Sloane Mathew A, Hesson Luke B, Nunez Andrea C, Thompson Bryony A, Ward Robyn L
Adult Cancer Program, Lowy Cancer Research Centre and Prince of Wales Clinical School, University of New South Wales, Sydney, NSW Australia.
Department of Genetics and Computational Biology, QIMR Berghofer Medical Research Institute, Brisbane, QLD Australia.
Clin Epigenetics. 2014 Dec 13;6(1):32. doi: 10.1186/s13148-014-0032-6. eCollection 2014.
Splicing is more efficient when coupled with transcription and it has been proposed that nucleosomes enriched in exons are important for splice site recognition. Lynch syndrome is a familial cancer syndrome that can be caused by the autosomal dominant inheritance of splice site mutations in the MutL homolog 1 (MLH1) gene. To better understand the role of nucleosomes in splicing, we used MLH1 splice site mutations in Lynch syndrome cases as a model to investigate if abnormal splicing was associated with altered nucleosome positioning at exon-intron boundaries.
Nucleosome Occupancy and Methylome sequencing (NOMe-seq) was used to determine the allele-specific positioning of nucleosomes around heterozygous splice site mutations in lymphoblastoid cells lines (LCLs) derived from six Lynch syndrome patients. These mutations were previously shown to cause exon skipping in five of the six patients. Allele-specific high-resolution nucleosome mapping across exons and exon-intron boundaries revealed high levels of nucleosomes across all regions examined. Alleles containing donor or acceptor splice site mutations showed no consistent alteration in nucleosome positioning or occupancy.
Nucleosomes were enriched at MLH1 exons in LCLs derived from Lynch syndrome patients, and in this model system the positioning of nucleosomes was unaltered at exon-intron boundaries containing splice site mutations. Thus, these splice site mutations alone do not significantly change the local organisation of nucleosomes.
剪接与转录偶联时效率更高，并且有人提出富含外显子的核小体对剪接位点识别很重要。林奇综合征是一种家族性癌症综合征，可由错配修复蛋白同源物1（MLH1）基因剪接位点突变的常染色体显性遗传引起。为了更好地理解核小体在剪接中的作用，我们以林奇综合征病例中的MLH1剪接位点突变为模型，研究异常剪接是否与外显子-内含子边界处核小体定位的改变有关。
使用核小体占据和甲基化组测序（NOMe-seq）来确定来自6名林奇综合征患者的淋巴母细胞系（LCL）中杂合剪接位点突变周围核小体的等位基因特异性定位。这些突变先前已被证明在6名患者中的5名中导致外显子跳跃。跨外显子和外显子-内含子边界的等位基因特异性高分辨率核小体图谱显示，在所有检测区域中核小体水平都很高。含有供体或受体剪接位点突变的等位基因在核小体定位或占据方面没有一致的改变。
在来自林奇综合征患者的LCL中，核小体在MLH1外显子处富集，并且在该模型系统中，在含有剪接位点突变的外显子-内含子边界处核小体的定位未改变。因此，仅这些剪接位点突变不会显著改变核小体的局部组织。
