非编码RNA HULC促进细胞增殖、迁移及侵袭的生物学作用及分子机制研究
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摘要
非编码RNA,即non-coding RNA(ncRNA),是指那些由DNA转录却并不翻译成蛋白的RNA,它们包含高密度的终止子或缺少有效的开放阅读框。它们能够通过不同的机制,如RNA-RNA碱基配对,RNA蛋白相互作用,RNA-DNA相互作用等,在细胞调控的各个层面中发挥功能,包括参与染色质修饰、转录调节、剪切加工、RNA稳定性调节等。近来还有研究发现,在一些肿瘤组织中,ncRNA有着过表达或者低表达的现象。提示在肿瘤的发生发展过程中,它们可能发挥着重要的作用。
HULC (Highly Up-reglated in Liver Cancer)是一种在肝癌中发现的ncRNA。该基因定位于6p24.3,HULC基因转录产物经过剪切和加工后形成一个500bp的RNA,具有类似于mRNA的polyA尾结构。但这个RNA序列中存在多个终止密码子,并不能编码蛋白。HULC RNA不仅能够在肝癌组织中检测到,还能够在肝癌病人的血液中检测到高表达,另外有研究表明,除了能在肝癌组织中检测到HULC RNA高表达外,在从结肠癌转移到肝脏的转移瘤中也能检测到HULC RNA。这提示我们它可能可以作为一种潜在的肝癌标志物应用到临床诊断中。但是,HULC RNA的功能及分子机制迄今仍未知。
在本研究中,我们发现HULC在内源性高表达的细胞系中并不存在基因组扩增,且HULC RNA半衰期在约6小时,属于较长效的RNA。并且HULC RNA能够促进细胞的体外增殖、迁移及侵袭,并能抑制顺铂诱导的凋亡。另外,我们还筛选出与HULC RNA特异性结合的蛋白YB-1。它是一种多功能蛋白,可以分别和DNA、RNA及其他蛋白相结合,参与转录调节、翻译调控、mRNA选择性剪接、mRNA稳定性、DNA的修复、细胞增殖等。我们的研究还发现在受到DNA损伤刺激后,HULC RNA能够影响YB-1蛋白的核定位,并影响YB-1蛋白对部分下游基因的调控。
Non-coding RNAs (ncRNAs), which transcript from DNA but do not encode protein, contain high density of stop codons or lack extensive "Open Reading Frame". They have been shown to be involved in the different levels of cell regulations including chromosome modification, transcription regulation, RNA splicing and RNA stability, via RNA-RNA basepair, RNA-protein interaction, RNA-DNA interaction. Recently, dysregulated expressions of ncRNAs have been reported in many types of cancers, indicating that they maybe play critical roles in tumorigenesis.
HULC (Highly Up-regulated in Liver Cancer), is an ncRNA characterized in hepatocelluar carcinoma (HCC). The HULC gene locates at 6p24.3 and the transcripts of this gene is spliced and polyadenylated like mRNA to form a 500bp RNA. Feature of the HULC RNA sequence is the high density of stop codons in the small potential reading frames and can not encode a protein product. HULC RNA can be detected in HCC tissues as well as in the blood of HCC patients at a high level. It has also been reported that HULC expression is not confined to HCC, but also to those colorectal carcinomas that metastasize to the liver. HULC RNA can therefore be used as specific molecular markers for HCC. However, little is known about the function of HULC RNA and its molecular mechanism.
In the present thesis we firstly show that endogenously high expression of HULC RNA in HepG2 cell is not caused by gene amplification and the half-life of HULC RNA is approximately 6 hours. Meanwhile, HULC RNA can promote cell proliferation, migration and invasion in vitro and also inhibit cisplatin-induced apoptosis. Moreover, we have shown for the first time that HULC RNA can specifically bind to YB-1 protein both in vitro and in vivo. YB1 protein appears to carry out multiple functions, including the roles in transcriptional and translational regulation, RNA splicing and masking, DNA repair, and cell proliferation. Our study further demonstrates that HULC RNA can promote nuclear localization of YB-1 protein after DNA damage stress, and is involved in the transcriptional regulation of its target genes.
HULC (Highly Up-reglated in Liver Cancer)是一种在肝癌中发现的ncRNA。该基因定位于6p24.3,HULC基因转录产物经过剪切和加工后形成一个500bp的RNA,具有类似于mRNA的polyA尾结构。但这个RNA序列中存在多个终止密码子,并不能编码蛋白。HULC RNA不仅能够在肝癌组织中检测到,还能够在肝癌病人的血液中检测到高表达,另外有研究表明,除了能在肝癌组织中检测到HULC RNA高表达外,在从结肠癌转移到肝脏的转移瘤中也能检测到HULC RNA。这提示我们它可能可以作为一种潜在的肝癌标志物应用到临床诊断中。但是,HULC RNA的功能及分子机制迄今仍未知。
在本研究中,我们发现HULC在内源性高表达的细胞系中并不存在基因组扩增,且HULC RNA半衰期在约6小时,属于较长效的RNA。并且HULC RNA能够促进细胞的体外增殖、迁移及侵袭,并能抑制顺铂诱导的凋亡。另外,我们还筛选出与HULC RNA特异性结合的蛋白YB-1。它是一种多功能蛋白,可以分别和DNA、RNA及其他蛋白相结合,参与转录调节、翻译调控、mRNA选择性剪接、mRNA稳定性、DNA的修复、细胞增殖等。我们的研究还发现在受到DNA损伤刺激后,HULC RNA能够影响YB-1蛋白的核定位,并影响YB-1蛋白对部分下游基因的调控。
Non-coding RNAs (ncRNAs), which transcript from DNA but do not encode protein, contain high density of stop codons or lack extensive "Open Reading Frame". They have been shown to be involved in the different levels of cell regulations including chromosome modification, transcription regulation, RNA splicing and RNA stability, via RNA-RNA basepair, RNA-protein interaction, RNA-DNA interaction. Recently, dysregulated expressions of ncRNAs have been reported in many types of cancers, indicating that they maybe play critical roles in tumorigenesis.
HULC (Highly Up-regulated in Liver Cancer), is an ncRNA characterized in hepatocelluar carcinoma (HCC). The HULC gene locates at 6p24.3 and the transcripts of this gene is spliced and polyadenylated like mRNA to form a 500bp RNA. Feature of the HULC RNA sequence is the high density of stop codons in the small potential reading frames and can not encode a protein product. HULC RNA can be detected in HCC tissues as well as in the blood of HCC patients at a high level. It has also been reported that HULC expression is not confined to HCC, but also to those colorectal carcinomas that metastasize to the liver. HULC RNA can therefore be used as specific molecular markers for HCC. However, little is known about the function of HULC RNA and its molecular mechanism.
In the present thesis we firstly show that endogenously high expression of HULC RNA in HepG2 cell is not caused by gene amplification and the half-life of HULC RNA is approximately 6 hours. Meanwhile, HULC RNA can promote cell proliferation, migration and invasion in vitro and also inhibit cisplatin-induced apoptosis. Moreover, we have shown for the first time that HULC RNA can specifically bind to YB-1 protein both in vitro and in vivo. YB1 protein appears to carry out multiple functions, including the roles in transcriptional and translational regulation, RNA splicing and masking, DNA repair, and cell proliferation. Our study further demonstrates that HULC RNA can promote nuclear localization of YB-1 protein after DNA damage stress, and is involved in the transcriptional regulation of its target genes.
引文
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