B细胞淋巴瘤内miR-17~92基因转录调控机制研究

During transcription of genes, histone modification plays an important role in regulation of miRNAs. A cluster of six microRNAs, miR-17~92, is processed from the transcript of C13orf25, a gene amplified in some lymphomas and solid tumors. Overexpression of miR-17~92 accelerates lymphomagenesis in mouse models. The microRNAs in the cluster target transcripts of genes, such as E2F1, PTEN, and BIM, which are important in cell proliferation and apoptosis. Recent studies have shown that C13orf25 is activated by MYC and E2F transcription factors; however, detailed examination of its transcriptional regulation has not been reported. We found that levels of the miRNAs in the cluster do not vary entirely in parallel with each other or with the primary RNA in B-cell lines or normal cells, suggesting that processing or stability of the miRNAs is differentially regulated. We also found that the inhibitory MYC family member MXI1 bound to the promoter and the mutation of putative SP1 sites reduced activity of promoter reporter constructs by 70%, suggesting that the SP/KLF family of transcription factors may be important in regulating this gene. Initially, SP1 was thought to have no role other than to constitutively activate housekeeping genes, but evidence has mounted for important roles in cell proliferation and tumorigenesis. Additional specificity is likely provided by its direct interaction with numerous other transcription factors, including several that may affect C13orf25 expression, including E2F1, OCT1, NFYA, and NFYC. Interactions with E2F2 and E2F3, ETS1, and MYC have also been identified, but these may be indirect. Thus, the SP1 sites may be important in promoting cooperative recruitment of SP1 along with other transcription factors that bind the promoter. The objective of current proposal is to investigate the role of SP/KLF family of transcription factors in the transcription of this gene and to determine the functional role of MYC family member in the transcription of this gene in lymphoma. The proposed study will be the first transcriptional mechanism study of the role of miR-17~92 in lymphoma. The study will not only define the role of SP/KLF family of transcription factors in transcriptional regulation of the miR-17~92 cluster, but also identify the inhibitory MYC family member as the potential targets for novel therapies. The results of these studies will not only advance our understanding of the role of oncogenic miRNAs in lymphoma development and progression, but also promise to provide proof-of-principle data for targeting miRNAs in lymphoma therapies.

在基因的转录过程中，组蛋白修饰的调控是重要的分子水平调控因素之一。研究表明，组蛋白修饰对miRNAs的调控起着重要作用。人类miR-17~92基因簇位于C13orf25基因的第三个内含子的中，它通常过度表达于不同类型的B细胞淋巴瘤中。如今很多研究注重于miR-17~92基因簇的功能，但是对于其转录调控的原理几乎未知。我们初步证实，Myc家族蛋白在调控miR-17~92基因转录的作用，证实了转录因子Mxi1直接结合在启动子区域。对启动子转录因子SP1的潜在结合点进行突变后，启动子活性减少了70%。此课题拟通过实验进一步证实，Sp1-like/KLF转录调控蛋白家族对miR-17~92基因簇的转录所起的重要作用，进一步研究MYC蛋白家族和组蛋白修饰调控在淋巴瘤中，对miR-17~92基因簇转录调控的机制。同时在机制研究的基础上，为寻找新的淋巴瘤生物治疗手段提供依据。

人类miR-17~92基因簇位于C13orf25基因的第三个内含子的中，它通常过度表达于不同类型的B细胞淋巴瘤中，包括Burkitt淋巴瘤，弥漫性大壁淋巴瘤，滤泡性淋巴瘤和套细胞淋巴瘤。已知miR-17~92基因可以被转录因子MYC和E2F激活，但是对于其进一步的转录调控的原理几乎未知。我们通过实验证实，Myc家族蛋白在调控miR-17~92基因转录的作用，证实了转录因子Mxi1直接结合在启动子区域。对启动子转录因子SP1的潜在结合点进行突变后，启动子活性减少了70%。发现Sp1-like/KLF转录调控蛋白家族对miR-17~92基因簇的转录起了重要调控作用，进一步研究了MYC蛋白家族和组蛋白修饰调控在淋巴瘤中对miR-17~92基因簇转录调控的机制。本课题是世界上首次以B细胞淋巴瘤为研究模型对癌基因miR-17~92基因簇的转录调机制进行的研究, 本课题不仅进一步加深理解了肿瘤微小RNA在淋巴瘤发生中的机制更为其作为淋巴瘤靶向治疗提供了理论支持。