长非编码RNA对肿瘤抑制蛋白SAFB1的调控及机制研究

The aberrant expression of long noncoding RNAs (lncRNAs) is associated with mouse and human tumors. Several lines of evidence support the notion that lncRNAs can function through the regulation on protein activity and protein complexes. The tumor suppressor protein SAFB1 plays a significant role in the molecular network responsible for the regulation of cancer-, especially breast cancer-related stress. It is generally believed that the SAFB1-associated regulatory network is composed of protein coding genes. Our current results, however, indicate that epigenetic mechanism, regulated by lncRNA, is also involved in the regulation of SAFB1 function, as well the construction of SAFB1 regulation network. With our preliminary studies, we plan to characterize the molecular features that determine the lncRNA-SAFB1 interactions; investigate the regulatory mechanism underlying the effect of lncRNAs on SAFB1 activity; clarify the role of lncRNA-SAFB1 interactions in the process of breast cancer. It is expected that the proposed work will improve our understanding of breast cancer from the perspective of epigenetics, and will provide new rationales for exploring novel strategies for diagnosis and treatment of breast cancer.

长非编码RNA（lncRNA）的异常表达与肿瘤的发生发展密切相关；lncRNA对蛋白质活性或对蛋白质复合物的调节是近年来lncRNA研究的重大发现之一，也可能是lncRNA发挥其生物学功能的重要途径。肿瘤抑制蛋白SAFB1在负责调控癌症，特别是乳腺癌相关应激状态的分子网络中处于重要地位。当前普遍认为SAFB1调控网络完全由蛋白质编码基因构成，但我们的研究表明以lncRNA为代表的表观调控机制也参与了SAFB1功能的调节以及SAFB1调控网络的构建。基于前期研究基础，我们在本项目中将从分析lncRNA与SAFB1蛋白相互作用发生及选择的分子基础入手，系统研究lncRNA对SAFB1蛋白功能的调控及相应的调控机制，阐明lncRNA-SAFB1蛋白相互作用在乳腺癌发生发展中的意义，为从表观遗传学角度阐释乳腺癌的本质奠定基础，也为探讨乳腺癌的诊断及治疗提供新的思路。

SAFB1(scaffold attachment factor B1)作为已知的肿瘤抑制基因与乳腺癌关系密切，同时，SAFB1也是RNA结合蛋白。越来越多的证据表明lncRNA (long non-coding RNA)具有重要的调控作用，并广泛参与细胞的各种生理进程，如增殖生长、凋亡、应激反应等。为了筛选与SAFB1相互作用的lncRNA，我们采用SELEX-Seq (Systematic Evolution of Ligands by Exponential Enrichment with deep sequencing)技术对MCF7、SKBR3、MDA-MB-231、T-47D四种乳腺癌细胞系的细胞核RNA进行筛选，并从其中筛选得到了高丰度的763条RNA序列以及4种SAFB1识别基序（motif）。实验验证了基序以及基序所在的二级结构参与RNA与SAFB1蛋白相互作用。. 我们重点研究了筛选得到的lncRNA scaRNA9，scaRNA9能与SAFB1、ERα直接相互作用，并且过表达scaRNA9能加强SAFB1与ERα之间的相互作用，下调scaRNA9减弱SAFB1与ERα之间的相互作用。敲低scaRNA9能使ERα的靶基因OPG的表达水平上调，PUMA的表达水平下调。同时敲低scaRNA9能抑制MCF7细胞增殖以及克隆形成能力。. 综上所述，我们通过SELEX-Seq筛选得到了SAFB1识别的RNA的4种基序，并且证实了基序和由基序折叠形成的二级结构是RNA与SAFB1相互作用的基础，为RNA与蛋白相互作用的分子机制提供了理论依据。并且具有生物学功能的scaRNA9可能成为潜在的肿瘤治疗靶点。