The role of asymmetric protein dimethylation during the human papillomavirus lifecycle

感染人乳头瘤病毒(HPV)是美国最常见的性传播感染。除了导致宫颈癌和其他肛门癌以及头颈癌发病率上升外，估计全世界约有5%的癌症是由HPV引起的。重要的是，持续性感染而不是急性感染是(宫颈癌)发生的主要风险因素。因此，了解促进或限制病毒持久性的病毒-宿主相互作用对HPV生物学和人类癌症具有重要意义。直到最近，研究感染后HPV生命周期的早期事件，以及这些事件如何有助于建立阶段的初始基因组扩增和病毒的长期持续，一直是具有挑战性的。我们使用单细胞基因组学方法来确定与病毒感染和持久性有关的细胞因素。我们的初步数据表明，精氨酸N-甲基转移酶1(PRMT1)蛋白是病毒感染原代人宫颈细胞的重要因素。PRMT1调节几种可能与HPV生命周期相关的细胞功能。我们证明，抑制PRMT1会导致高度失调的病毒基因剪接模式。从机制上讲，我们发现病毒mRNA的m6A修饰显著增加，特别是在内含子中。这些数据涉及到RBM15，M6A甲基转移酶复合体的一个调节亚单位。我们假设PRMT1调节RBM15控制的m6A在病毒mRNA上的沉积，以暂时调节病毒感染过程中的选择性剪接。我们将(1)确定不对称蛋白质双甲基化在病毒生命周期中的重要性。(2)确定RBM15在PRMT1介导的病毒m6A甲基化调控中的作用。(3)检测m6A甲基化对病毒mRNA剪接的影响。抑制PRMT1会导致病毒剪接的失调。这项建议解决了我们关于如何调节HPV剪接以实现最佳感染、建立和长期持久性的知识中的一个关键空白。我们发现PRMT1是病毒剪接的调节因子，并假设这是通过病毒内含子上的靶向M6A沉积而发生的。由于PRMT1是一个确定的可用药靶点，这些结果可能会导致针对HPV诱导的癌症的新疗法。

Infections with human papillomaviruses (HPVs) are the most common sexually transmitted infection in the US. In addition to causing cervical and other anogenital cancers and a rising incidence of head and neck cancer, HPVs are responsible for an estimated 5% of cancers worldwide. Importantly, persistent infection, and not an acute infection is the primary risk factor for (cervical) cancer development. Thus, understanding the virus-host interplay that promotes or restrict viral persistence has important implications for HPV biology and human cancers. Until recently, it has been challenging to study the immediate early events of the HPV lifecycle following infection and how these events contribute to initial genome amplification during the establishment phase and long-term viral persistence. We used a single-cell genomics approach to identify cellular factors involved in viral infection and persistence. Our preliminary data identify protein arginine N-methyltransferase 1 (PRMT1) as an important factor upon viral infection of primary human cervical cells. PRMT1 regulates several cellular functions that may be relevant to the HPV lifecycle. We demonstrate that PRMT1 inhibition leads to a highly dysregulated splicing pattern of the viral genes. Mechanistically, we identify a dramatic increase in m6A modifications of viral mRNA, specifically in introns. These data implicate RBM15, a regulatory subunit of the m6A methyltransferase complex. We hypothesize that PRMT1 regulates RBM15 controlled m6a deposition on viral mRNA to temporally regulate alternative splicing throughout viral infection. We will (1) Determine the importance of asymmetric protein dimethylation throughout the viral lifecycle. (2) Determine the role for RBM15 in the PRMT1 mediated regulation of m6a methylation of viral mRNA. (3) Determine the effects of m6a methylation on viral mRNA splicing. PRMT1 inhibition leads to dysregulated viral splicing. This proposal addresses a critical gap in our knowledge relating to how HPV splicing is regulated for optimal infection, establishment, and long-term persistence. We identify PRMT1 as a regulator of viral splicing and hypothesize that this occurs through targeted m6A deposition on viral introns. Since PRMT1 is an established druggable target, these results may lead to novel therapeutics for HPV-induced cancers.