Article

The Set2/Rpd3S Pathway Suppresses Cryptic Transcription without Regard to Gene Length or Transcription Frequency

Curriculum in Genetics and Molecular Biology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States of America.
PLoS ONE (Impact Factor: 3.53). 02/2009; 4(3):e4886. DOI: 10.1371/journal.pone.0004886
Source: PubMed

ABSTRACT In cells lacking the histone methyltransferase Set2, initiation of RNA polymerase II transcription occurs inappropriately within the protein-coding regions of genes, rather than being restricted to the proximal promoter. It was previously reported that this "cryptic" transcription occurs preferentially in long genes, and in genes that are infrequently transcribed. Here, we mapped the transcripts produced in an S. cerevisiae strain lacking Set2, and applied rigorous statistical methods to identify sites of cryptic transcription at high resolution. We find that suppression of cryptic transcription occurs independent of gene length or transcriptional frequency. Our conclusions differ with those reported previously because we obtained a higher-resolution dataset, we accounted for the fact that gene length and transcriptional frequency are not independent variables, and we accounted for several ascertainment biases that make cryptic transcription easier to detect in long, infrequently transcribed genes. These new results and conclusions have implications for many commonly used genomic analysis approaches, and for the evolution of high-fidelity RNA polymerase II transcriptional initiation in eukaryotes.

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    • "In spt6 mutants, a genome-wide assay revealed that cryptic initiation occurs at 1000 genes (Cheung et al. 2008). This level of cryptic initiation is likely an underestimate of the true level, as this study looked only at coding strands, and the method of detection would have found cryptic initiation only in genes transcribed at low levels (Cheung et al. 2008; Lickwar et al. 2009). Cryptic initiation has also been observed in several other mutants, including spt16 and set2 (Kaplan et al. 2003; Mason and Struhl 2003; Carrozza et al. 2005; Prather et al. 2005; Nourani et al. 2006; Li et al. 2007b; Xiao et al. 2007; Cheung et al. 2008; Imbeault et al. 2008), with spt6 and spt16 mutants having the strongest effects (Cheung et al. 2008). "
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    • "the results obtained through the use of this reporter plasmid could be due to events of cryptic initiation arising from within the reporter. No evidence for cryptic initiation has been found at HIS3 in previous genomic studies (Lickwar et al. 2009). However, to be certain that cryptic initiation was not occurring in the reporter, we examined HIS3 transcripts arising from within the reporter plasmids by Northern blot analysis. "
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    • "Arabidopsis thaliana 341 emerge between the H3K4me3-marked domains around the TSS and the H3K36me2 domains in the 3¢ portion of gene bodies (Figure 2e), such that the central part of longer genes associates with neither the H3K4me3 nor the H3K36me2 marks. This H3K36me2 pattern may be adverse to the faithful transcriptions as H3K36 methylation has been reported to be essential for suppressing cryptic transcription initiation within gene bodies in other eukaryotes (Li et al., 2007b; Lickwar et al., 2009). Cryptic transcription initiation may generate antisense or aberrant transcripts that can trigger siRNA silencing and subsequently impair normal transcription . "
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