Asparagine deprivation causes a reversible inhibition of Human Cytomegalovirus acute virus replication
Asparagine deprivation causes a reversible inhibition of Human Cytomegalovirus acute virus replication
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天冬酰胺剥夺会导致人巨细胞病毒急性病毒复制的可逆抑制
DOI：
10.1101/690016
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发表时间：
2019
期刊：
影响因子：
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通讯作者：
Lee C
中科院分区：
文献类型：
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作者：
Lee C
作为专性细胞内病原体，病毒依赖宿主细胞机制进行有效复制，宿主代谢为此目的被广泛操纵。高通量小干扰RNA （siRNA）筛选为鉴定新型宿主-病毒相互作用提供了一种系统的方法。在这里，我们报告了对人类巨细胞病毒（HCMV）重要宿主因子的大规模筛选，包括6,881个sirna。我们确定了47个前病毒因子和68个抗病毒因子参与了广泛的细胞过程，包括中介复合物、蛋白酶体功能和mRNA剪接。对其中一个命中点天冬酰胺合成酶（ASNS）的重点表征表明，HCMV复制严格要求天冬酰胺，这导致在DNA扩增开始之前病毒复制的早期阻断。这种作用是HCMV特有的，因为ASNS的敲除对单纯疱疹病毒1或甲型流感病毒的复制几乎没有影响，这表明这种限制不仅仅是由于蛋白质生产的失败。值得注意的是，添加外源天冬酰胺可以在感染后7天完全恢复病毒的复制，这表明尽管病毒的复制在早期阶段受到限制，但在初始感染后几天仍能保持完全复制的能力。本研究代表了对参与HCMV复制的宿主因子进行最全面的siRNA筛选，并鉴定出非必需氨基酸天冬酰胺是调节HCMV病毒复制的关键因子。这些结果对控制病毒潜伏期和HCMV患者的临床治疗具有重要意义。hcmv占实体器官移植患者相关并发症的60%以上。使用抗病毒药物（如更昔洛韦）进行预防性或预防性治疗，可减少早发性HCMV疾病的发生。然而，晚发性疾病仍然是一个重大问题，长期治疗，特别是在免疫系统受到抑制的患者中，大大增加了抗病毒药物耐药性的风险。自从更昔洛韦获得许可以来，很少有针对HCMV的抗病毒药物被开发出来，而且在这些药物中，相同的病毒基因往往是靶向的，这降低了这些药物对耐药菌株的有效性。另一种方法是针对病毒复制所必需的宿主基因。在这里，我们证明了HCMV的复制高度依赖于氨基酸天冬酰胺的水平，并且敲低一种参与天冬酰胺合成的关键酶会导致病毒复制的严重衰减。这些结果表明，通过饮食限制或化疗治疗来降低天冬酰胺水平可以限制患者的HCMV复制。
As obligate intracellular pathogens, viruses rely on the host cell machinery to replicate efficiently, with the host metabolism extensively manipulated for this purpose. High-throughput small interfering RNA (siRNA) screens provide a systematic approach for the identification of novel host-virus interactions. Here, we report a large-scale screen for host factors important for human cytomegalovirus (HCMV), consisting of 6,881 siRNAs. We identified 47 proviral factors and 68 antiviral factors involved in a wide range of cellular processes, including the mediator complex, proteasome function, and mRNA splicing. Focused characterization of one of the hits, asparagine synthetase (ASNS), demonstrated a strict requirement for asparagine for HCMV replication which leads to an early block in virus replication before the onset of DNA amplification. This effect is specific to HCMV, as knockdown of ASNS had little effect on herpes simplex virus 1 or influenza A virus replication, suggesting that the restriction is not simply due to a failure in protein production. Remarkably, virus replication could be completely rescued 7 days postinfection with the addition of exogenous asparagine, indicating that while virus replication is restricted at an early stage, it maintains the capacity for full replication days after initial infection. This study represents the most comprehensive siRNA screen for the identification of host factors involved in HCMV replication and identifies the nonessential amino acid asparagine as a critical factor in regulating HCMV virus replication. These results have implications for control of viral latency and the clinical treatment of HCMV in patients.IMPORTANCEHCMV accounts for more than 60% of complications associated with solid organ transplant patients. Prophylactic or preventative treatment with antivirals, such as ganciclovir, reduces the occurrence of early onset HCMV disease. However, late onset disease remains a significant problem, and prolonged treatment, especially in patients with suppressed immune systems, greatly increases the risk of antiviral resistance. Very few antivirals have been developed for use against HCMV since the licensing of ganciclovir, and of these, the same viral genes are often targeted, reducing the usefulness of these drugs against resistant strains. An alternative approach is to target host genes essential for virus replication. Here we demonstrate that HCMV replication is highly dependent on levels of the amino acid asparagine and that knockdown of a critical enzyme involved in asparagine synthesis results in severe attenuation of virus replication. These results suggest that reducing asparagine levels through dietary restriction or chemotherapeutic treatment could limit HCMV replication in patients.