更新于：2024-11-01

XRN1

更新于：2024-11-01

基本信息

别名

5'-3' exoribonuclease 1、SEP1、Strand-exchange protein 1 homolog

+ [1]

简介

Major 5'-3' exoribonuclease involved in mRNA decay. Required for the 5'-3'-processing of the G4 tetraplex-containing DNA and RNA substrates. The kinetic of hydrolysis is faster for G4 RNA tetraplex than for G4 DNA tetraplex and monomeric RNA tetraplex. Binds to RNA and DNA (By similarity). Plays a role in replication-dependent histone mRNA degradation. May act as a tumor suppressor protein in osteogenic sarcoma (OGS).

关联

项与 XRN1 相关的药物

XRN1(Accent)

靶点

XRN1

作用机制

XRN1 inhibitors

在研机构

Accent Therapeutics, Inc.初创企业

原研机构

Accent Therapeutics, Inc.初创企业

在研适应症

非小细胞肺癌 [+1]

非在研适应症-

最高研发阶段临床前

首次获批国家/地区-

首次获批日期1800-01-20

100 项与 XRN1 相关的临床结果

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100 项与 XRN1 相关的转化医学

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0 项与 XRN1 相关的专利（医药）

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544

项与 XRN1 相关的文献（医药）

2024-11-01·Molecular Biology of the Cell

Med13 is required for efficient P-body recruitment and autophagic degradation of Edc3 following nitrogen starvation

Article

作者: Hanley, Sara E. ; Friedson, Brittany ; Cooper, Katrina F. ; Willis, Stephen D.

The Cdk8 kinase module (CKM), a conserved, detachable unit of the Mediator complex, plays a vital role in regulating transcription and communicating stress signals from the nucleus to other organelles. Here, we describe a new transcription-independent role for Med13, a CKM scaffold protein, following nitrogen starvation. In Saccharomyces cerevisiae, nitrogen starvation triggers Med13 to translocate to the cytoplasm. This stress also induces the assembly of conserved membraneless condensates called processing bodies (P-bodies) that dynamically sequester translationally inactive messenger ribonucleoprotein particles. Cytosolic Med13 colocalizes with P-bodies, where it helps recruit Edc3, a highly conserved decapping activator and P-body assembly factor, into these conserved ribonucleoprotein granules. Moreover, Med13 orchestrates the autophagic degradation of Edc3 through a selective cargo-hitchhiking autophagy pathway that utilizes Ksp1 as its autophagic receptor protein. In contrast, the autophagic degradation of Xrn1, another conserved P-body assembly factor, is Med13 independent. These results place Med13 as a new player in P-body assembly and regulation following nitrogen starvation. They support a model in which Med13 acts as a conduit between P-bodies and phagophores, two condensates that use liquid-liquid phase separation in their assembly.

2024-10-10·The Journal of Physical Chemistry B

Magnesium Regulates RNA Ring Dynamics and Folding in Subgenomic Flaviviral RNA

Article

作者: Singh, Rishabh K. ; Roy, Susmita ; Kundu, Rimi ; Mainan, Avijit

Mosquito-borne flaviviruses including dengue, Zika, yellow fever, and regional encephalitis produce a large amount of short subgenomic flaviviral RNAs during infection. A segment of these RNAs named as xrRNA1 features a multi-pseudoknot (PK)-associated structure, which resists the host cell enzyme (XRN1) from degrading the viral RNA. We investigate how this long-range RNA PK folds in the presence of counterions, specifically in a mix of monovalent (K⁺) and divalent (Mg²⁺) salts at physiological concentrations. In this study, we use extensive explicit solvent molecular dynamics (MD) simulations to characterize the RNA ion environment of the folded RNA conformation, as determined by the crystal structure. This allowed us to identify the precise locations of various coordinated RNA-Mg²⁺ interactions, including inner-sphere/chelated and outer-sphere coordinated Mg²⁺. Given that RNA folding involves large-scale conformational changes, making it challenging to explore through classical MD simulations, we investigate the folding mechanism of xrRNA1 using an all-atom structure-based RNA model with a hybrid implicit-explicit treatment of the ion environment via the dynamic counterion condensation model, both with and without physiological Mg²⁺ concentration. The study reveals potential folding pathways for this xrRNA1, which is consistent with the results obtained from optical tweezer experiments. The equilibrium and free energy simulations both capture a dynamic equilibrium between the ring-open and ring-close states of the RNA, driven by a long-range PK interaction. Free energy calculations reveal that with the addition of Mg²⁺ ions, the equilibrium shifts more toward the ring-close state. A detailed analysis of the free energy pathways and ion-mediated contact probability map highlights the critical role of Mg²⁺ in bridging G50 and A33. This Mg²⁺-mediated connection helps form the long-range PK which in turn controls the transition between the ring-open and ring-close states. The study underscores the critical role of Mg²⁺ in the RNA folding transition, highlighting specific locations of Mg²⁺ contributing to the stabilization of long-range PK connections likely to enhance the robustness of Xrn1 resistance of flaviviral xrRNAs.

