1 Department of Molecular Biology, Faculty of Science, Aarhus University, Aarhus University2 Department of Molecular Biology and Genetics - Gene Expression and Gene Medicine, Department of Molecular Biology and Genetics, Science and Technology, Aarhus University3 Department of Molecular Biology and Genetics - Gene Expression and Gene Medicine, Department of Molecular Biology and Genetics, Science and Technology, Aarhus University
Processing of mRNA and proper formation of messenger ribonucleoprotein particles (mRNPs) require co-transcriptional loading of proteins onto nascent transcripts, which is critically dependent on the function of the THO/TREX complex, as well as on proper mRNA 3’-end formation.To better determine the role(s) of the THO complex, we have searched for mutant alleles that exhibit a genetic interaction with a strain carrying a deletion of the THO complex component MFT1. Our results suggest that the THO complex is functionally connected to the 3’end formation/mRNA export step. High-resolution transcriptional run-on experiments show that the transcription elongation defect observed in THO complex mutants occurs in close proximity with the cleavage/polyadenylation site sequence of the HSP104 gene. Furthermore, when another gene is fused to the 3’-end of the HSP104 ORF, the elongation defect is shifted to the new 3’-end. Mutations in several genes involved in mRNA export and in mRNP assembly lead to retention of mRNPs in transcription site-foci and to partial degradation of the mRNA by the nuclear exosome. Here, we demonstrate a prominent role of the rate of transcription in the constitution of an export-competent mRNP. We show that a transcription-defective allele of the Rad3p helicase, a component of the TFIIH transcription initiation factor, suppress several export-related phenotypes linked to mutation of Rna14p and members of the THO complex. Biochemical and genetic data indicate that mutation of Rad3p in the context of THO and rna14-3 mutants improves mRNP quality by acting upstream of transcription-site retention and nuclear degradation of the transcripts. As Rad3p mutant effects can be phenocopied by other mutations known to affect transcription and by the addition of transcription elongation drugs, our data suggest that a decreased transcription rate generally favors proper mRNP formation under challenging conditions.