Regulation and Function of Viral and Endogenous Circular RNA in Cancer

单链环状RNA（circRNA）在癌症中富集，但其调控和生物学功能 大多数circRNA的结构仍不清楚。我们的长期目标是了解人类 在感染性疾病和肿瘤性疾病中的乳头瘤病毒（HPV）衍生的和内源性circRNA。的 本建议的具体目标是使人们更全面地了解该职能， 头颈部鳞状细胞癌（HNSCC）中存在的circRNA的调节。我们的中央 一种假设是，许多circRNA编码的RNA在调节和功能上与它们不同， 相应的全长RNA以促进HNSCC的发展。在初步研究中， 我们开发了创新的circRNA-Seq和Polysome RNA-Seq方案，发现许多 与邻近的非肿瘤粘膜相比，具有编码潜力的circRNA在HNSCC肿瘤中富集。在 此外，我们发现了HPV 16环状E7 RNA（circE 7），它被翻译成E7癌蛋白， 一种新的工具来了解circRNA形成和功能的机制和生理调节。 有了这些创新的工具和广泛的初步证据支持可行性，我们建议扩大 我们对HNSCC中circRNA的理解有三个相关的目的。首先，一个全面的转录组学 将生成多达50个HPV+和50个HPV- HNSCC肿瘤和邻近组织的特征，包括 RNA-Seq方法富集circRNA。此外，在该转录组学分析中鉴定的circRNA 和circE 7将被评估其作为存档的验证队列中的预后生物标志物的潜力。 HNSCC。第二，使用HPV 16 circE 7基因座，我们将确定调节细胞凋亡的顺式和反式元件。 circRNA在HNSCC中的形成，并确定它们是如何生理调节的。第三，我们将联合收割机 HNSCC circRNA-Seq数据集与最近完成的多核糖体RNA-Seq，以识别内源性circRNA 可能会被翻译。这些优先候选人将通过以下方式进行验证和定性： 严格控制的过表达和敲低实验，包括使用circRNA衍生的 特异性肽抗体。调查小组包括circRNA代谢，HPV生物学， 病理学，统计遗传学，头颈癌和流行病学，从而确保我们有能力 以及执行提案的资源。完成该提案将提供一个全球性的和详细的 理解circRNA及其在HNSCC中的调控。

Single-stranded circular RNAs (circRNA) are enriched in cancers, yet the regulation and biological function of most circRNA remains unclear. Our long-term goal is to understand the role of both human papillomavirus (HPV)-derived and endogenous circRNAs in both infectious and neoplastic diseases. The specific goal of this proposal is to generate a more comprehensive understanding of the function and regulation of circRNA that are present in head and neck squamous cell carcinoma (HNSCC). Our central hypothesis is that many circRNA are coding RNA that differ in regulation and function from their corresponding full-length RNAs in ways that promote the development of HNSCC. In preliminary studies, we have developed innovative circRNA-Seq and Polysome RNA-Seq protocols and found that many circRNA with coding potential are enriched in HNSCC tumors compared to adjacent non-tumor mucosa. In addition, our discovery of HPV16 circular E7 RNA (circE7), which is translated to the E7 oncoprotein, offers a novel tool to understand the mechanistic and physiological regulation of circRNA formation and function. With these innovative tools and extensive preliminary evidence supporting feasibility, we propose to expand our understanding of circRNAs in HNSCC in three related aims. First, a comprehensive transcriptomic profile from up to 50 HPV+ and 50 HPV- HNSCC tumors and adjacent tissue will be generated, including RNA-Seq approaches that enrich for circRNAs. Moreover, circRNA identified in this transcriptomic profiling and circE7 will be assessed for their potential as prognostic biomarkers in a validation cohort of archived HNSCC. Second, using the HPV16 circE7 locus, we will determine the cis and trans elements that regulate circRNA formation in HNSCC and determine how they are physiologically regulated. Third, we will combine HNSCC circRNA-Seq datasets with recently completed polysome RNA-Seq to identify endogenous circRNA that are likely to be translated. These prioritized candidates will be validated and characterized through rigorously controlled overexpression and knockdown experiments including the use of circRNA-derived specific peptide antibodies. The investigative team includes experts in circRNA metabolism, HPV biology, pathology, statistical genetics, head and neck cancer, and epidemiology, thus ensuring we have the skills and resources to execute the proposal. Completion of the proposal will provide both a global and detailed understanding of circRNA and their regulation in HNSCC.