2024-10-01·Yeast

The 5‐Fluorouracil RNA Expression Viewer (5‐FU^R) Facilitates Interpreting the Effects of Drug Treatment and RRP6 Deletion on the Transcriptional Landscape in Yeast

Article

作者: Bretaudeau, Anthony ; Primig, Michael ; Sallou, Oliver ; Morillon, Antonin ; Szachnowski, Ugo ; Wery, Maxime ; Boudet, Mateo

ABSTRACTSaccharomyces cerevisiae is an excellent model to study the effect of external cues on cell division and stress response. 5‐Fluorocuracil (5‐FU) has been used to treat solid tumors since several decades. The drug was initially designed to interfere with DNA replication but was later found to exert its antiproliferative effect also via RNA‐dependent processes. Since 5‐FU inhibits the activity of the 3′−5′‐exoribonuclease Rrp6 in yeast and mammals, earlier work has compared the effect of 5‐FU treatment and RRP6 deletion at the transcriptome level in diploid synchronized yeast cells. To facilitate interpreting the expression data we have developed an improved 5‐Fluorouracil RNA (5‐FUR) expression viewer. Users can access information via genome coordinates and systematic or standard names for mRNAs and Xrn1‐dependent‐, stable‐, cryptic‐, and meiotic unannotated transcripts (XUTs, SUTs, CUTs, and MUTs). Normalized log2‐transformed or linear data can be displayed as filled diagrams, line graphs or color‐coded heatmaps. The expression data are useful for researchers interested in processes such as cell cycle regulation, mitotic repression of meiotic genes, the effect of 5‐FU treatment and Rrp6 deficiency on the transcriptome and expression profiles of sense/antisense loci that encode overlapping transcripts. The viewer is accessible at http://5fur.genouest.org.

项与 XRN1 相关的新闻（医药）

2023-10-10

·PRNewswire

Accent Therapeutics Unveils XRN1 Program and Provides Pipeline Update with Presentations at 35th AACR-NCI-EORTC Symposium

