Li Yanjie, Lu Yue, Polak Urszula, Lin Kevin, Shen Jianjun, Farmer Jennifer, Seyer Lauren, Bhalla Angela D, Rozwadowska Natalia, Lynch David R, Butler Jill Sergesketter, Napierala Marek
Department of Biochemistry and Molecular Genetics, UAB Stem Cell Institute, University of Alabama at Birmingham, 1825 University Blvd., Birmingham, AL 35294, USA.
Department of Epigenetics and Molecular Carcinogenesis, University of Texas MD Anderson Cancer Center, Science Park, Smithville, TX 78957, USA.
Hum Mol Genet. 2015 Dec 15;24(24):6932-43. doi: 10.1093/hmg/ddv397. Epub 2015 Sep 23.
Friedreich's ataxia (FRDA) is a severe neurodegenerative disease caused by homozygous expansion of the guanine-adenine-adenine (GAA) repeats in intron 1 of the FXN gene leading to transcriptional repression of frataxin expression. Post-translational histone modifications that typify heterochromatin are enriched in the vicinity of the repeats, whereas active chromatin marks in this region are underrepresented in FRDA samples. Yet, the immediate effect of the expanded repeats on transcription progression through FXN and their long-range effect on the surrounding genomic context are two critical questions that remain unanswered in the molecular pathogenesis of FRDA. To address these questions, we conducted next-generation RNA sequencing of a large cohort of FRDA and control primary fibroblasts. This comprehensive analysis revealed that the GAA-induced silencing effect does not influence expression of neighboring genes upstream or downstream of FXN. Furthermore, no long-range silencing effects were detected across a large portion of chromosome 9. Additionally, results of chromatin immunoprecipitation studies confirmed that histone modifications associated with repressed transcription are confined to the FXN locus. Finally, deep sequencing of FXN pre-mRNA molecules revealed a pronounced defect in the transcription elongation rate in FRDA cells when compared with controls. These results indicate that approaches aimed to reactivate frataxin expression should simultaneously address deficits in transcription initiation and elongation at the FXN locus.
弗里德赖希共济失调（FRDA）是一种严重的神经退行性疾病，由FXN基因内含子1中鸟嘌呤 - 腺嘌呤 - 腺嘌呤（GAA）重复序列的纯合扩增引起，导致frataxin表达的转录抑制。代表异染色质的翻译后组蛋白修饰在重复序列附近富集，而在FRDA样本中该区域的活性染色质标记则较少。然而，重复序列扩增对通过FXN的转录进程的直接影响及其对周围基因组环境的远程影响，是FRDA分子发病机制中两个尚未得到解答的关键问题。为了解决这些问题，我们对一大群FRDA和对照原代成纤维细胞进行了下一代RNA测序。这项全面分析表明，GAA诱导的沉默效应不影响FXN上游或下游相邻基因的表达。此外，在9号染色体的大部分区域未检测到远程沉默效应。此外，染色质免疫沉淀研究结果证实，与转录抑制相关的组蛋白修饰局限于FXN基因座。最后，FXN前体mRNA分子的深度测序显示，与对照相比，FRDA细胞中的转录延伸率存在明显缺陷。这些结果表明，旨在重新激活frataxin表达的方法应同时解决FXN基因座转录起始和延伸的缺陷。
