长链非编码RNA CAR intergenic 10在细胞衰老中的作用和机制

Cellular senescence contributes to aging and/or age-related diseases. While p53/p21 and p16/pRB pathways are well known to play critical roles in cellular senescence, the molecular basis for its initiation and maintenance remains to be fully elucidated. Long non-coding RNAs (lncRNAs), which were recently found to be pervasively transcribed in the genome, are emerging as new players in gene expression regulation. LncRNAs can interact with its partner DNA, RNA, and/or protein to exert their function. However, knowledge regarding the contributions of lncRNAs to cellular senescence remains scarce.. In order to explore the role of the lncRNAs in cellular senescence, we screened for the differentially expressed lncRNAs among replicative senescent normal human fibroblasts (NHF), H2O2-induced premature senescent NHF and young NHF by microarray and validated the candidate lncRNAs by Q-PCR. LncRNA CAR intergenic 10 was found to be lowly expressed in senescent NHF cells. Silencing lncRNA CAR 10 resulted in an increased percentage of senescent NHF cells. Thus lncRNA CAR 10 possibly helps to prevent the onset of cellular senescence. To test this, we will assess the effect of lncRNA CAR 10 on cellular senescence by overexpressing or silencing the lncRNA in NHF; interrogate the partner proteins through RNA-pulldown and RNA-RIP. This study will show whether lncRNA CAR 10 delays cellular senescence and provide important clues about its molecular partners and its biological function.

衰老是老年性疾病发生的主要原因之一，已有研究表明长链非编码RNA（lncRNAs）参与调控细胞衰老进程。课题组前期对复制性衰老和氧化应激早熟性衰老的人成纤维细胞（NHF）进行了lncRNA芯片分析，lncRNA CAR intergenic 10是我们从芯片中筛选出的在两种衰老方式中均低表达的lncRNA，初步功能实验显示lncRNA CAR 10可延缓NHF细胞衰老，但具体作用机制尚待深入探讨。本课题拟采用逆转录病毒表达载体及siRNA干扰技术确定lncRNA CAR 10在NHF细胞衰老中的功能；利用RNA-pulldown、RNA-RIP等技术结合蛋白质谱等手段筛选、鉴定lncRNA CAR 10直接结合的蛋白分子及可能调控的靶基因；阐明CAR 10调控细胞衰老的下游通路和分子机制。本研究从lncRNA角度阐述细胞衰老的分子机制,将为延缓并减少老年性疾病的发生提供重要依据。

长链非编码RNAs（lncRNAs）是一类具有重要调控功能的分子，能从表观遗传修饰、转录及转录后等水平参与胚胎发育、细胞分化和衰老等生命过程的调控。.衰老是老年性疾病发生的主要原因之一，已有研究表明长链非编码RNA（lncRNAs）参与调控细胞衰老进程。课题组对复制性衰老和氧化应激早熟性衰老的人成纤维细胞（NHF）进行了lncRNA芯片分析，lncRNA CAR intergenic 10是我们从芯片中筛选出的在两种衰老方式中均低表达的lncRNA，而lncRNA BC042152是在两种衰老方式中均高表达的一种lncRNA。敲低CAR 10的NHF细胞存活率显著降低，表明lncRNA CAR 10可延缓NHF细胞衰老。lncRNA CAR 10基因与其附近的ADAM12基因有部分序列重叠，在siRNA干扰CAR 10表达的NHF细胞中，ADAM12基因的表达量也随之下调，提示ADAM12基因在细胞衰老过程中可能也起到一定的作用。lncRNA BC042152与其附近的POU2F2基因以头-头相接的方式存在于第19号染色体，二者转录起始点之间间距仅仅218bp。这两个基因共享一组双向的（bi-directional）启动子序列，在衰老的NHF细胞中lncRNA BC042152和POU2F2均高表达。.研究表明非编码RNA（ncRNAs）在肿瘤发生发展中起重要的调控作用。我们在高级别浆液性卵巢癌中发现非编码RNA miR-130a和PVT1显著高表达。我们的研究表明，miR-130a通过靶向抑制TSC1表达，激活mTOR信号通路，进而增强卵巢癌细胞增殖、克隆形成、侵袭能力并促进EMT过程。长链非编码RNA PVT1编码产生的三个miRNAs（miR-1204、miR-1205和miR-1206）可促进卵巢癌细胞的增殖、克隆形成能力及侵袭能力；PVT1还可能通过ceRNA机制发挥生物学功能。.本项研究将为深入理解非编码RNAs在细胞衰老和肿瘤发生发展过程中的分子机制、寻找理想的药物靶点提供理论和实验基础。