Company developing potent and selective small molecule inhibitors of XRN1, a compelling oncology target to address tumors with high interferon signaling Presentations at Triple Meeting also demonstrate progress for novel DHX9 and potentially first-in-class ADAR1 programs LEXINGTON, Mass., Oct. 10, 2023 /PRNewswire/ -- Accent Therapeutics, a biopharmaceutical company pioneering a new class of small molecule precision cancer therapies, today announced it will present data unveiling a novel, previously undisclosed program in two poster presentations at the 35th AACR-NCI-EORTC Symposium held October 11-15, 2023 in Boston, Massachusetts. The company will also present data supporting development of its ADAR1 and DHX9 programs in two additional poster presentations. 5′→3′ exoribonuclease 1 (XRN1) is an enzyme that degrades single- and double-stranded RNA, playing roles in extracellular mRNA turnover and innate immunity through prevention of dsRNA accumulation and subsequent MDA5 and PKR-mediated immune signaling. Accent has identified XRN1 as a key vulnerability for therapeutic targeting of tumors with elevated type I interferon signaling and is developing small molecule inhibitors of this compelling oncology target, for which there are none currently disclosed. Two of the company's presentations at the meeting describe the preclinical validation of XRN1 as an oncology target and the subsequent characterization of selective inhibitors of XRN1. XRN1 knockout resulted in cell death due to accumulation of dsRNA and downstream activation of the MDA5 and PKR innate immune pathways in cells with intrinsically elevated type I interferon signaling, but not in those with low interferon signaling. Accent has identified novel, potent, and selective allosteric inhibitors that bind XRN1 with nanomolar target affinity without binding to XRN2. "We are excited to share our progress advancing our portfolio and to introduce our XRN1 inhibitor program as a potentially impactful approach to helping cancer patients," said Robert Copeland, Ph.D., President and Chief Scientific Officer of Accent Therapeutics. "These data demonstrate the versatility and promise of pursuing critical intracellular dependences of cancer biology to identify novel oncology targets with the potential to benefit broad patient populations." ADAR1 has emerged as a promising, yet challenging-to-drug, oncology target for which inhibition has been shown to induce downstream immune activation and subsequent cell death in cancer cells with high intrinsic type 1 interferon signaling—a trait that 15-30% of primary tumors share. Accent will also present its innovative suite of in vitro and cellular assays that enable identification of small molecule inhibitors of ADAR1. The resulting compounds are shown to potently inhibit cell growth in a cancer cell line with high interferon type 1 signaling. The company's final presentation at the meeting will highlight its progress developing small molecule inhibitors of the novel oncology target DHX9, an RNA helicase that has been reported to play important roles in the maintenance of genomic stability in multiple cancer types. The presentation will demonstrate that DHX9 inhibition in microsatellite instable tumors exhibiting defective mismatch repair induces replication stress and subsequent cell death. The poster presentations will be archived on the Accent website following the meeting. Details for the presentations are as follows: Title: Exoribonuclease XRN1 is a Therapeutic Vulnerability in Tumors with Intrinsically Elevated Type I Interferon Signaling Abstract Number: A163 Session & Location: Poster Session A – Exhibit Hall D Session Date & Time: Thursday, October 12, 12:30 pm – 4:00 pm ET Presenter: Maureen M. Lynes, Ph.D. Title: Discovery of Small Molecule Inhibitors of ADAR1 Abstract Number: A170 Session & Location: Poster Session A – Exhibit Hall D Session Date & Time: Thursday, October 12, 12:30 pm – 4:00 pm ET Presenter: Shane M. Buker, Ph.D. Title: Characterization of Selective, Allosteric Inhibitors of Human XRN1 Abstract Number: B073 Session & Location: Poster Session B – Exhibit Hall D Session Date & Time: Friday, October 13, 12:30 pm – 4:00 pm ET Presenter: Gordon J. Lockbaum, Ph.D. Title: DHX9 Inhibition as a Novel Therapeutic Modality in Microsatellite Instable Colorectal Cancer Abstract Number: C087 Session & Location: Poster Session C – Exhibit Hall D Session Date & Time: Saturday, October 14, 12:30 pm – 4:00 pm ET Presenter: Jennifer Castro About XRN1 XRN1 is an RNA exonuclease that plays important roles in innate immunity by preventing double-stranded RNA (dsRNA) accumulation. Tumors with intrinsic type I interferon signaling are acutely sensitive to XRN1 loss, making it a strong oncology target. Inhibitors of XRN1 have potential for use as monotherapy and in combination with immuno-oncology therapeutics. Accent is applying structure-based design approaches to develop potent and selective small molecule inhibitors of XRN1 to address cancer indications with high unmet medical need. About ADAR1 Adenosine deaminase acting on RNA 1 (ADAR1) is an enzyme that edits double-stranded RNA (dsRNA) to prevent recognition and downstream immune signaling. In tumor cells with high interferon signaling, small molecule inhibitors of ADAR1 have been shown to induce selective cell killing and immune sensitization, making it a highly attractive target for monotherapy and combination approaches with immunotherapies, such as checkpoint inhibitors. Accent is developing ADAR1 targeted therapies to address many solid tumor indications with significant unmet need, including non-small cell lung cancer, triple-negative breast cancer, ovarian cancer, and head and neck squamous cell carcinomas. About DHX9 Accent's lead program is a first-in-class DHX9 inhibitor with the potential to address high unmet need indications that lack targeted therapies; these include colorectal, endometrial, gastric, and other high microsatellite instable (MSI-H) cancers. Several additional undisclosed cancer types representing large patient populations are also being explored based on their sensitivities to DHX9 inhibition. DHX9 is a DNA/RNA helicase that has been reported to play important roles in replication, transcription, translation, RNA splicing, RNA processing, and maintenance of genomic stability. Hence, this enzyme represents a compelling novel oncology target as inhibition of DHX9 exploits key tumor vulnerabilities, resulting in cancer-specific death. Accent is currently conducting IND-enabling studies evaluating its DHX9 inhibitor. About Accent Therapeutics Accent Therapeutics is pioneering a new class of small molecule precision cancer therapies targeting critical intracellular dependencies that span multiple types of cancer. Building upon industry-leading expertise in RNA-modifying proteins (RMPs) and the systematic mapping of both the RMP space and adjacent high-value areas for drug discovery, the company is building a flexible model that allows for a diversity of approaches to developing potentially transformative biomarker-driven cancer medicines. Accent's therapies are designed for both novel and known, but suboptimally-addressed, high-impact oncogenic targets with the potential to benefit large patient populations with significant unmet need. For more information on Accent's mission to translate extraordinary science into life-changing therapeutics for patients living with cancer, visit or follow us on LinkedIn. SOURCE Accent Therapeutics, Inc.

