Ets-1-dependent expression of vascular endothelial growth factor receptors is activated by latency-associated nuclear antigen of Kaposi's sarcoma-associated herpesvirus through interaction with Daxx
Ets-1-dependent expression of vascular endothelial growth factor receptors is activated by latency-associated nuclear antigen of Kaposi's sarcoma-associated herpesvirus through interaction with Daxx
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DOI：
10.1074/jbc.m602026200
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发表时间：
2006-09-22
影响因子：
4.8
通讯作者：
Fukazawa, Hidesuke
中科院分区：
文献类型：
--
作者：
Murakami, Yuko;Yamagoe, Satoshi;Fukazawa, Hidesuke
血管内皮生长因子（VEGF）及其受体在卡波西肉瘤（KS）病灶中高表达，在血管生成中发挥关键作用。卡波西肉瘤相关疱疹病毒 (KSHV/HHV8) 的潜伏期相关核抗原 (LANA) 具有与病毒潜伏期和 KSHV 诱导的肿瘤发生相关的多种功能。在本报告中，我们通过对稳定表达 LANA 的 HeLa 细胞进行免疫共沉淀分析，将 Daxx 鉴定为 LANA 结合蛋白。 LANA 与感染 KSHV 的 PEL 细胞系中的 Daxx 相关。 LANA 和 Daxx 也在体外结合，表明存在直接相互作用。根据结合测定的结果，LANA 内含有富含 Glu/Asp 结构域的区域和 Daxx 内包括第二对两亲螺旋的中心区域促成了相互作用。为了解决这种相互作用的生理意义，我们重点研究 Daxx 介导的 VEGF 受体基因调控。我们发现 Daxx 抑制 Ets-1 依赖性 Flt-1/VEGF 受体 1 基因表达，而 LANA 在报告基因检测中抑制 Daxx 的抑制。流式细胞术和实时PCR分析表明，表达LANA的人脐静脉内皮细胞（HUVEC）中VEGF受体-1和-2的表达显着增加。免疫共沉淀和免疫印迹实验表明 LANA 结合的 Daxx 抑制 Daxx 和 Ets-1 之间的相互作用。染色质免疫沉淀分析表明 Daxx 与 HUVEC 中的 VEGF 受体 1 启动子相关，而 LANA 表达会减弱这种关联。这些结果表明，LANA 通过干扰 Daxx 和 Ets-1 之间的相互作用，导致 KS 病变中 VEGF 受体的高表达。
Vascular endothelial growth factor ( VEGF) and its receptors are highly expressed in Kaposi's sarcoma(KS) lesion and play a key role in angiogenesis. Latency-associated nuclear antigen ( LANA) of Kaposi's sarcoma-associated herpesvirus (KSHV/HHV8) has multiple functions related to viral latency and KSHV-induced oncogenesis. In this report, we have identified Daxx as a LANA-binding protein by co-immunoprecipitation analysis of HeLa cells stably expressing LANA. LANA associated with Daxx in a PEL cell line infected with KSHV. LANA and Daxx also bound in vitro, suggesting direct interaction. From the results of binding assays, a region containing the Glu/Asp-rich domain within LANA, and a central region including the second paired amphipathic helix within Daxx contributed to the interaction. To address the physiological significance of this interaction, we focused on a Daxx-mediated VEGF receptor gene regulation. We found that Daxx repressed Ets-1-dependent Flt-1/VEGF receptor-1 gene expression, and that LANA inhibited the repression by Daxx in a reporter assay. Analyses of flow cytometry and real-time PCR revealed that expression of VEGF receptor-1 and -2 in LANA-expressing human umbilical vein endothelial cells(HUVECs) significantly increased. Co-immunoprecipitation and immunoblotting experiments suggested that LANA-bound Daxx to inhibit the interaction between Daxx and Ets-1. Chromatin immunoprecipitation assays showed that Daxx associated with VEGF receptor-1 promoter in HUVECs, and that LANA expression reduced this association. These results suggested that LANA contributes to a high expression of VEGF receptors in KS lesion by interfering with the interaction between Daxx and Ets-1.