Replication of Human Endogenous Retroviruses in Modern Humans

人内源性逆转录病毒在现代人类中的复制

• 批准号: 7762721
• 负责人: David Michael Markovitz
• 金额: $ 143.53万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-28 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7762721
• 关键词: Antibiotic Resistance; Autoimmunity; Bioinformatics; Blood; Cancer Biology; Cancer Patient; Cell Line; Cells; Cloning; DNA Sequence; Disease; Elements; Endogenous Retroviruses; Genetic Materials; Genome; Green Fluorescent Proteins; HERVs; Human; Human Genetics; Human Genome; Individual; Inherited; Laboratories; Lead; Length; Life; Link; Malignant Neoplasms; Methods; Molecular; Mutation; Nuclear Magnetic Resonance; Pathogenesis; Patients; Proteins; RNA; Research Personnel; Role; Safety; Surrogate Markers; System; Techniques; Testing; Thinking; Vascular blood supply; Virus; conformational conversion; in vivo; public health relevance; transmission process; tumorigenesis

DESCRIPTION (provided by applicant): The functions of the approximately 98% of the human genome that do not encode human cellular proteins remain to be elucidated. Actively replicating endogenous retroviruses entered the human genome millions of years ago and became a stable part of the inherited genetic material, with retroviral elements presently making up approximately 8% of the modern human genome. These viruses subsequently acquired multiple mutations, leading to the widely-held assumption that they are no longer competent to replicate. However, in studying living patients rather than the standard cell lines, we have recently discovered surprising evidence suggesting that in certain patients with cancer HERV-K (HML-2), an endogenous retrovirus that is a relatively recent entrant into the human genome and has been linked to oncogenesis, might still be capable of replication. Replication and transmission of endogenous retroviruses is difficult to prove using standard techniques, however, as these viruses are already present in the genomes of all human cells. Therefore, we have assembled a diverse and expert group of investigators who will test the hypothesis that modern HERV-K (HML-2) can replicate using cutting-edge and complementary techniques. We will use a newly devised molecular system in which antibiotic resistance serves as a surrogate marker to assess whether we can passage virus from the blood of patients in the laboratory. We will also use high-throughput DNA sequencing and Bioinformatics to find full-length functional virus that is present in cancer patients but is not represented in the current draft of the human genome. An infectious clone that is representative of modern HERV-K (HML-2) will be made by using the information gained from the Bioinformatics studies and/or by cloning a full-length virus from the HERV-K RNA found in the blood of patients. This clone will then be tagged with green fluorescent protein and used to directly study replication. Building on our advances using Nuclear Magnetic Resonance (NMR) to visualize spatially correlated dynamics that direct RNA and protein conformational transitions, we will also develop NMR methods to visualize HERV-K (HML-2) replication in vivo. Proof that endogenous retroviruses can still replicate in modern humans will lead to a paradigm shift in thinking about these viruses, and will suggest a role for them in reshaping individual genomes. In addition, as increased expression of chromosomal endogenous retroviral sequences has been linked to cancer and autoimmunity, these findings will be relevant to understanding the pathogenesis of significant diseases. Finally, if replicating, infectious endogenous human retroviruses capable of causing disease can be found in the blood of given individuals, this information will have important implications for the safety of the blood supply. PUBLIC HEALTH RELEVANCE: Here we propose to prove that, contrary to the entrenched dogma, certain human endogenous retroviruses can still replicate in modern humans. Proof of this hypothesis would have substantial implications for human genetics, cancer biology, and the safety of the human blood supply.

描述（由申请人提供）：大约 98% 的人类基因组不编码人类细胞蛋白，其功能仍有待阐明。活跃复制的内源性逆转录病毒在数百万年前进入人类基因组，并成为遗传性遗传物质的稳定部分，逆转录病毒元件目前约占现代人类基因组的8%。这些病毒随后获得了多种突变，导致人们普遍认为它们不再具有复制能力。然而，在研究活体患者而不是标准细胞系时，我们最近发现了令人惊讶的证据，表明在某些癌症患者中存在 HERV-K (HML-2)，这是一种内源性逆转录病毒，相对较新进入人类基因组，并且已与肿瘤发生有关，可能仍然能够复制。然而，内源性逆转录病毒的复制和传播很难使用标准技术来证明，因为这些病毒已经存在于所有人类细胞的基因组中。因此，我们组建了一个多元化的专家研究小组，他们将测试现代 HERV-K (HML-2) 可以使用尖端和互补技术进行复制的假设。我们将使用一种新设计的分子系统，其中抗生素耐药性作为替代标记来评估我们是否可以在实验室中从患者的血液中传代病毒。我们还将利用高通量 DNA 测序和生物信息学来寻找癌症患者体内存在但在当前人类基因组草案中未出现的全长功能病毒。将利用从生物信息学研究中获得的信息和/或从患者血液中发现的 HERV-K RNA 克隆全长病毒来制作代表现代 HERV-K (HML-2) 的感染性克隆。然后，该克隆将被标记为绿色荧光蛋白，并用于直接研究复制。基于我们使用核磁共振 (NMR) 来可视化指导 RNA 和蛋白质构象转变的空间相关动力学的进展，我们还将开发 NMR 方法来可视化体内 HERV-K (HML-2) 复制。内源性逆转录病毒仍然可以在现代人类中复制的证据将导致对这些病毒的思考发生范式转变，并将表明它们在重塑个体基因组中的作用。此外，由于染色体内源性逆转录病毒序列表达的增加与癌症和自身免疫有关，这些发现将有助于了解重大疾病的发病机制。最后，如果可以在特定个体的血液中发现能够引起疾病的复制型、传染性内源性人类逆转录病毒，那么该信息将对血液供应的安全产生重要影响。 公共卫生相关性：在这里，我们建议证明，与根深蒂固的教条相反，某些人类内源性逆转录病毒仍然可以在现代人类中复制。这一假设的证明将对人类遗传学、癌症生物学和人类血液供应的安全产生重大影响。