AACR会议免疫疗法

2023-02-02

·生物谷

Cancer Res：靶向RNA外切酶XRN1增强肿瘤免疫治疗的疗效

免疫检查点阻断(ICB)抗体，如抗PD-1和抗CTLA4，已经改变了许多类型癌症的治疗格局。尽管取得了这一显著成就，但ICB疗法的应答率相对较低，只有一小部分患者能从治疗中受益。免疫检查点阻断(ICB)抗体，如抗PD-1和抗CTLA4，已经改变了许多类型癌症的治疗格局。尽管取得了这一显著成就，但ICB疗法的应答率相对较低，只有一小部分患者能从治疗中受益。最近的研究比较了PD-1反应性黑色素瘤和无反应性黑色素瘤样本的突变特征，确认了干扰素信号在控制ICB反应中的关键作用。在用PD-1抗体处理的免疫活性小鼠中进行的体内CRISPR文库筛选也得出了类似的结论。这些发现表明，激活肿瘤细胞固有的干扰素信号可以加强免疫检查点阻断治疗，为开发干预方法扩大ICB的应用提供了线索。图片来源：https://pubmed-ncbi-nlm-nih-gov.libproxy1.nus.edu.sg/36638333/ 近日，来自新加坡国立大学的研究者们在Cancer research杂志上发表了题为“Targeting RNA exonuclease XRN1 potentiates efficacy of cancer immunotherapy”的文章，该研究发现为开发XRN1抑制剂并探索其与癌症免疫治疗相结合的潜在临床应用提供了分子基础。尽管免疫检查点阻断(ICB)治疗取得了显著的临床疗效，但应答率相对较低，只有一小部分患者能从治疗中受益。异常的RNA积聚可以介导干扰素信号并刺激免疫反应，提示靶向RNA降解机制可能使肿瘤细胞对免疫治疗敏感。考虑到这一点，研究者确定了一种RNA外切核酸酶XRN1，作为抑制RNA衰退和刺激抗肿瘤免疫的潜在治疗靶点。沉默XRN1抑制了同基因免疫活性小鼠的肿瘤生长，并增强了免疫治疗效果，而沉默XRN1本身并不影响免疫缺陷小鼠的肿瘤生长。从机制上讲，XRN1缺失激活了干扰素信号和病毒防御途径；这两条途径在调节免疫逃避方面都起着决定性的作用。异常RNA传感信号蛋白(RIG-I/MAV)介导了干扰素基因的表达，因为它们中的每一个缺失都会钝化XRN1沉默细胞中抗病毒/干扰素信号的上调。对泛癌CRISPR筛查数据的分析表明，XRN1沉默触发的干扰素信号是一种普遍现象，提示XRN1沉默的作用可能扩展到多种类型的癌症。总体而言，XRN1缺失触发了异常的RNA介导的干扰素信号，突显了异常RNA传感通路在调节免疫反应中的重要性。 XRN1缺失增强肿瘤组织中免疫细胞的侵袭图片来源：https://pubmed-ncbi-nlm-nih-gov.libproxy1.nus.edu.sg/36638333/ 综上所述，研究者发现XRN1的缺失触发了一个强烈的异常--RNA介导的干扰素信号，并显著增强了免疫治疗的效果。本研究强调了异常RNA传感通路在肿瘤细胞内源性免疫反应中的重要性，为开发XRN1抑制剂并探索其在肿瘤免疫治疗中的潜在临床应用提供了分子基础。（生物谷 Bioon.com）参考文献 Xue-Bin Ran et al. Targeting RNA exonuclease XRN1 potentiates efficacy of cancer immunotherapy. Cancer Res. 2023 Jan 13;CAN-21-3052. doi: 10.1158/0008-5472.CAN-21-3052.

