Investigating regulation and function of the cytosine deaminase APOBEC3A during cell cycle re-entry

研究细胞周期再进入过程中胞嘧啶脱氨酶 APOBEC3A 的调节和功能

• 批准号: BB/V010271/2
• 负责人: Tim Fenton
• 金额: $ 64.06万
• 依托单位: University of Southampton
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2022
• 资助国家: 英国
• 起止时间: 2022 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FV010271%2F2
• 关键词: Investigating; regulation; function; cytosine; deaminase

One way in which our bodies fight viral infections is to attack viral genomes (either DNA or RNA, depending on the virus) when viruses enter our cells and attempt to replicate. One protein involved in this immune response, APOBEC3A (A3A) modifies cytosine, one of the four building blocks of DNA and RNA, causing errors (mutations) and breaks in viral genes. A3A is not normally found at high levels in our cells except in certain specialised immune cells called macrophages. It is switched on in response to infection and plays an important role in protecting us from a range of viruses, however it is also activated in inflammatory conditions including eczema and psoriasis. Although A3A helps to defend us from viral infections, this protection comes at a cost, as we and others have shown that it can turn against our own genes, generating mutations that cause cancer. Numerous studies have confirmed that this process occurs in a large proportion of cancers, particularly those arising in the tissues (epithelia) that line the mouth and throat, lung, breast, bladder and cervix. Not only does A3A mutate our DNA during cancer development but it appears that A3A can continue to act in this rogue fashion while patients are receiving chemotherapy; driving drug-resistance and ultimately, treatment failure. This knowledge has stimulated initiatives in academia and industry to develop A3A inhibitors; drugs that could block this mutagenic activity in patients receiving chemotherapy, thereby preventing tumours from becoming resistant to the therapy. While this approach holds the potential to improve outcomes for millions of cancer patients, there is much we do not yet know about the way in which A3A is controlled and about the functions that it performs in the normal, healthy epithelial cells from which A3A-mutated cancers develop. Without this knowledge, we have little idea of what triggers rogue A3A activity, or what the side-effects of inhibiting A3A activity might be in patients. In this proposal, we set out a series of experiments to address these questions, based on three key findings that we have made from studying A3A in cultured human epithelial cells. 1) We have discovered that by mimicking a wound-healing response in epithelial cells, we can switch the A3A gene on to levels far higher than those previously seen in these cells and that remains at very high levels as these cells replicate their DNA, a time at which the DNA is potentially vulnerable to rogue A3A activity. 2) By deleting the A3A gene in epithelial cells, we have uncovered a previously unknown role for A3A in regulating the rate at which these cells divide, a role that is critical to understand if we are to anticipate the effects of targeting A3A for cancer therapy. 3) Recent studies have demonstrated that A3A can modify many cellular messenger RNAs, the intermediate transcripts that allow our genes to be translated into proteins but the significance of this activity remains unclear. We observe a very strong induction of this RNA-editing activity when we activate A3A in epithelial cells.Based on these novel observations, we will use our unique tools to identify in detail how the A3A gene is switched on in epithelial cells that have been stimulated to divide and will establish the role that A3A plays in regulating this process. We will conduct a comprehensive survey of A3A-mediated RNA editing events and will test our hypothesis that this activity allows A3A to change the rate at which key proteins are made.This project will result in a step-change in our knowledge of A3A regulation and function, its importance in normal epithelial biology and in pathologies ranging from inflammation to viral infections and cancer. This knowledge will be vital if we are to successfully harness A3A as a drug target. It will also address a fundamental question regarding the role of A3A-mediated RNA editing in controlling how our genes are expressed.

我们的身体对抗病毒感染的一种方法是，当病毒进入我们的细胞并试图复制时，攻击病毒基因组（DNA 或 RNA，具体取决于病毒）。 APOBEC3A (A3A) 是参与这种免疫反应的一种蛋白质，它会修饰胞嘧啶（DNA 和 RNA 的四个组成部分之一），从而导致病毒基因出现错误（突变）和断裂。除了某些称为巨噬细胞的特殊免疫细胞外，A3A 在我们的细胞中通常不会出现高水平。它会针对感染而启动，并在保护我们免受多种病毒侵害方面发挥重要作用，但它也会在包括湿疹和牛皮癣在内的炎症条件下被激活。虽然 A3A 有助于保护我们免受病毒感染，但这种保护是有代价的，因为我们和其他人已经证明它可以对抗我们自己的基因，产生导致癌症的突变。大量研究已经证实，这一过程发生在大部分癌症中，特别是那些出现在口腔和咽喉、肺癌、乳腺癌、膀胱和子宫颈的组织（上皮细胞）中的癌症。 A3A 不仅会在癌症发展过程中使我们的 DNA 发生突变，而且在患者接受化疗时，A3A 似乎还会继续以这种异常的方式发挥作用。导致耐药性并最终导致治疗失败。这些知识激发了学术界和工业界开发 A3A 抑制剂的积极性；可以阻断接受化疗的患者的这种诱变活性的药物，从而防止肿瘤对治疗产生耐药性。虽然这种方法有可能改善数百万癌症患者的治疗结果，但我们对 A3A 的控制方式以及它在正常、健康的上皮细胞（A3A 突变癌症是从这些细胞中发展而来）中发挥的功能知之甚少。如果没有这些知识，我们几乎不知道是什么触发了异常的 A3A 活性，或者抑制 A3A 活性可能会给患者带来什么副作用。在本提案中，我们基于在培养的人上皮细胞中研究 A3A 所获得的三个关键发现，开展了一系列实验来解决这些问题。 1) 我们发现，通过模仿上皮细胞中的伤口愈合反应，我们可以将 A3A 基因的开启水平远远高于之前在这些细胞中观察到的水平，并且在这些细胞复制其 DNA 时保持在非常高的水平，此时 DNA 可能容易受到流氓 A3A 活性的影响。 2) 通过删除上皮细胞中的 A3A 基因，我们发现了 A3A 在调节这些细胞分裂速率方面的先前未知的作用，如果我们要预测以 A3A 为靶点进行癌症治疗的效果，这一作用对于了解这一作用至关重要。 3) 最近的研究表明，A3A 可以修饰许多细胞信使 RNA，即允许我们的基因翻译成蛋白质的中间转录物，但这种活性的意义仍不清楚。当我们激活上皮细胞中的 A3A 时，我们观察到这种 RNA 编辑活性的强烈诱导。基于这些新的观察结果，我们将使用我们独特的工具来详细识别 A3A 基因如何在被刺激分裂的上皮细胞中开启，并将确定 A3A 在调节这一过程中所起的作用。我们将对 A3A 介导的 RNA 编辑事件进行全面调查，并将检验我们的假设，即这种活动使 A3A 能够改变关键蛋白质的生成速率。该项目将导致我们对 A3A 调节和功能、其在正常上皮生物学以及从炎症到病毒感染和癌症等病理学中的重要性的认识发生重大变化。如果我们要成功利用 A3A 作为药物靶点，这些知识将至关重要。它还将解决一个基本问题，即 A3A 介导的 RNA 编辑在控制我们的基因表达方式中的作用。

项目成果

Differentiation signals induce APOBEC3A expression via GRHL3 in squamous epithelia and squamous cell carcinoma

分化信号通过 GRHL3 在鳞状上皮和鳞状细胞癌中诱导 APOBEC3A 表达

• DOI: 10.21203/rs.3.rs-3997426/v1
• 发表时间: 2024
• 作者: Fenton T