Translatome and transcriptome analysis of TMA20 (MCT-1) and TMA64 (eIF2D) knockout yeast strains

Abstract

TMA20 (MCT-1), TMA22 (DENR) and TMA64 (eIF2D) are eukaryotic translation factors involved in ribosome recycling and re-initiation. They operate with P-site bound tRNA in post-termination or (re-)initiation translation complexes, thus participating in the removal of 40S ribosomal subunit from mRNA stop codons after termination and controlling translation re-initiation on mRNAs with upstream open reading frames (uORFs), as well as de novo initiation on some specific mRNAs. Here we report ribosomal profiling data of S.cerevisiae strains with individual deletions of TMA20 , TMA64 or both TMA20 and TMA64 genes. We provide RNA-Seq and Ribo-Seq data from yeast strains grown in the rich YPD or minimal SD medium. We illustrate our data by plotting differential distribution of ribosomal-bound mRNA fragments throughout uORFs in 5′-untranslated region (5′ UTR) of GCN4 mRNA and on mRNA transcripts encoded in MAT locus in the mutant and wild-type strains, thus providing a basis for investigation of the role of these factors in the stress response, mating and sporulation. We also document a shift of transcription start site of the APC4 gene which occurs when the neighboring TMA64 gene is replaced by the standard G418-resistance cassette used for the creation of the Yeast Deletion Library. This shift results in dramatic deregulation of the APC4 gene expression, as revealed by our Ribo-Seq data, which can be probably used to explain strong genetic interactions of TMA64 with genes involved in the cell cycle and mitotic checkpoints. Raw RNA-Seq and Ribo-Seq data as well as all gene counts are available in NCBI Gene Expression Omnibus (GEO) repository under GEO accession GSE122039 ( https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE122039 ).

Translational induction of ATF4 during integrated stress response requires noncanonical initiation factors eIF2D and DENR

“…In Saccharomyces cerevisae , the homologs of eIF2D , DENR, and MCTS-1 (Tma64 Tma22, and Tma20, respectively) have more prominent roles in ribosome recycling as evidenced by increased translation of downstream of ORFs and uORFs 58 . The translation of GCN4, the S. cerevisiae equivalent of ATF4, also increases upon the loss of these recycling factors 59 . Together, these findings suggest that the in vivo roles of eIF2D, DENR, and MCTS have diverged during evolution, with metazoan factors playing more prominent roles in translational initiation of ATF4 mRNA.…”

uORF-mediated translational control: recently elucidated mechanisms and implications in cancer

“…A recent report revealed that co-deletion of the yeast eIF2D (Tma64) and a ribosome recycling factor (Tma20 or Tma22) results in translation reinitiation at downstream AUG codons after translation termination and hence promotes translation of uORF-containing reporters [118]. Moreover, eIF2D knockout altered ribosome profiling reads in uORFs of GCN4, suggesting a regulatory role for eIF2D in translation of uORFcontaining mRNAs [119]. Whether cell stress or oncogenic signaling modulates the activity of these factors and hence influences uORF activation or CDS translation remains to be investigated.…”

In vivo chromatin organization on native yeast telomeric regions is independent of a cis-telomere loopback conformation

“…However, it has become clear that these marker insertions and analyses for TPE for example may be difficult [27]. Also, marker insertion at any location in the genome can affect the transcriptional potential of an area surrounding the marker and hence the chromatin configuration may not reflect that of the native state [28]. Genome wide analyses of nucleosome positioning suggested a low content of nucleosomes across the X elements [29], whereas a parallel study suggested positioned nucleosomes over those same areas [30].…”