免疫疗法基因疗法

2022-07-15

·生物制品圈

推荐阅读|mRNA 5'端质量控制

摘要：所有真核生物的mRNA都在5'端被加帽。mRNA加帽与转录同步进行，该过程分为三步。加帽过程的中间体，和未加帽的5’-三磷酸RNA一样，对去帽和某些已知因素导致的降解都具有抵抗性，导致人们认为mRNA5’端最终都会完成加帽过程。这种观点最近才发生了重大的变化。一个新的酶家族，包括酵母蛋白Rai1, Dxo1/Ydr370C和哺乳动物蛋白DXO/Dom3Z已经得到确认和鉴定。这些酶可以将加帽异常的mRNA转化为5'-单磷酸RNA，后者进而被5'-3'外切核糖核酸酶降解。其中一些酶本身也具有5'-3'外切核糖核酸酶活性，可以直接清除未正确加帽的mRNA。对这些酶的研究，使人们认识到不是所有mRNA加帽过程都会完成，并确定了真核生物mRNA加帽过程的质量控制机制。本文就这一领域的最新进展作一简要综述。 1.引言所有真核生物的mRNA的5’端都会被加帽，这一过程发生在转录起始后不久。帽子结构包含N7位点的甲基化鸟苷酸，并通过一个特殊的5’-5’焦磷酸键，与RNA的5’端核苷酸连接（m7GpppN)。加帽对mRNA的稳定性、剪接、聚腺苷酸化、出核，以及翻译效率至关重要。帽子结构的去除是一个高度调控的过程，由脱帽酶Dcp2和Nudt16催化完成。这些酶将7-甲基-二磷酸鸟苷从mRNA上酶切下来，生成5’-单磷酸mRNA。接下来，该5’-单磷酸结构会被5’-3’外切核糖核酸酶Xrn1特异性识别和酶切，引发RNA降解并脱帽。加帽过程分三步进行。第一步，起始转录产物的5’端三磷酸被转化成二磷酸键；接下来一个单磷酸鸟苷分子连接到该二磷酸末端；最后，鸟嘌呤碱基N7位点发生甲基化，从而形成成熟的帽子结构。在酵母中，加帽过程由单独的三种酶催化完成，而在哺乳动物中，第一、二步反应是由一个双功能的酶来催化的。加帽过程中间体以及未加帽的初始转录产物，可抵抗由Xrn1酶引起的降解，因为该酶特异性识别5’单磷酸位点。而脱帽酶 Dcp2/Nudt16识别的是成熟的、甲基化的帽子结构，所以并不能将这些加帽中间体或未加帽的初始转录产物转化成可降解的单磷酸RNA。鉴于这些原因，人们曾认为mRNA总是会被正常加帽，因为非正常加帽的mRNA分子并不能被一些已知的因素降解，从而被注意到。近年来，该观点发生了翻天覆地的转变。酵母中Rai1蛋白的酶功能得到确认，它可以将初始转录产物或者未甲基化的帽子结构（加帽中间体）5’端的三磷酸基团，转化成单磷酸基团，进而被降解。这些发现引发了mRNA加帽过程的质量控制机制。因此，我们将简要回顾这一领域的最新进展。2.mRNA加帽过程质量控制机制的发现酵母Rai1蛋白（Rat1相互作用蛋白1），已经被认为可以结合并激活5’-3’外切核糖核酸酶Rat1/Xrn2，Rat1/Xrn2在聚合酶II（PolII）转录终止中起重要作用。Rai1的首个三维结构是以Rat1/Xrn2复合体的形式被测定的。但是，其结构表明Rai1具有某种酶活活性。在表面观察到一个大口袋，远离Rat1/Xrn2结合区域（图1a）。这个大口袋区域包含诸多保守位点，提示其可能作为活性位点存在。随后的研究一致表明Rai1具有焦磷酸酶活性，且都归因于这个大口袋活性位点。该活性将5’-三磷酸RNA转化为5’-单磷酸RNA。