Project description:The Fil1 transcription factor regulates the response to amino acid starvation. Here we analyse the effects of overexpressing the fil1 coding sequence and of its deregulated expression. The following strains were used: 1] Strains expressing the fil1 gene under the control of the thiamine-repressible nmt1 promoter (nmt-fil1) 2] Strains in which 6 uORFs (upstream Open Reading Frames) in the fil1 5'-UTR have been inactivated, leading to deregulated translation of the fil1 mRNA (6uORF_mutant)
Project description:CHX is an inhibitor of translation elongation often used in ribosome profiling experiments. There is evidence that CHX treatment of cells may cause artefacts in the distribution of ribosomes on mRNAs. We investigate this possibility in S. pombe by performing ribosome profiling in the presence and absence of this drug.
Project description:Translation complexes are stabilised in vivo using formaldehyde. Cells are lysed, and cell extracts are treated with RNase I to produce protected RNA fragments (FPs or footprints). Extracts are then run through sucrose gradients to separate small ribosomal subunits and full ribosomes. FPs are isolated from each of these fractions and analysed by sequencing. Note this is a modified version of ribo-seq (a.k.a. ribosome profiling)
Project description:RNA-sequencing of S. pombe wild type, ppa1 mutants, and tor1 mutants, grown under condition of normal nitrogen and under nitrogen starvation.
Project description:To examine whether naturally occurring duplications are altering gene expression we chose eight pairs of closely related strains (less than 150 SNPS between each pair) that contained at least one unshared duplication and performed pair-wise two-colour microarray analysis.
Project description:Target of Rapamycin Complex 1 (TORC1) signaling promotes growth and ageing. Inhibition of TORC1 leads to a down-regulation of factors that stimulate protein translation, which in turn contributes to longevity. TORC1-dependent post-transcriptional regulation of protein translation has been well studied, while analogous transcriptional regulation is less understood. Here we screened fission yeast deletion mutants for resistance to Torin1, which inhibits TORC1 and cell growth. Cells lacking the GATA transcription factor Gaf1 (gaf1Δ) grew normally even in high doses of Torin1. The gaf1Δ mutation shortened the chronological lifespan of non-dividing cells and diminished the longevity triggered by Torin1 treatment. Expression profiling and genome-wide binding experiments showed that, upon TORC1 inhibition, Gaf1 directly up-regulated genes for small-molecule metabolic pathways and indirectly repressed genes for protein translation. Surprisingly, Gaf1 bound to, and down-regulated the tRNA genes, so also functions as a transcription factor for genes transcribed by RNA polymerase III. Thus, Gaf1 controls the transcription of both coding and tRNA genes to inhibit translation and growth downstream of TORC1.