PHD and TFIIS-Like domains of the Bye1 transcription factor determine its multivalent genomic distribution.

The BYpass of Ess1 (Bye1) protein is a putative S. cerevisiae transcription factor homologous to the human cancer-associated PHF3/DIDO family of proteins. Bye1 contains a Plant Homeodomain (PHD) and a TFIIS-like domain. The Bye1 PHD finger interacts with tri-methylated lysine 4 of histone H3 (H3K4me3) while the TFIIS-like domain binds to RNA polymerase (Pol) II. Here, we investigated the contribution of these structural features to Bye1 recruitment to chromatin as well as its function in transcriptional regulation. Genome-wide analysis of Bye1 distribution revealed at least two distinct modes of association with actively transcribed genes: within the core of Pol II- and Pol III-transcribed genes concomitant with the presence of the TFIIS transcription factor and, additionally, with promoters of a subset of Pol II-transcribed genes. Specific loss of H3K4me3 abolishes Bye1 association to gene promoters, but doesn't affect its binding within gene bodies. Genetic interactions suggested an essential role of Bye1 in cell fitness under stress conditions compensating the absence of TFIIS. Furthermore, BYE1 deletion resulted in the attenuation of GAL genes expression upon galactose-mediated induction indicating its positive role in transcription regulation. Together, these findings point to a bimodal role of Bye1 in regulation of Pol II transcription. It is recruited via its PHD domain to H3K4 tri-methylated promoters at early steps of transcription. Once Pol II is engaged into elongation, Bye1 binds directly to the transcriptional machinery, modulating its progression along the gene

Translational activators and mitoribosomal isoforms cooperate to mediate mRNA-specific translation in Schizosaccharomyces pombe mitochondria

International audienceMitochondrial mRNAs encode key subunits of the oxidative phosphorylation complexes that produce energy for the cell. In Saccharomyces cerevisiae, mitochondrial translation is under the control of translational activators, specific to each mRNA. In Schizosaccharomyces pombe, which more closely resembles the human system by its mitochondrial DNA structure and physiology, most translational activators appear to be either lacking, or recruited for post-translational functions. By combining bioinformatics, genetic and biochemical approaches we identified two interacting factors, Cbp7 and Cbp8, controlling Cytb production in S. pombe. We show that their absence affects cytb mRNA stability and impairs the detection of the Cytb protein. We further identified two classes of Cbp7/Cbp8 partners and showed that they modulated Cytb or Cox1 synthesis. First, two isoforms of bS1m, a protein of the small mitoribosomal subunit, that appear mutually exclusive and confer translational specificity. Second, a complex of four proteins dedicated to Cox1 synthesis, which includes an RNA helicase that interacts with the mitochondrial ribosome. Our results suggest that S. pombe contains, in addition to complexes of translational activators, a heterogeneous population of mitochondrial ribosomes that could specifically modulate translation depending on the mRNA translated, in order to optimally balance the production of different respiratory complex subunits

Two RNA Polymerase I Subunits Control the Binding and Release of Rrn3 during Transcription▿ †

Rpa34 and Rpa49 are nonessential subunits of RNA polymerase I, conserved in species from Saccharomyces cerevisiae and Schizosaccharomyces pombe to humans. Rpa34 bound an N-terminal region of Rpa49 in a two-hybrid assay and was lost from RNA polymerase in an rpa49 mutant lacking this Rpa34-binding domain, whereas rpa34Δ weakened the binding of Rpa49 to RNA polymerase. rpa34Δ mutants were caffeine sensitive, and the rpa34Δ mutation was lethal in a top1Δ mutant and in rpa14Δ, rpa135(L656P), and rpa135(D395N) RNA polymerase mutants. These defects were shared by rpa49Δ mutants, were suppressed by the overexpression of Rpa49, and thus, were presumably mediated by Rpa49 itself. rpa49 mutants lacking the Rpa34-binding domain behaved essentially like rpa34Δ mutants, but strains carrying rpa49Δ and rpa49-338::HIS3 (encoding a form of Rpa49 lacking the conserved C terminus) had reduced polymerase occupancy at 30°C, failed to grow at 25°C, and were sensitive to 6-azauracil and mycophenolate. Mycophenolate almost fully dissociated the mutant polymerase from its ribosomal DNA (rDNA) template. The rpa49Δ and rpa49-338::HIS3 mutations had a dual effect on the transcription initiation factor Rrn3 (TIF-IA). They partially impaired its recruitment to the rDNA promoter, an effect that was bypassed by an N-terminal deletion of the Rpa43 subunit encoded by rpa43-35,326, and they strongly reduced the release of the Rrn3 initiation factor during elongation. These data suggest a dual role of the Rpa49-Rpa34 dimer during the recruitment of Rrn3 and its subsequent dissociation from the elongating polymerase

Bye1 is associated with the Pol II Rpb1 subunit <i>in vivo</i>: (A) Co-immunoprecipitation of Bye1-Myc, HA-TFIIS and Rpb1 from cross-linked cellular extracts; (B) Bye1 and TFIIS interact with the jaw domain (aa 1154–1252) of Rpb1 by Y2H.

<p>All three domains, TFIIS-like, PHD and SPOC, are essential for Rpb1-Bye1 interaction: overlay of the Y190 strain transformed with either empty G_AD/G_BD vectors or G_AD fused to the Rpb1 jaw domain and G_BD fused to Bye1 or TFIIS. Left panel: control for the autoactivated β-galactosidase activity. Right panel: interactions are observed for Rpb1 and TFIIS and for Rpb1 and Bye1 the whole and the PHD and SPOC deleted proteins.</p

Bye1 association with class II genes requires active transcription: (A) Quantification of Pol II enrichment in <i>rpb1-1</i> mutant and Bye1-Myc enrichment in <i>RPB1 WT</i> and <i>rpb1-1</i> strains at 30°C and 37°C by ChIP; (B) Quantification of Bye1 enrichment at the heat-induced <i>HSP104</i> locus at 28°C and 37°C by ChIP.

<p>An intergenic genomic locus is used for a relative occupancy calculation; 3′ORF and mORF stand for the 3′end and middle coding regions.</p