和Xrn1一样，Rai1相关的外切核糖核酸酶Rat1/Xrn2特异性地识别5’-单磷酸的RNA底物，而不能识别和酶切5’-三磷酸RNA。Rai1酶活的确认提示这类RNA可被Rai1-Rat1/Xrn2复合物降解。在这一线索的基础上，后续的研究表明，Rai1还可以催化加帽中间体-未甲基化帽子结构的降解。在帽子结构甲基化缺陷菌株里，敲除Rai1基因显著增加了mRNA的半衰期，说明细胞里的Rai1蛋白在非正常加帽mRNA的清除过程中发挥了关键的作用。重要的是，更多的研究表明mRNA加帽是一个高度调控的过程，并不总能正确完成，这与之前的设想完全相反。在寡营养条件下，虽然Rai1基因敲除菌株的加帽机制是完整的，但非正常mRNA的相对比重依然显著增加。综上的这些证据揭示了mRNA加帽的质量控制机制。图1 Rai1及其同系物催化反应的结构基础(a)Rai1的结构(由PyMOL生成，网址http://www.pymol.org)。红色圈出的为活性位点，箭头所示为Rai1与Rat1/Xrn2相互作用的区域。(b) DXO/Dom3Z与帽子结构模拟物m7GpppG形成的复合物的结构。(c) DXO/Dom3Z与寡核苷酸复合物的结构。(b)和(c)的结构大致有一定相关性，即沿垂直轴旋转了45。(d) DXO/Dom3Z的活性位点。图中展示了帽子结构和第二个核苷酸的位置，粉色框里表示的是第一个核苷酸的位置。红色字母“W”表示进攻水或羟基，红色箭头表示亲核攻击的方向。图(b) - (d)引用自Jiao et al.(2013)。(注:此图例中对颜色的解释，请读者参考本文网络版) 3.酵母和哺乳动物中mRNA加帽过程的质量控制Rai1的同源蛋白存在于大多数真核生物中。尽管这些同源蛋白的序列相似度并不是很高，但Rai1活性位点相关的残基序列高度保守。这意味着Rai1同源蛋白也具有相似的酶活性，真核生物中的mRNA加帽控制机制是一致的。迄今为止，已被研究的Rai1同源蛋白有2个，这使得研究者对酵母中mRNA加帽质量控制机制有了更深入的认识，并推动了哺乳动物中这一机制的确认和表征。大多数的真核生物中仅有一种Rai1同源蛋白，但在某些真菌（如酿酒酵母）中，还存在其它种类的Rai1同源蛋白。该蛋白（Ydr370C）与Rai1序列相似度为20%。与Rai1类似，Ydr370C对含未甲基化帽子结构的RNA也具有脱帽活性。此外，Ydr370C还具有5’-3’核酸外切酶活性。正因为Ydr370C可以独立降解未甲基化mRNA，因此被称作脱帽外切核糖核酸酶1，即Dxo1。和Rai1不同，未检测到Dxo1/Ydr370C具有任何焦磷酸化酶活性。在正常生长条件下，敲除Rai1或Dxo1/Ydr370C都没有检测到mRNA水平的变化。但是，同时敲除这两种基因则会观察到未正确加帽mRNA比例的显著增加，表明这两种蛋白在酵母mRNA加帽质量控制过程中都发挥着必要的作用，且某种程度上来说二者是互补的。哺乳动物中Rai1的同源蛋白是Dom3Z。研究发现，它是Rai1和Dxo1/Ydr370C的杂合形式，同时拥有对5’-三磷酸或未甲基化帽子RNA的焦磷酸化酶和脱帽酶活性，且具有5’-3’外切核糖核酸酶活性，因此后续更名为DXO。这些活性使得DXO/Dom3Z可以独立降解未加帽的、或者未甲基化加帽的RNA。敲除细胞中的DXO/Dom3Z，对成熟加帽的mRNA水平没有影响，但观察到未正确加帽mRNA的显著增加。