H. M. Bligh Cancer Research Laboratories, Department of Microbiology and Immunology, Chicago Medical School, Rosalind Franklin University of Medicine and Science, North Chicago, IL 60064, USA.
Proc Natl Acad Sci U S A. 2012 May 8;109(19):E1163-72. doi: 10.1073/pnas.1119592109. Epub 2012 Apr 16.
Kaposi's sarcoma-associated herpesvirus (KSHV), etiologically associated with Kaposi's sarcoma, uses integrins (α3β1, αVβ3, and αVβ5) and associated signaling to enter human dermal microvascular endothelial cells (HMVEC-d), an in vivo target of infection. KSHV infection activated c-Cbl, which induced the selective translocation of KSHV into lipid rafts (LRs) along with the α3β1, αVβ3, and xCT receptors, but not αVβ5. LR-translocated receptors were monoubiquitinated, leading to productive macropinocytic entry, whereas non-LR-associated αVβ5 was polyubiquitinated, leading to clathrin-mediated entry that was targeted to lysosomes. Because the molecule(s) that integrate signal pathways and productive KSHV macropinocytosis were unknown, we immunoprecipitated KSHV-infected LR fractions with anti-α3β1 antibodies and analyzed them by mass spectrometry. The tyrosine kinase EphrinA2 (EphA2), implicated in many cancers, was identified in this analysis. EphA2 was activated by KSHV. EphA2 was also associated with KSHV and integrins (α3β1 and αVβ3) in LRs early during infection. Preincubation of virus with soluble EphA2, knockdown of EphA2 by shRNAs, or pretreatment of cells with anti-EphA2 monoclonal antibodies or tyrosine kinase inhibitor dasatinib significantly reduced KSHV entry and gene expression. EphA2 associates with c-Cbl-myosin IIA and augmented KSHV-induced Src and PI3-K signals in LRs, leading to bleb formation and macropinocytosis of KSHV. EphA2 shRNA ablated macropinocytosis-associated signaling events, virus internalization, and productive nuclear trafficking of KSHV DNA. Taken together, these studies demonstrate that the EphA2 receptor acts as a master assembly regulator of KSHV-induced signal molecules and KSHV entry in endothelial cells and suggest that the EphA2 receptor is an attractive target for controlling KSHV infection.
卡波济肉瘤相关疱疹病毒（KSHV）与卡波济肉瘤的病因有关，它利用整合素（α3β1、αVβ3 和 αVβ5）和相关信号进入人类真皮微血管内皮细胞（HMVEC-d），这是感染的体内靶标。KSHV 感染激活了 c-Cbl，c-Cbl 诱导 KSHV 与 α3β1、αVβ3 和 xCT 受体一起选择性转位到脂筏（LRs），但不与 αVβ5 转位。LR 转位的受体被单泛素化，导致有效的巨胞饮进入，而非 LR 相关的 αVβ5 被多泛素化，导致网格蛋白介导的进入，该进入被靶向溶酶体。由于整合信号途径和 KSHV 巨胞饮作用的分子尚不清楚，我们用抗-α3β1 抗体免疫沉淀 KSHV 感染的 LR 级分，并通过质谱分析它们。在这项分析中，鉴定出酪氨酸激酶 EphrinA2（EphA2），它与许多癌症有关。KSHV 激活了 EphA2。EphA2 也在感染早期与 KSHV 和整合素（α3β1 和 αVβ3）一起存在于 LRs 中。病毒与可溶性 EphA2 预孵育、shRNA 敲低 EphA2、或用抗 EphA2 单克隆抗体或酪氨酸激酶抑制剂 dasatinib 预处理细胞，均可显著降低 KSHV 的进入和基因表达。EphA2 与 c-Cbl-myosin IIA 相关，并增强 KSHV 在 LRs 中诱导的Src 和 PI3-K 信号，导致 KSHV 疱疹的泡状形成和巨胞饮作用。EphA2 shRNA 使巨胞饮相关信号事件、病毒内化和 KSHV DNA 的有效核转位消失。综上所述，这些研究表明，EphA2 受体是 KSHV 诱导的信号分子和内皮细胞中 KSHV 进入的主要组装调节剂，并表明 EphA2 受体是控制 KSHV 感染的有吸引力的靶点。
