78 research outputs found
F-box proteins: more than baits for the SCF?
Regulation of protein stability through the ubiquitin proteasome system is a key mechanism underlying numerous cellular processes. The ubiquitin protein ligases (or E3) are in charge of substrate specificity and therefore play a pivotal role in the pathway. Among the several different E3 enzyme families, the SCF (Skp1-Cullin-F box protein) is one of the largest and best characterized. F-box proteins, in addition to the loosely conserved F-box motif that binds Skp1, often carry typical protein interaction domains and are proposed to recruit the substrate to the SCF complex. Strikingly, genomes analysis revealed the presence of large numbers of F-box proteins topping to nearly 700 predicted in Arabidopsis thaliana. Recent evidences in various species suggest that some F-box proteins have functions not directly related to the SCF complex raising questions about the actual connection between the large F-box protein family and protein degradation, but also about their origins and evolution
Anticodon wobble uridine modification by Elongator at the crossroad of cell signaling, differentiation and diseases
First identified 20 years ago as an RNA polymerase II-associated putative histone acetyltransferase, the conserved Elongator complex has since been recognized as the central player of a complex, regulated, and biologically relevant epitranscriptomic pathway targeting the wobble uridine of some tRNAs. Numerous studies have contributed to three emerging concepts resulting from anticodon modification by Elongator: the codon-specific control of translation, the ability of reprogramming translation in various physiological or pathological contexts, and the maintenance of proteome integrity by counteracting protein aggregation. These three aspects of tRNA modification by Elongator constitute a new layer of regulation that fundamentally contributes to gene expression and are now recognized as being critically involved in various human diseases
Construction of a set of vectors allowing inducible production of siRNA in Schizosaccharomyces pombe.
RNA interference (RNAi) is a sequence-specific gene silencing mechanism. It is induced by the formation of dsRNA that are recognised by the Dicer complex and processed into 21-23 long oligonucleotides called siRNA (short interfering RNA). Subsequently, RISC (RNA-Inducing Silencing Complex) binds siRNA that targets the complex towards its homologous mRNA (DYKXHOORN et al., 2003) which is eventually degraded.
In contrast to budding yeast, the entire pathway is conserved in the fission yeast Schizosaccharomyces pombe, making it a valuable organism to both study physiological RNAi and to use it as a inducible gene knock-down tool.
In an attempt to apply this method in the fission yeast, we are using three different approaches to produce siRNA. In each case, a vector containing a regulatable promoter activated in presence of tetracycline (tTA') (GOSSEN et al., 1995) is generated and the ura4 marker required for growth on medium lacking uracil serves as reporter. First, a vector expressing the full lenght antisens RNA of ura4 (800 nucleotides) (RAPONI and ARNDT, 2003) is used. Second, we are trying to generate much shorter dsRNA where both strands are linked by either a short hairpin of 25 nucleotides (BRUMMELKAMP et al., 2002) or a longer one of 350 nucleotides (SCHRAMKE and ALLSHIRE, 2003). The ability of these different dsRNA to induce silencing of ura4 will be presented
Translational control of cell division by elongator
SummaryElongator is required for the synthesis of the mcm5s2 modification found on tRNAs recognizing AA-ending codons. In order to obtain a global picture of the role of Elongator in translation, we used reverse protein arrays to screen the fission yeast proteome for translation defects. Unexpectedly, this revealed that Elongator inactivation mainly affected three specific functional groups including proteins implicated in cell division. The absence of Elongator results in a delay in mitosis onset and cytokinesis defects. We demonstrate that the kinase Cdr2, which is a central regulator of mitosis and cytokinesis, is under translational control by Elongator due to the Lysine codon usage bias of the cdr2 coding sequence. These findings uncover a mechanism by which the codon usage, coupled to tRNA modifications, fundamentally contributes to gene expression and cellular functions
Histone H2B ubiquitylation represses gametogenesis by opposing RSC-dependent chromatin remodeling at the ste11 master regulator locus.
In fission yeast, the ste11 gene encodes the master regulator initiating the switch from vegetative growth to gametogenesis. In a previous paper, we showed that the methylation of H3K4 and consequent promoter nucleosome deacetylation repress ste11 induction and cell differentiation (Materne et al., 2015) but the regulatory steps remain poorly understood. Here we report a genetic screen that highlighted H2B deubiquitylation and the RSC remodeling complex as activators of ste11 expression. Mechanistic analyses revealed more complex, opposite roles of H2Bubi at the promoter where it represses expression, and over the transcribed region where it sustains it. By promoting H3K4 methylation at the promoter, H2Bubi initiates the deacetylation process, which decreases chromatin remodeling by RSC. Upon induction, this process is reversed and efficient NDR (nucleosome depleted region) formation leads to high expression. Therefore, H2Bubi represses gametogenesis by opposing the recruitment of RSC at the promoter of the master regulator ste11 gene. DOI: http://dx.doi.org/10.7554/eLife.13500.00
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Reciprocal regulation of TORC signaling and tRNA modifications by Elongator enforces nutrient-dependent cell fate
Nutrient availability has a profound impact on cell fate. Upon nitrogen starvation, wild-type fission yeast cells uncouple cell growth from cell division to generate small, round-shaped cells that are competent for sexual differentiation. The TORC1 (TOR complex 1) and TORC2 complexes exert opposite controls on cell growth and cell differentiation, but little is known about how their activity is coordinated. We show that transfer RNA (tRNA) modifications by Elongator are critical for this regulation by promoting the translation of both key components of TORC2 and repressors of TORC1. We further identified the TORC2 pathway as an activator of Elongator by down-regulating a Gsk3 (glycogen synthase kinase 3)-dependent inhibitory phosphorylation of Elongator. Therefore, a feedback control is operating between TOR complex (TORC) signaling and tRNA modification by Elongator to enforce the advancement of mitosis that precedes cell differentiation
Production d'anticorps contre la diméthylase (Dim1p) de l'ARNr 18S de Saccharomyces cerevisiae
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