Identification of Multiple Proteins Coupling Transcriptional Regulation to Genome Stability in Arabidopsis thaliana

Eukaryotic genomes are heavily regulated by epigenetic marks that often act to modulate the transcriptional control of genetic elements. In Arabidopsis thaliana the ATXR5 and ATXR6 histone methyltransferases, and their cognate H3K27 monomethylation mark, act in transcriptional silencing while also maintaining genome stability by preventing generation of excess DNA corresponding to pericentromeric heterochromatin. In this study we characterize the atxr5 atxr6 transcriptome and its relationship to the DNA damage response which suggests that the atxr5 atxr6 transcriptional defects may be epistatic to the genome instability defects in the mutants. In addition we isolate several factors that modulate both the transcriptional and genomic instability phenotypes of atxr5 atxr6 mutants, which suggest a mechanism for atxr5 atxr6-induced extra DNA involving conflicts between the replicative and transcriptional processes in the cell. PolyA RNA sequencing (RNA-seq), whole-genome resequencing (DNA-seq), and whole-genome bisulfite sequencing (methyl-seq) was performed on Arabidospsis thaliana mutant and wildtype plants. DNA-seq was used to characterize DNA copy number and map EMS-induced mutations, RNA-seq was used to quantify transcript abundance and map EMS-induced mutations, and methyl-seq was used to assess DNA methylation. Details of the relationship between samples in this series and figures in the associated manuscript can be found in Supplemental Table 4 of the associated manuscript. Unless otherwise noted in the description all lines are ecotype Columbia, and all genotypes should be assumed homozygous unless otherwise indicated with a '/'.

真核基因组广泛受表观遗传标记调控，此类标记往往参与调控遗传元件的转录控制。在拟南芥（Arabidopsis thaliana）中，ATXR5与ATXR6组蛋白甲基转移酶（histone methyltransferases）及其同源的H3K27单甲基化标记，既可介导转录沉默，又可通过阻止着丝粒异染色质区域产生过量DNA，维持基因组稳定性。本研究对atxr5 atxr6双突变体的转录组及其与DNA损伤应答的关联进行了表征，结果提示atxr5 atxr6突变体的转录缺陷可能与基因组不稳定缺陷存在上位性关系。此外，本研究筛选获得数个可调控atxr5 atxr6突变体转录与基因组不稳定表型的因子，这为atxr5 atxr6诱导的过量DNA产生机制提供了线索——该机制可能涉及细胞内DNA复制与转录过程的冲突。本研究对拟南芥突变体与野生型植株开展了PolyA RNA测序（PolyA RNA sequencing, RNA-seq）、全基因组重测序（whole-genome resequencing, DNA-seq）以及全基因组亚硫酸氢盐测序（whole-genome bisulfite sequencing, methyl-seq）。其中，全基因组重测序（DNA-seq）用于表征DNA拷贝数变异并定位乙基甲磺酸（Ethyl Methanesulfonate, EMS）诱导的突变；PolyA RNA测序（RNA-seq）用于定量转录本丰度并定位EMS诱导的突变；全基因组亚硫酸氢盐测序（methyl-seq）则用于检测DNA甲基化水平。本数据集系列中样本与关联手稿中图件的关联细节，可查阅关联手稿的补充表4。除非本描述中另有说明，所有株系均为哥伦比亚生态型（Columbia ecotype）；若基因型未以'/'标注，则默认该基因型为纯合状态。