Department of Microbiology and Immunology, Pennsylvania State University College of Medicine, Hershey, Pennsylvania, USA.
Department of Microbiology and Immunology, Pennsylvania State University College of Medicine, Hershey, Pennsylvania, USA
J Virol. 2018 May 29;92(12). doi: 10.1128/JVI.02096-17. Print 2018 Jun 15.
Secondary envelopment of human cytomegalovirus (HCMV) occurs through a mechanism that is poorly understood. Many enveloped viruses utilize the endosomal sorting complexes required for transport (ESCRTs) for viral budding and envelopment. Although there are conflicting reports on the role of the ESCRT AAA ATPase protein VPS4 in HCMV infection, VPS4 may act in an envelopment role similar to its function during other viral infections. Because VPS4 is normally recruited by the ESCRT-III complex, we hypothesized that ESCRT-III subunits would also be required for HCMV infection. We investigated the role of ESCRT-III, the core ESCRT scission complex, during the late stages of infection. We show that inducible expression of dominant negative ESCRT-III subunits during infection blocks endogenous ESCRT function but does not inhibit virus production. We also show that HCMV forms enveloped intracellular and extracellular virions in the presence of dominant negative ESCRT-III subunits, suggesting that ESCRT-III is not involved in the envelopment of HCMV. We also found that as with ESCRT-III, inducible expression of a dominant negative form of VPS4A did not inhibit the envelopment of virions or reduce virus titers. Thus, HCMV does not require the ESCRTs for secondary envelopment. However, we found that ESCRT-III subunits are required for efficient virus spread. This suggests a role for ESCRT-III during the spread of HCMV that is independent of viral envelopment. Human cytomegalovirus (HCMV) is a prevalent opportunistic pathogen in the human population. For neonatal and immunocompromised patients, HCMV infection can cause severe and possibly life-threatening complications. It is important to define the mechanisms of the viral replication cycle in order to identify potential targets for new therapies. Secondary envelopment, or acquisition of the membrane envelope, of HCMV is a mechanism that needs further study. Using an inducible fibroblast system to carefully control for the toxicity associated with blocking ESCRT-III function, this study determines that the ESCRT proteins are not required for viral envelopment. However, the study does discover a nonenvelopment role for the ESCRT-III complex in the efficient spread of the virus. Thus, this study advances our understanding of an important process essential for the replication of HCMV.
人类巨细胞病毒（HCMV）的二次包膜形成机制尚不清楚。许多包膜病毒利用内体分选复合物必需运输（ESCRTs）进行病毒出芽和包膜。尽管关于 ESCRT AAA ATPase 蛋白 VPS4 在 HCMV 感染中的作用存在相互矛盾的报告，但 VPS4 可能在包膜作用中类似于其在其他病毒感染中的功能。由于 VPS4 通常被 ESCRT-III 复合物募集，我们假设 ESCRT-III 亚基也需要 HCMV 感染。我们研究了 ESCRT-III，即核心 ESCRT 分裂复合物，在感染后期的作用。我们表明，在感染过程中诱导表达显性失活的 ESCRT-III 亚基会阻断内源性 ESCRT 功能，但不会抑制病毒产生。我们还表明，在存在显性失活的 ESCRT-III 亚基的情况下，HCMV 形成包膜的细胞内和细胞外病毒粒子，这表明 ESCRT-III 不参与 HCMV 的包膜形成。我们还发现，与 ESCRT-III 一样，诱导表达显性失活形式的 VPS4A 也不会抑制病毒粒子的包膜形成或降低病毒滴度。因此，HCMV 不需要 ESCRTs 进行二次包膜形成。然而，我们发现 ESCRT-III 亚基是病毒有效传播所必需的。这表明 ESCRT-III 在 HCMV 的传播中发挥作用，而与病毒包膜形成无关。人类巨细胞病毒（HCMV）是人群中普遍存在的机会性病原体。对于新生儿和免疫功能低下的患者，HCMV 感染可导致严重且可能危及生命的并发症。确定病毒复制周期的机制对于确定新疗法的潜在靶点非常重要。HCMV 的二次包膜形成，或包膜的获得，是一个需要进一步研究的机制。本研究使用可诱导的成纤维细胞系统仔细控制与阻断 ESCRT-III 功能相关的毒性，确定 ESCRT 蛋白不是病毒包膜形成所必需的。然而，该研究确实发现 ESCRT-III 复合物在病毒的有效传播中具有非包膜作用。因此，本研究加深了我们对 HCMV 复制过程中一个重要过程的理解。
