptiste Vannier, from the MRC London Institute of Medical Sciences and the Institute of Clinical Sciences at Imperial, said: "In the past we have had to rely on looking at indirect signs of the effect of these helicases, but now we take a look at them directly inside live cells."

来自MRC伦敦医学科学研究所和帝国理工学院临床科学研究所的Jean-Baptiste Vannier博士说：“过去，我们不得不依靠观察这些解旋酶作用的间接迹象，但现在我们直接在活细胞内看一下它们。”

They also examined the ability of other molecules to interact with G-quadruplexes in living cells. If a molecule introduced to a cell binds to this DNA structure, it will displace the DAOTA-M2 probe and reduce its lifetime, i.e. how long the fluorescence lasts.

他们还研究了其他分子与活细胞中G-四链体相互作用的能力。如果引入细胞的分子与该DNA结构结合，它将取代DAOTA-M2探针并缩短其寿命，即荧光持续多长时间。

This allows interactions to be studied inside the nucleus of living cells, and for more molecules, such as those which are not fluorescent and can't be seen under the microscope, to be better understood.

这样可以更好地理解在活细胞核内进行的相互作用，以及更多分子（例如不发荧光且无法在显微镜下看到的分子）的相互作用。

Professor Ramon Vilar, from the Department of Chemistry at Imperial, explained: "Many researchers have been interested in the potential of G-quadruplex binding molecules as potential drugs for diseases such as cancers. Our method will help to progress our understanding of these potential new drugs."

帝国理工大学化学系的拉蒙·维拉尔（Ramon Vilar）教授解释说：“许多研究人员对G四联体结合分子作为潜在的药物治疗癌症等疾病的潜力感兴趣。我们的方法将有助于增进我们对这些潜在新疾病的认识。毒品。”

Peter Summers, another lead author from the Department of Chemistry at Imperial, said: "This project has been a fantastic opportunity to work at the intersection of chemistry, biology and physics. It would not have been possible without the expertise and close working relationship of all three research groups."

帝国理工学院化学系的另一位主要作者彼得·萨默斯（Peter Summers）表示：“这个项目是在化学，生物学和物理学的交汇处工作的绝好机会。没有他的专业知识和紧密的工作关系是不可能的。三个研究小组。”

The three groups intend to continue working together to improve the properties of

这三个小组打算继续共同努力，以改善

their probe and to explore new biological problems and shine further light on the roles G- quadruplexes play inside our living cells. The research was funded by Imperial's Excellence Fund for Frontier Research.

他们的探针，并探索新的生物学问题，并进一步阐明G-四链体在我们的活细胞中的作用。这项研究由帝国帝国卓越研究基金资助。

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