Broad non-coding transcription suggests genome surveillance by RNA Polymerase V

Eukaryotic genomes are pervasively transcribed, yet most transcribed sequences lack conservation or known biological functions. In Arabidopsis thaliana RNA polymerase V (Pol V) produces non-coding transcripts, which base-pair with small interfering RNA (siRNA) and allow specific establishment of RNA-directed DNA methylation (RdDM) on transposable elements. Here, we show that Pol V transcribes much more broadly than previously expected, including subsets of both heterochromatic and euchromatic regions. At already established RdDM targets Pol V and siRNA work together to maintain silencing. In contrast, some euchromatic sequences do not give rise to siRNA but are covered by low levels of Pol V transcription, which is needed to establish RdDM de novo if a transposon is reactivated. We propose a model where Pol V surveils the genome to make it competent to silence newly activated or integrated transposons. This indicates that pervasive transcription of non-conserved sequences may serve an essential role in maintenance of genome integrity. Immunoprecipitation followed by Analysis of RNA Ends (IPARE) with an anti-NRPE1 antibody was performed on 24 samples, which include 11 different genotypes (Col-0:6replicates; nrpe1:5replicates; spt5l,ago4,dms5-1,drd3-3: 2replicates each, drd1,dms3,suvh2, suvh9, suvh2/suvh9: 1replicate each)

真核基因组存在广泛的转录现象，但绝大多数转录序列既无保守性，亦未发现明确生物学功能。在拟南芥（Arabidopsis thaliana）中，RNA聚合酶V（Pol V）可生成非编码转录本，这类转录本可与小干扰RNA（siRNA）发生碱基配对，从而介导转座元件上RNA指导的DNA甲基化（RdDM）的特异性建立。本研究发现，Pol V的转录范围远超前述预期，涵盖异染色质与常染色质区域的多个亚组。在已建立RdDM的靶位点上，Pol V与siRNA协同维持基因沉默。与之相对，部分常染色质序列无法产生siRNA，但存在低水平的Pol V转录；若转座元件被重新激活，该类转录是从头建立RdDM的必要条件。我们提出如下模型：Pol V可对基因组进行监视，使基因组具备沉默新激活或新整合转座元件的能力。这表明非保守序列的广泛转录可能在维持基因组完整性中发挥关键作用。研究团队针对24份样本开展了基于抗NRPE1抗体的免疫沉淀结合RNA末端分析（Immunoprecipitation followed by Analysis of RNA Ends, IPARE）实验，样本涵盖11种不同基因型（Col-0：6个生物学重复；nrpe1：5个生物学重复；spt5l、ago4、dms5-1、drd3-3：各2个生物学重复；drd1、dms3、suvh2、suvh9、suvh2/suvh9：各1个生物学重复）。