HCMV GPCR functions during latency and reactivation

人类巨细胞病毒（HCMV）是一种广泛传播的病原体，感染了美国大多数人口。这种病毒对发育中的胎儿以及缺乏有效免疫系统的儿童和成人构成重大威胁，往往导致严重疾病和死亡。一旦个体感染了HCMV，病毒就会在宿主体内终生存在，在造血室中处于潜伏或静止状态。在严重的免疫压力时期，病毒重新激活到活跃状态，允许传播和随后的疾病。除了immuno-naïve和血清阴性的器官移植受者外，HCMV的原发性感染很少引起疾病，而是再激活导致严重的并发症。因此，为了预防hcmv相关疾病，我们必须全面了解病毒潜伏期和再激活。在潜伏期，HCMV编码四种病毒G蛋白偶联受体（vgpcr）中的三种：US28、UL33和UL78。我们之前的研究表明，us28介导的信号是建立和维持潜伏期所必需的，并调节特定细胞靶点的表达，这些靶点调节主要即时早期（MIE）位点，这是潜伏到裂解开关的主要调节因子。我们的研究结果还揭示了UL33和UL78对病毒潜伏期有效再激活的要求。因此，我们假设vgpcr调节特定的宿主信号通路来调节潜伏期和再激活之间的平衡。为了探索这一假设，我们将利用新的方法，结合我们的病毒重组库，以及体外和离体潜伏期模型系统。在目的1中，我们将定义us28介导的信号传导的潜在机制，以及它如何影响潜伏期沉默因子在MIE位点的招募。在目标2中，我们将确定UL33信号如何影响MIE基因座的转激活和随后的再激活，以及描述US28和UL78之间的相互作用如何从潜伏复制到裂解复制的平衡。综上所述，本文提出的实验将有助于进一步了解vgpcr在潜伏期和再激活过程中的生物学功能，为未来研究开发新的治疗方法来预防HCMV再激活和疾病奠定基础。

Human cytomegalovirus (HCMV) is a wide-spread pathogen, infecting the majority of the population in the United States. This virus poses a significant threat to developing fetuses as well as to children and adults who lack a competent immune system, often causing severe disease and mortality. Once individuals acquire an HCMV infection, the virus remains with the host for life, in a latent or quiescent state in the hematopoietic compartment. During times of severe immunological stress, the virus reactivates to its active state, allowing for dissemination and subsequent disease. With the exception of the immuno-naïve and sero-negative organ transplant recipients, primary infection with HCMV rarely causes disease, but rather it is reactivation that leads to significant complications. Thus, to prevent HCMV-associated disease, we must gain a complete understanding of viral latency and reactivation. During latency, HCMV encodes three of the four viral G protein-coupled receptors (vGPCRs): US28, UL33, and UL78. We previously showed US28-mediated signaling is required for establishing and maintaining latency and modulates the expression of specific cellular targets that regulate the Major Immediate Early (MIE) locus, a master regulator in the latent-to-lytic switch. Our findings also reveal a requirement for UL33 and UL78 for efficient viral reactivation from latency. Therefore, we hypothesize the vGPCRs modulate specific host signaling pathways to regulate the balance between latency and reactivation. To explore this hypothesis, we will take advantage of novel approaches coupled with our arsenal of viral recombinants, as well as both in vitro and ex vivo latency model systems. In Aim 1, we will define mechanisms underlying US28-mediated signaling and how this impacts the recruitment of silencing factors to the MIE locus during latency. In Aim 2, we will determine how UL33 signaling impacts the transactivation of the MIE locus and subsequent reactivation, as well as delineate how the interaction between US28 and UL78 tips the balance from latent-to-lytic replication. In sum, the experiments proposed herein will lead to a greater understanding of the vGPCRs’ biological functions during latency and reactivation, laying the foundation for future studies to develop novel therapeutics to prevent HCMV reactivation and disease.