新LncRNA(REAT1)靶向NOD1调控鸡T淋巴细胞增殖和功能的分子机制

Reticuloendotheliosis virus (REV) can induce avian immune organ or blood T lymphocytes damage and lead to immune suppression, resulting in huge economic losses to the poultry industry. However, the pathogenic mechanism and host immune response at molecular level remain unclear. In the previous study we found a novel LncRNA (REAT1) targeting NOD1 by analyzing the transcriptome with RNA-seq in the well-established models of REV infected lymphocyte and SPF chicken. Furthermore, we observed NOD1 up-regulation and MAPK activation in the immune responses induced by REV infection. In this project we propose that LncRNA (REAT1) targeting NOD1 gene may regulate MAPK pathways and then control the proliferation and function of T lymphocytes in REV infection. To confirm our hypothesis, CHIRP-seq, ChIP-seq, mass spectrometry and dual luciferase reporter system will be performed to identify the role of REAT1 in the innate immune response of REV infection with in vivo and in vitro models. In particular, the molecular mechanism of REAT1 in regulating NOD1 expression and activating MAPK signaling pathway will be fully investigated to reveal how REAT1 regulates the proliferation and function of T lymphocytes after REV infection. This study will provide fundamental understanding on the molecular mechanism of LncRNA (REAT1) in immune regulation to against REV infection.

禽网状内皮组织增生症病毒（REV）可引起禽类免疫器官或血液T淋巴细胞损伤，导致机体免疫抑制，给家禽业造成巨大经济损失。但是REV的致病分子机理与机体免疫应答机理尚不清楚。本项目前期建立了REV感染的SPF鸡和T淋巴细胞模型，并通过RNA-seq测序发现新LncRNA (REAT1)靶向NOD1；而NOD1在鸡血液T淋巴细胞感染REV后表达上调，并激活MAPK信号通路。推测REAT1可能通过NOD1介导MAPK通路调节T淋巴细胞增殖和功能。基于前期基础，以REAT1为切入点，联合使用CHIRP-seq、ChIP-seq、质谱、双荧光素酶报告系统等技术，从体内、外两个层面明确REAT1调控鸡REV感染的先天免疫反应，重点探究REAT1通过靶向NOD1调节MAPK信号通路，调控REV病毒感染后鸡T淋巴细胞增殖和功能的分子机制，为全面揭示鸡REV感染的分子致病机理奠定新思路和理论依据。

禽网状内皮组织增生症病毒（Reticuloendotheliosis virus，REV）感染鸡群后能够引起鸡群不同程度的免疫抑制状态。REV不仅严重影响鸡群的生产性能，更制约着家禽业的经济发展。机体先天性免疫应答系统能够抵御80%以上的病毒入侵过程，有效保证机体被病毒入侵后的内环境稳定。基于前期研究结果发现鸡感染REV后一个新的LncRNA（REAT1）被激活，并通过靶向调控机体的先天性免疫应答机制中NLRs通路的重要成员NOD1基因表达，激活NOD-like受体、MAPK和AP1-NF-kB信号通路，进而调控下游免疫相关因子的表达和分泌，以抵御和应答REV病毒的侵入。但REAT1能够靶向调控NOD1表达并激活MAPK和NF-kB信号通路，从而影响T淋巴细胞以免疫应答鸡REV感染的分子机制尚不清楚。. 项目的研究内容主要包括（1）明确REAT1这一新的非编码RNA在REV感染后鸡T淋巴细胞中的免疫调控功能；（2）阐明REAT1调控靶基因NOD1表达，进而调控鸡T淋巴细胞增殖和功能的分子机制。通过利用5’-RACE、3’-RACE、构建其过表达载体和干扰载体、细胞转染、双荧光素酶报告系统检测、活体病毒转染等技术与实验，系统研究了REAT1这一新的非编码RNA在REV感染后鸡T淋巴细胞的免疫调节功能及其调控靶基因NOD1表达的分子机理等内容，结果表明REAT1与NOD1基因存在靶向调控作用，REAT1表达增加能够上调靶基因NOD1表达，并促进免疫因子IL-8分泌水平，继而通过NOD-like受体、MAPK和AP1-NF-kB等信号通路影响淋巴细胞免疫应答鸡REV感染的功能和分子机理。. 目前，禽网状内皮组织增生症感染率在我国已达到20%-30%，因此寻找针对REV行之有效的预防和治疗措施已成为当务之急。当下可行的病原学和血清学检测，阻断REV的垂直传播的方法，治标不治本、效果不理想。因此本项目以家禽抗病毒的免疫系统为核心，深入探究REV的致病机理，相关研究成果将为最终揭示REV感染宿主的免疫抑制和致病机理增添新的依据，也将为未来的高抗病力鸡的生物育种提供思路和方向。