Manchester Institute of Biotechnology, The University of Manchester, Manchester, United Kingdom.
School of Chemistry, Faculty of Science and Engineering, The University of Manchester, Manchester, United Kingdom.
mBio. 2018 Jun 19;9(3):e01112-18. doi: 10.1128/mBio.01112-18.
The UL69 protein from human cytomegalovirus (HCMV) is a multifunctional regulatory protein and a member of the ICP27 protein family conserved throughout herpesviruses. UL69 plays many roles during productive infection, including the regulation of viral gene expression, nuclear export of intronless viral RNAs, and control of host cell cycle progression. Throughout the ICP27 protein family, an ability to self-associate is correlated with the functions of these proteins in transactivating certain viral genes. Here, we determined the domain boundaries of a globular ICP27 homology domain of UL69, which mediates self-association, and characterized the oligomeric state of the isolated domain. Size exclusion chromatography coupled with multiangle light scattering (SEC-MALS) revealed that residues 200 to 540 form a stable homo-tetramer, whereas a shorter region comprising residues 248 to 536 forms a homo-dimer. Structural analysis of the UL69 tetramer by transmission electron microscopy (TEM) revealed a dimer-of-dimers three-dimensional envelope with bridge features likely from a region of the protein unique to betaherpesviruses. The data provide a structural template for tetramerization and improve our understanding of the structural diversity and features necessary for self-association within UL69 and the ICP27 family. Human cytomegalovirus (HCMV) infection is widespread in the human population but typically remains dormant in an asymptomatic latent state. HCMV causes disease in neonates and adults with suppressed or impaired immune function, as the virus is activated into a lytic state. All species of herpesvirus express a protein from the ICP27 family which functions as a posttranscriptional activator in the lytic state. In HCMV, this protein is called UL69. The region of sequence conservation in the ICP27 family is a folded domain that mediates protein interactions, including self-association and functions in transactivation. All members thus far analyzed homo-dimerize, with the exception of UL69, which forms higher-order oligomers. Here, we use biochemical and structural data to reveal that UL69 forms stable tetramers composed of a dimer of dimers and determine a region essential for cross-dimer stabilization.
人巨细胞病毒（HCMV）的 UL69 蛋白是一种多功能调节蛋白，也是疱疹病毒家族中 ICP27 蛋白家族的成员。UL69 在有性感染过程中发挥多种作用，包括病毒基因表达的调节、无内含子的病毒 RNA 的核输出以及宿主细胞周期进程的控制。在整个 ICP27 蛋白家族中，自我缔合的能力与这些蛋白在转录激活某些病毒基因方面的功能相关。在这里，我们确定了介导自我缔合的 UL69 的 ICP27 同源结构域的球形结构域的边界，并对分离的结构域的寡聚状态进行了表征。大小排阻色谱法与多角度光散射（SEC-MALS）相结合表明，残基 200 至 540 形成稳定的同源四聚体，而包含残基 248 至 536 的较短区域形成同源二聚体。通过透射电子显微镜（TEM）对 UL69 四聚体的结构分析表明，二聚体-二聚体的三维信封具有桥接特征，可能来自于β疱疹病毒特有的蛋白区域。该数据为四聚化提供了结构模板，并提高了我们对 UL69 和 ICP27 家族内自我缔合的结构多样性和必要特征的理解。人巨细胞病毒（HCMV）在人群中广泛传播，但通常处于无症状潜伏状态。HCMV 会导致新生儿和免疫功能低下或受损的成年人发病，因为病毒被激活进入裂解状态。所有疱疹病毒物种都表达 ICP27 家族的一种蛋白，该蛋白在裂解状态下作为转录后激活物发挥作用。在 HCMV 中，这种蛋白称为 UL69。ICP27 家族中序列保守的区域是一个折叠结构域，介导蛋白相互作用，包括自我缔合和转录激活功能。迄今为止分析的所有成员都形成同源二聚体，除了 UL69，它形成更高阶的寡聚体。在这里，我们使用生化和结构数据揭示了 UL69 形成稳定的四聚体，由二聚体组成的二聚体，并且确定了对交叉二聚体稳定至关重要的区域。
