Institute for Molecular Virology and McArdle Laboratory for Cancer Research, University of Wisconsin-Madison, Madison, Wisconsin, USA.
Institute for Molecular Virology and McArdle Laboratory for Cancer Research, University of Wisconsin-Madison, Madison, Wisconsin, USA
J Virol. 2019 May 15;93(11). doi: 10.1128/JVI.00430-19. Print 2019 Jun 1.
The human cytomegalovirus (HCMV) UL138 protein downregulates the cell surface expression of the multidrug resistance-associated protein 1 (MRP1) transporter. We examined the genetic requirements within UL138 for MRP1 downregulation. We determined that the acidic cluster dileucine motif is essential for UL138-mediated downregulation of MRP1 steady-state levels and inhibition of MRP1 efflux activity. We also discovered that the naturally occurring UL138 protein isoforms, the full-length long isoform of UL138 and a short isoform missing the N-terminal membrane-spanning domain, have different abilities to inhibit MRP1 function. Cells expressing the long isoform of UL138 show decreased MRP1 steady-state levels and fail to efflux an MRP1 substrate. Cells expressing the short isoform of UL138 also show decreased MRP1 levels, but the magnitude of the decrease is not the same, and they continue to efficiently efflux an MRP1 substrate. Thus, the membrane-spanning domain, while dispensable for a UL138-mediated decrease in MRP1 protein levels, is necessary for a functional inhibition of MRP1 activity. Our work defines the genetic requirements for UL138-mediated MRP1 downregulation and anticipates the possible evolution of viral escape mutants during the use of therapies targeting this function of UL138. HCMV UL138 curtails the activity of the MRP1 drug transporter by reducing its steady-state levels, leaving cells susceptible to killing by cytotoxic agents normally exported by MRP1. It has been suggested in the literature that capitalizing on this UL138-induced vulnerability could be a potential antiviral strategy against virally infected cells, particularly those harboring a latent infection during which UL138 is one of the few viral proteins expressed. Therefore, identifying the regions of UL138 required for MRP1 downregulation and predicting genetic variants that may be selected upon UL138-targeted chemotherapy are important ventures. Here we present the first structure-function examination of UL138 activity and determine that its transmembrane domain and acidic cluster dileucine Golgi sorting motif are required for functional MRP1 downregulation.
人巨细胞病毒（HCMV）UL138 蛋白下调多药耐药相关蛋白 1（MRP1）转运蛋白的细胞表面表达。我们研究了 UL138 内调控 MRP1 下调的遗传要求。我们确定酸性簇二亮氨酸基序是 UL138 介导的 MRP1 稳态水平下调和抑制 MRP1 外排活性所必需的。我们还发现，天然存在的 UL138 蛋白同工型，即 UL138 的全长长同工型和缺少 N 端跨膜结构域的短同工型，具有不同抑制 MRP1 功能的能力。表达 UL138 长同工型的细胞显示 MRP1 稳态水平降低，并且不能外排 MRP1 底物。表达 UL138 短同工型的细胞也显示 MRP1 水平降低，但降低的幅度不同，并且它们继续有效地外排 MRP1 底物。因此，跨膜结构域虽然对于 UL138 介导的 MRP1 蛋白水平降低不是必需的，但对于 MRP1 活性的功能抑制是必需的。我们的工作定义了 UL138 介导的 MRP1 下调的遗传要求，并预测了在针对 UL138 此功能的治疗中可能发生病毒逃逸突变体的进化。HCMV UL138 通过降低其稳态水平来抑制 MRP1 药物转运体的活性，使细胞易受通常由 MRP1 外排的细胞毒性剂的杀伤。文献中提出，利用这种 UL138 诱导的脆弱性可能是针对病毒感染细胞的潜在抗病毒策略，特别是在潜伏感染期间，UL138 是少数表达的病毒蛋白之一。因此，确定 UL138 下调 MRP1 所需的区域并预测在针对 UL138 的化学疗法选择时可能出现的遗传变异是重要的。在这里，我们首次对 UL138 活性进行了结构功能检查，并确定其跨膜结构域和酸性簇二亮氨酸高尔基体分拣基序是功能性 MRP1 下调所必需的。