这些都表明在哺乳动物中存在mRNA加帽控制机制，且DXO/Dom3Z在其中发挥了关键作用。细胞中，Rai1-Rat1/Xrn2复合物、Dxo1/Ydr370C和DXO/Dom3Z的位置是不同的。Rai1-Rat1/Xrn2仅分布在细胞核中，DXO/Dom3Z主要在细胞核中，而Dxo1/Ydr370C则主要在细胞质中。分布位置的不同，说明mRNA加帽过程质量控制发生的位置不同，在哺乳动物中该过程发生在细胞核内，而在酵母中则细胞核内和细胞质都会发生。这可能也是酵母和哺乳动物细胞中mRNA出核方式不同的原因。在这两种有机体中，都存在一个保守的TREX（转录/导出）复合物，在mRNA出核中发挥关键作用。在酵母中，TREX募集是伴随着转录过程的，而在哺乳动物中，TREX复合物的募集是剪接和5’-帽子结构依赖性的。因此，哺乳动物mRNA出核的过程其实扮演着一个额外的mRNA加帽控制检查点的角色，它可以阻止非正常加帽的mRNA进入到细胞质中。酵母中缺乏这一机制，可能需要依靠细胞质中的Dxo1/Ydr370C来保证正确的加帽。体外实验中，Rai1对含有成熟甲基化帽子结构mRNA的脱帽活性最低，而Dox1/Ydr370C和DXO/Dom3Z的活性则相对极高。而在细胞内该脱帽活性被抑制，因为帽结合蛋白会优先与成熟帽子结构结合从而保护其免遭降解。体外试验中，不论细胞核还是细胞质中的帽结合蛋白（CBP20和elF4E），都可以有效抑制DXO/Dom3Z的脱帽活性。除此之外，DXO/Dom3Z对三甲基化帽子（m2,2,7GpppN）RNA也具有脱帽活性，并可能调节细胞内三甲基化加帽富鸟苷酸的小核RNA。4.Rai1及其同系物催化的结构基础Rai1及其同系物的结构包含两个高度扭曲的β-片和几个α-螺旋，他们覆盖了β-片的一些暴露的表面。活性位点位于整个蛋白结构的中心，位于β-片之间的交界面 (Fig. 1a)）。活性位点包含四个高度保守的基序，在DXO/Dom3Z中对应的是Arg132（基序I），Gly188-Tyr-Phe-Glu192（基序II，GΦXΦE，Φ为芳香族或疏水性氨基酸残基，X为任意氨基酸），Glu234-Val-Asp236（基序III，EhD，h为疏水性残基），Glu253-Leu-Lys255（基序IV，EhK）。Rai1及其同系物的结构与其它已知的蛋白结构上不具有同源性，但和D-(D/E)XK核酸酶的结构略有关联。尽管如此，Rai1同系物和这些核酸酶在序列上几乎没有相似性，且相似度主要限于基序III（EhD）和基序IV（EhK）上的天冬氨酸残基，在所有这些酶中，该残基作为金属离子结合位点。突变研究表明，Rai1及其同系物具有的所有三种酶活性，即：脱帽、焦磷酸化和5’-3’外切核酸酶活性，均来自这一活性位点。脱帽酶Dcp2是将m7Gpp帽子从mRNA5’端酶切下来，与之不同的是，Rai1及其同系物酶切的是第一和第二核苷酸之间的磷酸二酯键，从而释放出甲基化或非甲基化的GpppN帽子。对DXO/Dom3Z结构的研究，为Rai1及其同系物的酶催化机理提供了分子学基础。DXO/Dom3Z与模拟帽子m7GpppG（图1b）及寡核苷酸（图1c）形成的复合物的结构表明，加帽mRNA分子结合活性位点口袋，其中帽子（m7GpppG）和第一个核苷酸位于一侧，RNA分子的剩余部分位于另一侧，催化体系还包含位于中间的两个金属离子。