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Broad noncoding transcription suggests genome surveillance by RNA polymerase V
Friday, 2020/12/04 | 08:23:53

Masayuki Tsuzuki,  Shriya Sethuraman,  Adriana N. Coke,  M. Hafiz Rothi,  Alan P. Boyle, and Andrzej T. Wierzbicki

PNAS December 1, 2020 117 (48) 30799-30804

Significance

Eukaryotic genomes are pervasively transcribed, yet most transcribed sequences lack conservation or known biological functions. We show that a specialized plant-specific RNA polymerase V broadly transcribes the Arabidopsis genome. We propose a model where Pol V transcription surveils the genome and is required to recognize and repress newly inserted or reactivated transposons. Our results indicate that pervasive transcription of nonconserved sequences may serve an essential role in maintenance of genome integrity.

Abstract

Eukaryotic genomes are pervasively transcribed, yet most transcribed sequences lack conservation or known biological functions. In Arabidopsis thaliana, RNA polymerase V (Pol V) produces noncoding 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 nonconserved sequences may serve an essential role in maintenance of genome integrity.

 

See https://www.pnas.org/content/117/48/30799

 

Figure 1: Pol V transcribes more broadly than expected. (A) The IPARE method of detecting Pol V transcription. RT refers to reverse transcription, UMI refers to unique molecule identifiers. (B) Genome browser screenshot of a region transcribed by Pol V (Chromosome 1, 1878280 to 1883250). TAIR10 genome annotation, DNA methylation in CHH contexts, Pol V IPARE, and annotated Pol V transcribed regions data are shown. Data for individual biological replicates are shown in SI Appendix, Fig. S1C. (C) IPARE reads from Col-0 wild type cover a greater proportion of the genome than known features of RdDM including siRNA (28), annotated TEs (19), and CHH DMRs. Red bars indicate percentage of the genome covered by specific features. (D) HMM identifies Pol V-transcribed regions of the genome. Boxplot shows IPARE signal using combined data from three biological replicates comparing bins identified as Pol V transcribed (states 0 and 1) or non-Pol V transcribed (states 2 and 3). (E) Pol V IPARE signal depends on the enzymatic activity of Pol V. Boxplots show RPM-normalized IPARE signal levels at Pol V-transcribed and non-Pol V-transcribed regions in Col-0, nrpe1 (null allele), dms5-1 (early termination allele of NRPE1), and drd3-3 (catalytic active site point mutant of NRPE1). Asterisk indicates Wilcoxon test P < 2.2e-16. (F) Pol V IPARE signal depends on the activity of the DDR complex. Boxplots show RPM-normalized IPARE signal levels at Pol V-transcribed and non-Pol V-transcribed regions in Col-0 and nrpe1 as well as DDR subunit mutants drd1 and dms3. Asterisk indicates Wilcoxon test P < 2.2e-16.

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