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Accession IconSRP092767

Bidirectional terminators in Saccharomyces cerevisiae prevent cryptic transcription from invading neighbouring genes

Organism Icon Saccharomyces cerevisiae
Sample Icon 12 Downloadable Samples
Technology Badge IconIllumina HiSeq 2000

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Transcription can be quite disruptive for chromatin so cells have evolved mechanisms to preserve chromatin integrity during transcription, hence preventing the emergence of cryptic transcript from spurious promoter sequences. How these transcripts are regulated and processed by cells remains poorly characterized. Notably, very little is known about the termination of cryptic transcription. Here we used RNA-Seq to identify and characterize cryptic transcripts in Spt6 mutant cells (spt6-1004) in Saccharomyces cerevisiae. We found polyadenylated cryptic transcripts running both sense and anti-sense relative to genes in this mutant. Cryptic promoters were enriched for TATA boxes, suggesting that the underlying DNA sequence defines the location of cryptic promoters. While intragenic sense cryptic transcripts terminate at the terminator of the genes that host them, we found that anti-sense cryptic transcripts preferentially terminate at the 3’-end of upstream genes. These findings led us to demonstrate that most terminators in yeast are bidirectional, leading to termination and polyadenylation of transcripts coming from either direction. We propose that S. cerevisiae has evolved this mechanism in order to prevent spurious transcription from invading neighbouring genes, a feature particularly critical for organisms with small compact genomes. Overall design: Cells from spt16-1004 and its respective WT strain were grown to an OD600 of 0.5 at 30°C and shifted to 37°C for 80 min before RNA extraction. Total RNA was extracted using the hot phenol method. Prior to library preparation, total RNA was either depleted for ribosomal RNA using the Ribo-zero Gold yeast kit (Epicentre-Illumina) or enriched for polyadenylated RNA using the NEBnext Poly(A) kit (New England Biolabs). Strand specific RNA-seq libraries were prepared using the KAPA stranded RNA-Seq library preparation kit prior to paired-end sequencing on an Illumina Hi-Seq2000. Reads were mapped to the sacCer3 assembly of the S. cerevisiae genome using Tophat2 (23). Intron length range was set at 50 to 1000 bp and a reference annotation file was provided to guide the assembly.
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