Functional analysis epigenetics regulators in carcinogenesis

人们一直致力于了解基因突变所致恶性转化的机制。如今，对癌症表观遗传突变的研究也是必要的。在这个项目中，我们专注于表观遗传学调节因子(组蛋白修饰酶和组蛋白变体)，它们在化学诱导的肝癌发生过程中上调。组蛋白修饰酶过表达在肝癌发生发展中的作用我们通过RT-PCR、DNA芯片和Western印迹分析鉴定了在肝癌发生过程中诱导的组蛋白修饰酶。其中，组蛋白乙酰转移酶调节ras癌基因突变所介导的转化活性。组蛋白脱乙酰酶是另一种调节剂，它能促进软琼脂的锚定非依赖性生长。组蛋白修饰酶过表达对肿瘤标志物基因表达的影响。谷胱甘肽转移酶胎盘型在…过程中被显著诱导更易发生肝癌，被公认为可靠的肿瘤标志物。转录因子Nrf2/MafK异源二聚体是肝癌发生过程中对GST-P表达最重要的正向调节因子之一。我们发现，在包括增生性结节在内的肝脏核提取液中过表达的组蛋白乙酰转移酶之一，作为Nrf2/MafK异源二聚体的辅助因子，诱导GST-p的表达。酵母中组蛋白变异体的功能分析：我们发现组蛋白变异体中的一种组蛋白变异体H_2A.Z在肝癌发生过程中被诱导。为了深入了解H_2A·Z的分子功能，我们在酿酒酵母中获得了H_2A·Z缺失菌株。H_2A.Z缺失菌株对遗传毒性试剂敏感。此外，我们观察了镍化合物的敏感性，镍化合物降低了四种组蛋白的乙酰化水平，增加了H_2A和H_2B的泛素化以及H_3赖氨酸9的二甲基化，发现镍毒性是通过染色质重塑因子介导的H_2AZ掺入染色质而出现的，而不是H_2AZ的乙酰化。较少

Great effort has been directed towards understanding the mechanism of malignant transformation derived from genetic mutations. Nowadays, studies on epigenetic mutations in cancer are also required. In this project, we focused on epigenetics modulators (histone modification enzymes and histone variants), up-regulated during chemically induced hepatocarcinogenesis.1. Effect of over expression of histone modification enzymes on the malignant transformation.We identified histone modification enzymes, which induced during hepatocarcinogenesis by RT-PCR, DNA microarray, and Western blot analyses. One of them, histone acetyltransferase, regulated the transformation activity mediated by mutated ras oncogene. Histone deacetylase, one of other modulators, promoted anchorage independent growth in soft agarose.2. Effect of over expression of histone modification enzymes on expression of the tumor marker gene.Glutathione transferase placental form (GST-P) is specifically dramatically induced during … More hepatocarcinogenesis and recognized as a reliable tumor marker. Transcriptional factor Nrf2/MafK heterodimer is one of the most important positive regulators on GST-P expression during hepatocarcinogenesis. We revealed that one of histone acetyltransferase, which was over-expressed in nuclear extracts from livers including hyperplastic nodules, acted as a cofactor of Nrf2/MafK heterodimer and induced expression of GST-P.3. Functional analysis of the histone variant in yeast.We found that H2A.Z, one of histone variants, was induced during hepatocarcinogenesis. Tb gain insights of molecular function of H2A.Z, we generated a H2A.Z deletion strain in Saccharomyces cerevisiae. The H2A.Z deletion strain was sensitive to genotoxic reagents. Further, we observed the sensitivity to nickel compounds, which decrease acetylation levels of all four histones and increase ubiquitylation of H2A and H2B and dimethylation of H3 lysine 9, and revealed that the nickel toxicity appeared with the incorporation of H2AZ into chromatin mediated by the chromatin remodeling factor, but not acetylation of H2AZ. Less