XIAP基因3’UTR的异常可变性剪切对肝细胞癌恶性进展的作用和分子机制

Study on miRNAs is one of the hottest fields in scientific research in recent years. However, the overwhelm studies on miRNAs did not bring clear and consistent understanding to people. There are even confusing and contradictory reports existed in the cancer research of miRNAs, suggesting there may be a complex network in regulation of miRNAs. Most recently, the shortening of 3’UTR by alternative cleavage and polyadenylation was found to be a widespread phenomenon and genetic regulation to activate oncogenes in human cancer cells. As a result, the shorter mRNA isoforms exhibit increased stability and typically producing more protein, through the loss of microRNA-mediated repression. In our previous study, we demonstrated that the recognized anti-apoptosis gene, XIAP, also can suppress tumor cell autophagy. Furthermore, our preliminary study revealed that there are some transcripts with short 3’UTR of XIAP in common hepatocelluar cancer cell lines, which leaded to more products of protein by short mRNA. By the reviewing the references and bioinformatics analysis, we presume that XIAP mRNA with short 3’UTR has more efficiency of translation and lead to increased functions by losing the control of certain miRNAs. At the same time, those miRNAs which are just free from 3’UTR of XIAP will bind to other target genes and lead to alteration of protein expression and functions of those genes. In current study, we will set out to verify the exact types of 3’UTR shortening of XIAP in HCC and investigate the functional and clinical significance of those short 3’UTR. We will also to reveal the functional changes and identify key factors of XIAP-related competing endogenous RNAs network by XIAP 3’UTR shortening. We hope our study would give new evidence for better understanding of miRNA-mediated tumor regulation.

近年来miRNA研究如火如荼，然而大量研究报道带来的是更加迷惑甚至矛盾的结果，其背后可能存在着竞争性调控网络和机制。新近研究发现肿瘤细胞mRNA异常的可变性剪切可以导致其3’UTR截短从而逃避miRNA的调控。前期发现，公认的凋亡抑制因子XIAP在肝细胞癌中表达上调并参与细胞自噬，其机制不明朗；同时在肝细胞癌细胞系中XIAP基因存在着3’UTR截短的mRNA，并且截短型较全长型翻译效率更高。结合生物学信息分析和文献，推测：截断3’UTR的XIAP可以逃逸相应不同miRNA调控而上调表达，同时游离miRNA可能又作用于其他基因3’UTR并引起相应蛋白表达和功能的改变。本研究拟在体内外实验中研究截断型XIAP3’UTR的功能和临床意义，并且进一步发掘3’UTR长度变化引起的局部“竞争性内源RNA”网络改变的关键因子及其功能的改变，为加深对miRNA参与肿瘤调控的认识提供新的理论基础。

X连锁凋亡抑制蛋白（XIAP）通过抑制caspase活性参与了细胞内源性凋亡的调控，同时XIAP在人类恶性肿瘤的发生发展中发挥了重要作用。目前大量研究显示微小RNA可以通过与靶基因的非编码区，即3'UTR结合来降解或转录后抑制靶基因，显示基因的3'UTR并非“垃圾区域”，也可以通过基因调控在各种生理和病理过程发挥重要作用。本研究首先发现稳定转染XIAP3'UTR的肝细胞癌细胞的XIAP和HMGA2表达显著上调，提示其可能作为一个竞争性内源RNA（ceRNA）的角色；进一步我们发现let-7a-5p均可以与 XIAP和HMGA2的3'UTR直接结合；体外实验显示过表达XIAP3'UTR可以促进SMMC-7721和HepG2细胞活力、增殖、克隆形成、侵袭和迁移能力；并且，我们证实XIAP3'UTR对肝细胞癌细胞的功能是通过let-7a-5p介导HMGA2的表达实现。因此，本研究发现XIAP3'UTR可以作为一个ceRNA吸附内源性miRNA——let-7a-5p从而解除其对靶基因HMGA2的抑制，发挥促进肝细胞癌进展的功能。另一方面，本项目发现miR-23a可以直接与XIAP的3'UTR相结合，通过抑制XIAP的表达而促进肿瘤细胞的自噬；在临床肝细胞癌样本中，我们发现miR-23a和XIAP的表达呈显著负相关，并且两者表达与肿瘤的临床病理特征密切相关，提示miR-23a/XIAP通路可能参与了肝细胞癌的发生发展。