Pathogen Molecular Biology Department, London School of Hygiene and Tropical Medicine, University of London, Keppel St., London WC1E 7HT, UK.
University Clinic of Freiburg, Department of Hematology and Oncology, Freiburg, Germany.
J Gen Virol. 2013 Jul;94(Pt 7):1624-1635. doi: 10.1099/vir.0.050153-0. Epub 2013 Mar 27.
Human monocytes expressing CCR2 with CD14 and CD16 can mediate antigen presentation, and promote inflammation, brain infiltration and immunosenescence. Recently identified roles are in human immunodeficiency virus infection, tuberculosis and parasitic disease. Human herpesvirus 6B (HHV-6B) encodes a chemokine, U83B, which is monospecific for CCR2, and is distinct from the related HHV-6A U83A, which activates CCR1, CCR4, CCR5, CCR6 and CCR8 on immune effector cells and dendritic cells. These differences could alter leukocyte-subset recruitment for latent/lytic replication and associated neuroinflammatory pathology. Therefore, cellular interactions between U83A and U83B could help dictate potential tropism differences between these viruses. U83A specificity is maintained in the 38-residue N-terminal spliced-truncated form. Here, we sought to determine the basis for the chemokine receptor specificity differences and identify possible applications. To do this we first analysed variation in a natural host population in sub-Saharan Africa where both viruses are equally prevalent and compared these to global strains. Analyses of U83 N-terminal variation in 112 HHV-6A and HHV-6B infections identified 6/38 U83A or U83B-specific residues. We also identified a unique single U83A-specific substitution in one U83B sequence, 'U83BA'. Next, the variation effects were tested by deriving N-terminal (NT) 17-mer peptides and assaying activation of ex vivo human leukocytes, the natural host and cellular target. Chemotaxis of CCR2+ leukocytes was potently induced by U83B-NT, but not U83BA-NT or U83A-NT. Analyses of the U83B-NT activated population identified migrated CCR2+, but not CCR5+, leukocytes. The U83BA-NT asparagine-lysine14 substitution disrupted activity, thus defining CCR2 specificity and acting as a main determinant for HHV-6A/B differences in cellular interactions. A flow-cytometry-based shape-change assay was designed, and used to provide further evidence that U83B-NT could activate CCR2+CD14+CD16+ monocytes. This defines a potential antiviral target for HHV-6A/B disease and novel peptide immunomodulator for proinflammatory monocytes.
表达 CCR2 的人单核细胞与 CD14 和 CD16 可以介导抗原呈递，并促进炎症、脑浸润和免疫衰老。最近发现的作用是在人类免疫缺陷病毒感染、结核病和寄生虫病中。人类疱疹病毒 6B（HHV-6B）编码一种趋化因子 U83B，它是 CCR2 的单特异性趋化因子，与相关的 HHV-6A U83A 不同，后者激活免疫效应细胞和树突状细胞上的 CCR1、CCR4、CCR5、CCR6 和 CCR8。这些差异可能改变潜伏/裂解复制和相关神经炎症病理学的白细胞亚群募集。因此，U83A 和 U83B 之间的细胞相互作用可以帮助决定这些病毒之间潜在的嗜性差异。38 个残基的 N 端剪接截断形式保持 U83A 的特异性。在这里，我们试图确定趋化因子受体特异性差异的基础，并确定可能的应用。为此，我们首先分析了在两种病毒同样流行的撒哈拉以南非洲的自然宿主人群中的变异，并将这些与全球株进行了比较。对 112 例 HHV-6A 和 HHV-6B 感染中 U83 的 N 端变异进行分析，确定了 6/38 个 U83A 或 U83B 特异性残基。我们还在一个 U83B 序列中发现了一个独特的 U83A 特异性取代，即“U83BA”。接下来，通过推导 N 端（NT）17 个氨基酸肽并检测其对体外人类白细胞、天然宿主和细胞靶标的激活作用，测试了变异的影响。U83B-NT 强烈诱导 CCR2+白细胞的趋化作用，但 U83BA-NT 或 U83A-NT 则不能。对 U83B-NT 激活的群体的分析鉴定出迁移的 CCR2+，而不是 CCR5+白细胞。U83BA-NT 的天冬酰胺-赖氨酸 14 取代破坏了活性，从而确定了 CCR2 的特异性，并成为 HHV-6A/B 细胞相互作用差异的主要决定因素。设计了一种基于流式细胞术的形态变化测定法，并用于提供进一步的证据，表明 U83B-NT 可以激活 CCR2+CD14+CD16+单核细胞。这为 HHV-6A/B 疾病定义了一个潜在的抗病毒靶点，并为促炎单核细胞提供了一种新型的肽免疫调节剂。
