Versatile functions of LANA in KSHV pathogenesis

 申请者描述(申请人提供)：本申请由中国科学院上海巴斯德研究所研究员中国申请，是应PAR-14-080《包括艾滋病在内的传染病国际研究(R01)》的要求提交的。随着艾滋病毒感染，KSHV和KS已成为世界范围内的一个重要健康问题。艾滋病毒人群中KSHV相关肿瘤发病率的增加引起了国际关注，特别是在中国等欠发达国家。阐明KSHV诱导肿瘤发生的机制将为肿瘤治疗提供新的靶点。在KS皮损中，大多数肿瘤细胞潜伏感染KSHV并表达病毒 潜伏相关核抗原(LANA)等蛋白质，提示病毒潜伏感染在肿瘤发生中起重要作用。利用串联亲和纯化(TAP)技术，我们鉴定了Krüppel相关盒结构域相关蛋白-1(KAP1)是一种新的LANA结合蛋白。KAP1作为转录抑制因子，通过募集组蛋白脱乙酰酶和甲基转移酶复合体来改变表观遗传状态。我们先前的研究表明，LANA将KAP1招募到KSHV基因组的RTA启动子区域，参与了LANA对RTA的转录抑制。由于KAP1是一个重要的转录抑制因子，我们假设LANA能够通过与KAP1的相互作用来调节宿主基因的表达。在这个项目中，我们将确定LANA是否以及如何通过下调细胞肿瘤抑制基因的表达来促进KSHV感染细胞的促增殖基因签名。另一方面，我们以前的研究表明，LANA通过BMP激活的细胞核内持续的p-Smad1上调BMP-Smad1-ID信号，并增强其对BMP靶基因ID启动子的负载量。我们发现ID蛋白在KSHV转化的KMM细胞中显著上调，并在人KS皮损中大量表达。值得注意的是，BMP-Smad1-ID通路的遗传抑制在体外和体内显著阻止了KSHV转化细胞的致癌表型。有趣的是，ID蛋白在胚胎发育和肿瘤形成过程中也在血管生成中发挥关键作用。因此，我们建议确定LANA是否调节BMP-Smad1-ID信号以促进肿瘤血管生成，因为丰富的血管生成是KS的特征，也是KS进展的关键因素。此外，我们还将确定LANA如何调控BMP诱导的p-Smad1激活的终止过程，以及ID蛋白对KMM细胞血管生成表型的作用机制。最后，我们将使用不同的BMP信号抑制剂来评价它们对肿瘤致瘤性和促血管生成的抑制效果。 KMM细胞的表型。通过我们的研究，我们希望对LANA在KSHV相关恶性肿瘤中的多功能功能提供新的见解。

 DESCRIPTION (provided by applicant): This application by investigators at the Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, P.R. China, is submitted in response to PAR-14-080 'International Research in Infectious Diseases, including AIDS (R01)'. Along with HIV infection, KSHV and KS have emerged as an important health problem worldwide. The increase in incidence of KSHV-associated tumors in the HIV population is of international concern especially in under-developed countries, such as China. Studies towards the elucidation of the mechanism for KSHV-induced tumorigenesis will provide new therapeutic targets. In KS lesions, the majority of tumor cells are latently infected by KSHV and express viral proteins such as latency associated nuclear antigen (LANA), suggesting an essential role for viral latent infection in tumorigenesis. Using tandem affinity purification (TAP) technology, we identified Krüppel associated box domain associated protein-1 (KAP1) as a novel LANA-binding protein. KAP1 functions as a transcriptional repressor and can change epigenetic state by recruiting histone deacetylase and methyltransferase complex. We previously showed that LANA recruited KAP1 to the RTA promoter region of the KSHV genome, which was involved in transcriptional repression of RTA by LANA. Since KAP1 is an important transcription corepressor, we hypothesize that LANA is capable of regulating global host gene expression through interaction with KAP1. In this project, we will determine whether and how LANA contributes to the pro-proliferative gene signature of KSHV infected cell by down-regulating expression of cellular tumor repressor genes. On the other hand, our previous studies showed that LANA up-regulated BMP-Smad1-Id signaling through sustained BMP-activated p-Smad1 in the nucleus and enhanced its loading on the promoter of BMP target gene Id. We showed that Id proteins were significantly up-regulated in KSHV transformed KMM cells and abundantly expressed in human KS lesions. Strikingly, genetic inhibition of the BMP-Smad1-Id pathway significantly blocked the oncogenic phenotype of KSHV-transformed cells in vitro and in vivo. Interestingly, Id proteins also play critical roles in angiogenesis both during embryogenesis and tumor formation. Thus, we proposed to determine whether LANA modulates BMP-Smad1-Id signaling to facilitate tumor angiogenesis, since abundant angiogenesis is the feature of KS and is the key factor for KS progress. Moreover, we will also determine how LANA regulates the termination process of BMP-induced p-Smad1 activation and the mechanisms by which of Id proteins contribute to angiogenic phenotype of KMM cells. Finally, we will use various BMP signaling inhibitors to evaluate their inhibition efficacy on the tumorigenicity and pro-angiogenic phenotype of KMM cells. Through our studies, we expect to reveal novel insights onto the versatile functions of LANA in KSHV-related malignancies.