Defective XRN3‐mediated transcription termination in Arabidopsis affects the expression of protein‐coding genes

Abstract

Arabidopsis thaliana contains two nuclear XRN2/3 5'-3' exonucleases that are homologs of yeast and human Rat1/Xrn2 proteins involved in the processing and degradation of several classes of nuclear RNAs and in transcription termination of RNA polymerase II. Using strand-specific short read sequencing we show that knockdown of XRN3 leads to an altered expression of hundreds of genes and the accumulation of uncapped and polyadenylated read-through transcripts generated by inefficiently terminated Pol II. Our data support the notion that XRN3-mediated changes in the expression of a subset of genes are caused by upstream read-through transcription and these effects are enhanced by RNA-mRNA chimeras generated in xrn3 plants. In turn, read-through transcripts that are antisense to downstream genes may trigger production of siRNA. Our results highlight the importance of XRN3 exoribonuclease in Pol II transcription termination in plants and show that disturbance in this process may significantly alter gene expression.

Characterization of Arabidopsis thaliana promoter bidirectionality and antisense RNAs by depletion of nuclear RNA decay enzymes

“…These pathways may involve the nuclear 5'-3' exonucleases XRN2 and XRN3. XRN2 has documented functions in ribosomal RNA cleavage (Zakrzewska-Placzek et al, 2010) , while XRN3 has more identified nuclear substrates, including RNA polymerase II-associated RNA and cleavage products by DICER-LIKE enzymes, the former giving rise to defects in transcription termination in xrn3 mutants (Krzyszton et al, 2018) . In addition, the heptameric nuclear Lsm complex, Lsm2-8, a well-established pre-mRNA splicing factor (Bouveret et al, 2000;Tharun et al, 2000) , also plays roles in nuclear RNA decapping and, ultimately, decay, presumably via 5'-3' exonucleolysis (Golisz et al, 2013) .…”

Transcriptional read-through of the long non-coding RNA SVALKA governs plant cold acclimation

“…Detection in nuclear exosome mutants and nascent transcript preparation gives asCBF1 characteristics of a cryptic lncRNA that is quickly cleaved and degraded in wild-type. The XRN3 is thought to represent the 5'-to-3' torpedo exonuclease mediating transcriptional termination in Arabidopsis 30,31 . However, we did not observe the asCBF1 RNA or CBF1 mis-regulation in the xrn3-3 mutant, suggesting that this effect is XRN3 independent (Supplementary Fig.…”

“…Nascent transcript cleavage creates the 3'-end of the full length mRNA substrate for poly-adenylation, and a free 5'-end is attacked by 5' to 3' exonucleases eliciting transcription termination according to the torpedo model 29,30 . To detect any read-through transcription from SVK, we purified nascent RNA associating with RNAPII complexes (Supplementary Fig.…”

Secondary sulfur metabolism in cellular signalling and oxidative stress responses

“…Inhibition of XRNs by PAP (Dichtl et al, 1997), leads to the activation of 25 % of the high light stress transcriptome, including genes encoding antioxidant enzymes such as Ascorbate Peroxidase 2 (APX2) (Estavillo et al, 2011). Recent work by Crisp et al (2018) and Krzyszton et al (2018) provided valuable insights into the paradox of how specificity in the regulation of nuclear gene expression by PAP is achieved, given that the XRNs are not transcription factors which bind to specific DNA motifs. PAP-mediated inhibition of XRNs leads to inefficient removal of Pol II upon completion of transcription a particular gene, thereby facilitating polymerase read-through to the downstream gene and increasing transcription of this downstream gene.…”