Project description:THO/TREX is a conserved nuclear complex that functions in mRNP biogenesis and prevents transcription-associated recombination. Whether or not it has a ubiquitous role in the genome is an open question. ChIP-chip studies reveal that the Hpr1 component of THO and the Sub2 RNA-dependent ATPase have genome wide-distributions at active ORFs in yeast. In contrast to RNAPII, evenly distributed from promoter to termination regions, THO and Sub2 are absent at promoters and distributed in a sharp 5’→3’ gradient. Importantly, ChIP-chips reveal an over-recruitment of Rrm3 in active genes in THO mutants that is reduced by overexpression of RNase H1. Our work establishes a genome-wide function for THO-Sub2 in transcription elongation and mRNP biogenesis that function to prevent the accumulation of transcription-mediated replication obstacles, including R-loops.
Project description:THO/TREX is a conserved nuclear complex that functions in mRNP biogenesis and prevents transcription-associated recombination. Whether or not it has a ubiquitous role in the genome is an open question. ChIP-chip studies reveal that the Hpr1 component of THO and the Sub2 RNA-dependent ATPase have genome wide-distributions at active ORFs in yeast. In contrast to RNAPII, evenly distributed from promoter to termination regions, THO and Sub2 are absent at promoters and distributed in a sharp 5M-bM-^@M-^YM-bM-^FM-^R3M-bM-^@M-^Y gradient. Importantly, ChIP-chips reveal an over-recruitment of Rrm3 in active genes in THO mutants that is reduced by overexpression of RNase H1. Our work establishes a genome-wide function for THO-Sub2 in transcription elongation and mRNP biogenesis that function to prevent the accumulation of transcription-mediated replication obstacles, including R-loops. ChIP-chip studies were perfomed with tagged forms of the Hpr1 component of THO (Hpr1-FLAG), the Sub2 RNA-dependent ATPase of TREX (Sub2-FLAG), the Rpb3 subunit of RNA polymerase II (Rpb3-PK) and the Rrm3 protein (Rrm3-FLAG) in the yeast S. cerevisiae.
Project description:Crosslinked DNA from wild type cells or from mutant of the THO complex was analyzed by tiling arrays to precisely detect DNA targets of THO in yeast.
Project description:THO/TREX is a conserved nuclear complex that functions in mRNP biogenesis at the interface of transcription-RNA export with a key role in preventing transcription-associated genome instability. We used microarrays to analyze the impact of different THO/TREX mutations on gene expression and found that THO-Sub2 deletions have a high functional impact on highly expressed, long and G+C-rich genes regardless of gene function.
Project description:RNA helicases constitute a large protein family implicated in cellular RNA homeostasis and disease development. Here we show that the RNA helicase Ighmbp2, linked to the neuromuscular disorder SMARD1 (DSMA1) associates with polysomes and impacts on translation of cellular mRNAs containing short, GC-rich and highly structured 5’UTRs. Absence of Ighmbp2 causes ribosome stalling at the start codon of target mRNAs, leading to their reduced translation efficiency. The main mRNA targets of Ighmbp2-mediated regulation encode for components of the THO complex that links mRNA production and nuclear export. Accordingly, failure of Ighmbp2 regulation of the THO complex causes perturbations of the cellular transcriptome and its encoded proteome. Ablation of essential THO complex subunits phenocopies these perturbations. Thus, Ighmbp2 is an upstream regulator of the THO complex that impacts on cellular mRNA homeostasis. Of note, Ighmbp2 dependent regulation of the THO complex is also observed in astrocytes derived from DSMA1 patients, suggesting that de-regulated mRNA metabolism contributes to SMARD1 etiology.
Project description:THO/TREX is a conserved nuclear complex that functions in mRNP biogenesis at the interface of transcription-RNA export with a key role in preventing transcription-associated genome instability. We used microarrays to analyze the impact of different THO/TREX mutations on gene expression and found that THO-Sub2 deletions have a high functional impact on highly expressed, long and G+C-rich genes regardless of gene function. S. cerevisiae strains were grown in YPAD liquid culture, total RNA was isolated and hybridized on Affymetrix microarrays.