Arg132（基序I）的侧链识别帽子结构的磷酸基团，金属离子与基序II、III、IV的侧链残基，第一第二核苷酸之间的磷酸基团，以及基序IVLeu254主链的羧基发生配位作用。基于这一结构信息，有人提出：其中一个金属离子激活水分子或羟基离子，其随后攻击第一和第二核苷酸之间的易裂磷酸键，在该基团5’端进行切割并释放游离GpppN。Lys255（基序IV）的侧链稳定了催化过程中的氧负离子中间体（图1d）。这种双金属离子催化机制在核酸酶中广泛存在。DXO/Dom3Z的结构表明，结合帽子和第一核苷酸的活性位点，也包含了5’末端核苷酸（pN1）或焦磷酸基团，催化5’端单磷酸或三磷酸RNA降解释放出pN1或焦磷酸基团，产生5’-3’前核糖核酸酶和焦磷酸酶活性。因此，Rai1及其同系物的三种看似不同的活性都使用了相同的催化机制，并且催化的产物取决于不同底物的不同结合模式。5.结束语与以往的认知不同，如今已知真核生物中mRNA的加帽过程并不总是能被完成。非正确加帽可能来自于加帽过程中的偶发错误，而加帽质量控制机制可以纠正这些错误。且/或，后续的证据表明，这是加帽过程主动调控的结果。例如，已有报道指出importin-α激活帽子甲基化。Myc调节许多mRNA帽子甲基化，而营养缺陷条件导致帽子甲基化比例普遍降低。mRNA加帽的调控因子和质量控制机制一起，可能为基因表达提供另一层调控。最近的研究极大地促进了我们对真核生物中mRNA加帽质量控制的了解。然而，目前对这一过程的理解还远远不够。例如：Rai1、Dxo1/Ydr370C和DXO/Dom3Z底物特异性的分子基础仍不清晰。是否可以通过已知的加帽质量控制因素清楚5’-二磷酸RNA，即加帽过程中产生的另一种中间体，还有待观察。重要的是，关于Rai1及其同系物如何与其它因素相互作用，以协调加帽质量控制与其它细胞过程，我们知之甚少。例如，在酵母中，已经证明Rai1-Rat1/Xrn2复合物与PolII的转录机制相关，且Rat1/Xrn2会在转录过程中就清除掉不正确加帽的初级转录产物。在哺乳动物中是否也是如此还未可知，如果也是，那么是哪些因素负责调动DXO/Dom3Z。这些问题的答案都还有待进一步研究。mRNA加帽在诸多细胞过程中发挥着重要的作用，而加帽过程质量控制机制的明确，为帽子结构紊乱异常及探究帽子结构在这些细胞过程中的功能提供了一种新颖的思路。DXO/Dom3Z的敲除就是一个例证，使得人们认识到，在哺乳动物中mRNA加帽与mRNA剪接和多聚腺苷酸化的关联比以往认为的还要密切。在这个方向上的进一步研究将带来对mRNA帽子结构生理作用更深入的理解。原文来源：mRNA quality control at the 5' end.J Zhejiang Univ Sci B.2014 May;15(5):438-43.识别微信二维码，添加生物制品圈小编，符合条件者即可加入微信群！请注明：姓名+研究方向！版权声明本公众号所有转载文章系出于传递更多信息之目的，且明确注明来源和作者，不希望被转载的媒体或个人可与我们联系(cbplib@163.com)，我们将立即进行删除处理。所有文章仅代表作者观点，不代表本站立场。

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