Characterization of Arabidopsis thaliana promoter Bidirectionality and Antisense RNAs by Depletion of Nuclear RNA Decay Pathways

In animals, transcription by RNA polymerase II initiates bidirectionally from gene promoters to produce pre-mRNAs on the forward strand and promoter upstream transcripts (PROMPTs) on the reverse strand. PROMPTs are degraded by the nuclear exosome. Previous studies based on nascent RNA approaches concluded that Arabidopsis thaliana does not produce PROMPTs. Here, we used steady-state RNA sequencing in mutants defective in nuclear RNA decay, including the exosome, to reassess the existence of A. thaliana PROMPTs. While PROMPTs are overall rare in A. thaliana, ~100 cases of exosome-sensitive PROMPTs were identified. Such PROMPTs are sources of small interfering RNAs in exosome-deficient mutants, perhaps explaining why plants have evolved mechanisms to suppress PROMPTs. In addition, we found ~200 transcription start sites within 39-UTR-encoding regions that produce long unspliced, exosome-sensitive antisense RNAs. The previously characterized non-coding (nc) RNA that regulates expression of the key seed dormancy regulator, DELAY OF GERMINATION1, is a typical representative of this class of RNAs. Transcription factor genes are overrepresented among loci with exosome-sensitive antisense RNAs, suggesting a potential for widespread control of gene expression via this class of ncRNAs. Lastly, we assess the use of alternative promoters in A. thaliana and compare the accuracy of existing TSS annotations.