Widespread bidirectional promoters are the major source of cryptic transcripts in yeast

Abstract

Pervasive and hidden transcription is widespread in eukaryotes, but its global level, the mechanisms from which it originates and its functional significance are unclear. Cryptic unstable transcripts (CUTs) were recently described as a principal class of RNA polymerase II transcripts in Saccharomyces cerevisiae. These transcripts are targeted for degradation immediately after synthesis by the action of the Nrd1-exosome-TRAMP complexes. Although CUT degradation mechanisms have been analysed in detail, the genome-wide distribution at the nucleotide resolution and the prevalence of CUTs are unknown. Here we report the first high-resolution genomic map of CUTs in yeast, revealing a class of potentially functional CUTs and the intrinsic bidirectional nature of eukaryotic promoters. An RNA fraction highly enriched in CUTs was analysed by a 3' Long-SAGE (serial analysis of gene expression) approach adapted to deep sequencing. The resulting detailed genomic map of CUTs revealed that they derive from extremely widespread and very well defined transcription units and do not result from unspecific transcriptional noise. Moreover, the transcription of CUTs predominantly arises within nucleosome-free regions, most of which correspond to promoter regions of bona fide genes. Some of the CUTs start upstream from messenger RNAs and overlap their 5' end. Our study of glycolysis genes, as well as recent results from the literature, indicate that such concurrent transcription is potentially associated with regulatory mechanisms. Our data reveal numerous new CUTs with such a potential regulatory role. However, most of the identified CUTs corresponded to transcripts divergent from the promoter regions of genes, indicating that they represent by-products of divergent transcription occurring at many and possibly most promoters. Eukaryotic promoter regions are thus intrinsically bidirectional, a fundamental property that escaped previous analyses because in most cases divergent transcription generates short-lived unstable transcripts present at very low steady-state levels.

Divergent transcription: A new feature of active promoters

“…The average CUT length and location of the 3' SAGE CUT clusters suggests that CUT transcription predominantly originates within these regions. 1 These results suggest that a majority of CUTs may represent by-products of divergent transcription occurring at many and possibly most promoters.…”

“…The 3' SAGE tags from both libraries were highly organized in clusters and 85% of the clusters from the CUT RNA library did not correspond to annotated genes. 1 In order to determine the relationship between CUTs and annotated genes, CUT clusters were subdivided into categories based on their association and orientation with respect to annotated genes. 63% of CUTs fall between tandem genes; of these, 79% are transcribed in the opposite orientation with respect to these genes.…”

“…In contrast to 3' SAGE mRNA clusters, 3' SAGE CUT clusters are highly correlated with the start of known genes. 1 Most promoters contain a NFR, which facilitates transcription by allowing RNAPII to bind the DNA. The average CUT length and location of the 3' SAGE CUT clusters suggests that CUT transcription predominantly originates within these regions.…”

“…Results from a number of laboratories using different experimental techniques have found evidence for transcription occurring divergently from many promoters in yeast, 1,2 and mammals. [3][4][5] Two recent studies in mammals have shown that polymerase occupancy is nearly equivalent in both directions; however, one study demonstrates that the polymerase that is upstream and antisense to the direction of genes does not productively elongate.…”

Gene expression changes in normal haematopoietic cells

“…81,82 These cutting edge technologies have rapidly advanced our understanding of how genomic changes impact expression, identifying thousands of new polymorphisms and promoter loci. 78,83,84 Currently, there are a number of different techniques to examine DNA methylation and histone marks -two major epigenetic mechanisms for controlling transcription. DNA methylation changes were first examined using restriction enzyme digest/PCR assays.…”