Powers et al. 2022 (PRJNA854601)

General Details

Title loss of DBP1 during meiosis in yeast
Organism
Number of Samples 4
Release Date 2022/06/30 00:00
Sequencing Types
Protocol Details

Study Links

Repository Details

SRA SRP384366
ENA SRP384366
GEO GSE207267
BioProject PRJNA854601

Publication

Title
Authors Powers EN, Chan C, Doron-Mandel E, Llacsahuanga Allcca L, Kim Kim J, Jovanovic M, Brar GA
Journal eLife
Publication Date 2022 Dec 12
Abstract Targeted selection-based genome-editing approaches have enabled many fundamental discoveries and are used routinely with high precision. We found, however, that replacement of DBP1 with a common selection cassette in budding yeast led to reduced expression and function for the adjacent gene, MRP51 , despite all MRP51 coding and regulatory sequences remaining intact. Cassette-induced repression of MRP51 drove all mutant phenotypes detected in cells deleted for DBP1 . This behavior resembled the 'neighboring gene effect' (NGE), a phenomenon of unknown mechanism whereby cassette insertion at one locus reduces the expression of a neighboring gene. Here, we leveraged strong off-target mutant phenotypes resulting from cassette replacement of DBP1 to provide mechanistic insight into the NGE. We found that the inherent bidirectionality of promoters, including those in expression cassettes, drives a divergent transcript that represses MRP51 through combined transcriptional interference and translational repression mediated by production of a long undecoded transcript isoform (LUTI). Divergent transcript production driving this off-target effect is general to yeast expression cassettes and occurs ubiquitously with insertion. Despite this, off-target effects are often naturally prevented by local sequence features, such as those that terminate divergent transcripts between the site of cassette insertion and the neighboring gene. Thus, cassette-induced off-target effects can be eliminated by the insertion of transcription terminator sequences into the cassette, flanking the promoter. Because the driving features of this off-target effect are broadly conserved, our study suggests it should be considered in the design and interpretation of experiments using integrated expression cassettes in other eukaryotic systems, including human cells. © 2022, Powers et al.
PMC PMC9754628
PMID 36503721
DOI
Run Accession Study Accession Scientific Name Cell Line Library Type Treatment GWIPS-viz Trips-Viz Reads BAM BigWig (F) BigWig (R)
SRR19908120 PRJNA854601 Saccharomyces cerevisiae NA_SK1 Ribo-Seq 0.0
SRR19908119 PRJNA854601 Saccharomyces cerevisiae NA_SK1 Ribo-Seq 0.0
SRR19908118 PRJNA854601 Saccharomyces cerevisiae NA_SK1 Ribo-Seq 0.0
SRR19908117 PRJNA854601 Saccharomyces cerevisiae NA_SK1 Ribo-Seq 0.0
Run Accession Study Accession Scientific Name Cell Line Library Type Treatment GWIPS-viz Trips-Viz Reads BAM BigWig (F) BigWig (R)

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