Non-canonical chimeric proteins generated during Adenovirus infection

腺病毒感染期间产生的非典型嵌合蛋白

• 批准号: 10448505
• 负责人: Matthew D. Weitzman
• 金额: $ 26.4万
• 依托单位: CHILDREN'S HOSP OF PHILADELPHIA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-09 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10448505
• 关键词: Address; Adenovirus Infections; Adenovirus Protein; Adenoviruses; Affect; Alternative Splicing; Cell Line; Cell physiology; Cells; Chimeric Proteins; Code; Complement; Complex; DNA; DNA Packaging; DNA Viruses; DNA biosynthesis; DNA-Binding Proteins; Data; Development; Family; Gene Expression; Gene Expression Regulation; Generations; Genes; Genetic; Genetic Transcription; Genome; Goals; Host Defense; Individual; Infection; Knowledge; Messenger RNA; Modeling; Molecular; Morphogenesis; Morphology; Nuclear; Open Reading Frames; Outcome; Pathway interactions; Phase; Poly A; Polyadenylation; Positioning Attribute; Production; Proteins; Proteome; RNA; RNA Polymerase II; RNA Processing; RNA Splicing; Reagent; Research; Resolution; Seminal; Series; Signal Transduction; Site; System; Technical Expertise; Technology; Transcript; Transcriptional Regulation; Translations; Viral; Viral Genes; Viral Genome; Viral Packaging; Viral Proteins; Virus; Virus Diseases; Virus Replication; base; ds-DNA; gene product; human DNA; mutant; novel; polypeptide; promoter; protein function; transcriptome; viral DNA; viral RNA; virus host interaction

PROJECT SUMMARY Adenovirus (AdV) and other human DNA viruses with limited genome size have maximized gene expression and coding potential through the use of alternative splicing and polyadenylation. The AdV double-stranded DNA genome has served as a powerful system for seminal discoveries in gene regulation and RNA processing. There are five early transcription units (E1A, E1B, E2, E3 and E4) which are transcribed by host RNA polymerase II and processed by cellular machinery. Each viral transcription unit is driven by a designated promoter and gives rise to multiple mRNAs generated by alternative splicing. The canonical proteins produced from each viral transcript have been assigned functions through the study of infection with viral mutants. The E2 region encodes three proteins involved in viral DNA replication. The E4 region encodes multifunctional proteins involved in regulation of transcription, splicing and translation of viral mRNAs, as well as antagonizing intrinsic cellular defenses. We combined short-read and direct long-read sequencing technologies to define viral transcripts during AdV infection, and thus generated the most complete annotation of the virus transcriptome. Our analysis identified a number of non-canonical chimeric proteins that fuse polypeptides from different early transcriptional units. Here we focus on one of these unexpected gene products that we have designated E4orf6/DBP since it spans the E2 and E4 regions. Our preliminary data show that this novel chimeric protein is robustly expressed, and we propose that it could provide a yet unidentified function during AdV infection. In Aim 1 we will evaluate how the transcript is generated, and the importance of its interacting partners. In Aim 2 we will determine functions of the fusion protein, and its impact on infection. We have generated or accumulated all reagents and technical expertise necessary to define the functions of the E4orf6/DBP chimeric protein. Our findings challenge the concept of discrete early transcriptional units with their own designated promoter and poly(A) site, and suggest that studying individual open reading frames may not reflect the true complexity of the viral transcriptome. Our studies aim to uncover molecular mechanisms that regulate virus-host interactions by providing a deeper understanding of non-canonical fusion proteins that expand the diversity of the viral proteome. The proposal will generate a model approach for interrogating functions of non-canonical chimeric proteins generated by viruses with complex viral proteomes.

项目摘要 腺病毒（AdV）和其他基因组大小有限的人类DNA病毒具有最大化的基因表达 和编码潜力。AdV双链 DNA基因组作为一个强大的系统，在基因调控和RNA方面有着开创性的发现。 处理.宿主转录的早期转录单位有E1 A、E1 B、E2、E3和E4 RNA聚合酶II和处理的细胞机器。每个病毒转录单位由一个指定的 启动子并产生通过选择性剪接产生的多个mRNA。产生的典型蛋白质 通过对病毒突变体感染的研究，每个病毒转录本的基因都被赋予了功能。的 E2区编码三种参与病毒DNA复制的蛋白。E4区编码多功能 参与调节病毒mRNA转录、剪接和翻译的蛋白质，以及拮抗 内在的细胞防御我们结合了短读和直接长读测序技术， AdV感染过程中的病毒转录本，从而产生了最完整的病毒注释 转录组我们的分析鉴定了许多非典型的嵌合蛋白，其融合来自 不同的早期转录单位在这里，我们集中在这些意想不到的基因产物之一，我们有 命名为E4 orf 6/DBP，因为它跨越E2和E4区域。我们的初步数据显示这部小说 嵌合蛋白是稳健的表达，我们提出，它可以提供一个尚未确定的功能， 腺病毒感染。在目标1中，我们将评估转录本是如何产生的，以及其相互作用的重要性。 伙伴在目标2中，我们将确定融合蛋白的功能及其对感染的影响。我们有 产生或积累了所有必要的试剂和技术专长，以确定的功能， E4 orf 6/DBP嵌合蛋白。我们的发现挑战了离散早期转录单位的概念， 他们自己指定的启动子和poly（A）位点，并建议研究个人的开放阅读框架， 并不能反映病毒转录组的真实复杂性。我们的研究旨在揭示分子机制 通过提供对非典型融合蛋白的更深入理解来调节病毒-宿主相互作用， 扩大病毒蛋白质组的多样性。该提案将产生一个审讯的模式方法， 由具有复杂病毒蛋白质组的病毒产生的非典型嵌合蛋白的功能。