Size selected NET‐Seq reveals a conserved architecture of transcription
units around yeast genes.
Abstract
Genomes from yeast to human are subject to pervasive transcription. A
single round of pervasive transcription is sufficient to alter local
chromatin conformation, nucleosome dynamics and gene expression, but
experimentally is can be hard to distinguish from background signals.
Size selected native elongating transcript sequencing (ssNET-Seq) was
developed to precisely map transcription units (TU) independent of
expression levels. RNAPII-associated nascent transcripts are
fractionation into different size ranges before library construction.
When anchored to the transcription start sites (TSS) of annotated genes,
the combined pattern of the output metagenes define the expected
reference pattern for a TU. Bioinformatic pattern matching to the
reference identified 9542 TU in Saccharomyces cerevisiae, of
which 47% are coding and 53% are non-coding. 3113 (33%) are newly
identified unannotated non-coding TU. Anchoring all TU to the TSS or
polyadenylation site (PAS) of annotated coding regions reveals
distinctive architectures of linked pairs of divergent TU approximately
200nt apart. The Reb1 transcription factor is enriched 30nt downstream
of the PAS only when an upstream (TSS-60nt) non-coding TU co-occurs with
a downstream (TSS+150nt) coding TU and supports nucleosome depletion in
the generation of the pervasive nascent transcriptome. The potential for
extensive transcriptional interference is evident from low abundance
unannotated TUs with variable TSS (median-240nt) initiating within a
500nt window upstream of, and transcribing over, the promoters of
protein coding genes. This study confirms a highly interleaved yeast
genome with different types of transcription units altering the
chromatin landscape in distinctive ways, with the potential to exert
extensive regulatory control.