158 research outputs found
Reconstitution of archaeal H/ACA small ribonucleoprotein complexes active in pseudouridylation
Pseudouridine (Ψ) are frequently modified residues in RNA. In Eukarya, their formation is catalyzed by enzymes or by ribonucleoprotein complexes (RNPs) containing H/ACA snoRNAs. H/ACA sRNA and putative ORFs for H/ACA sRNP proteins (L7Ae, aCBF5, aNOP10 and aGAR1) were found in Archaea. Here, by using Pyrococcus abyssi recombinant proteins and an in vitro transcribed P.abyssi H/ACA sRNA, we obtained the first complete in vitro reconstitution of an active H/ACA RNP. Both L7Ae and the aCBF5 RNA:Ψ synthase bind directly the sRNA; aCBF5 also interacts directly and independently with aNOP10 and aGAR1. Presence of aCBF5, aNOP10 and a U residue at the pseudouridylation site in the target RNA are required for RNA target recruitment. In agreement, we found that the aCBF5–aNOP10 pair is the minimal set of proteins needed for the formation of a particle active for pseudouridylation. However, particles more efficient in targeted pseudouridylation can be formed with the addition of proteins L7Ae and/or aGAR1. Although necessary for optimal activity, the conserved ACA motif in the sRNA was found to be not essential
Identification of determinants in the protein partners aCBF5 and aNOP10 necessary for the tRNA:Ψ55-synthase and RNA-guided RNA:Ψ-synthase activities
Protein aNOP10 has an essential scaffolding function in H/ACA sRNPs and its interaction with the pseudouridine(Ψ)-synthase aCBF5 is required for the RNA-guided RNA:Ψ-synthase activity. Recently, aCBF5 was shown to catalyze the isomerization of U55 in tRNAs without the help of a guide sRNA. Here we show that the stable anchoring of aCBF5 to tRNAs relies on its PUA domain and the tRNA CCA sequence. Nonetheless, interaction of aNOP10 with aCBF5 can counterbalance the absence of the PUA domain or the CCA sequence and more generally helps the aCBF5 tRNA:Ψ55-synthase activity. Whereas substitution of the aNOP10 residue Y14 by an alanine disturbs this activity, it only impairs mildly the RNA-guided activity. The opposite effect was observed for the aNOP10 variant H31A. Substitution K53A or R202A in aCBF5 impairs both the tRNA:Ψ55-synthase and the RNA-guided RNA:Ψ-synthase activities. Remarkably, the presence of aNOP10 compensates for the negative effect of these substitutions on the tRNA: Ψ55-synthase activity. Substitution of the aCBF5 conserved residue H77 that is expected to extrude the targeted U residue in tRNA strongly affects the efficiency of U55 modification but has no major effect on the RNA-guided activity. This negative effect can also be compensated by the presence of aNOP10
A partial localisation of the binding sites of the 50 S subunit proteins L1, L20 and L23 on 23 S ribosomal RNA of Escherichia coli
Recherche des gènes d'ARN non codant
La masse considérable de données brutes extraite des programmes de séquençage nécessite de nouvelles techniques d'analyse. La première étape visant à annoter les séquences génomiques est la recherche de régions codant des protéines (ORF pour Open Reading Frame). Cependant les gènes d'ARN non codant (ARNnc), qui ne produisent pas de protéines mais des ARN fonctionnels en tant que tels, ne présentent pas les signaux utilisés pour la détection d'ORF. La recherche systématique des gènes d'ARNnc requiert de ce fait le développement d'outils appropriés, ce qui représente un challenge de premier ordre dans l'ère post génomique. Nous proposons ainsi d'utiliser une méthode issue de l'apprentissage statistique basée sur les machines à vecteurs support (SVM) qui est applicable à l'ensemble des séquences génomiques. Cette approche a été validée par la recherche de snoRNA à boîtes C/D ou H/ACA dans le génome de la levure S. cerevisiae et dans les génomes d'Archaea du genre Pyrococcus
A Multi-Site Constraint Programming Model of Alternative Splicing Regulation
Alternative splicing is a key process in post-transcriptional regulation, by which several kinds of mature RNA can be obtained from the same premessenger RNA. The resulting combinatorial complexity contributes to biological diversity, especially in the case of the human immunodeficiency virus HIV-1. Using a constraint programming approach, we develop a model of the alternative splicing regulation in HIV-1. Our model integrates different scales (single site vs. multiple sites), and thus allows us to exploit several types of experimental data available to us
Pengaruh Kesadaran Politik terhadap Pasrtisipasi Politik Masyarakat Kecamatan Tenayan Raya dalam Pemilihan Umum Walikota Pekanbaru Tahun 2017
One manifestation of political consciousness is political participation in the election of the mayor. Political participation based on political consciousness will encourage people to exercise their right to vote rationally and in accordance with the aspirations of the people. But in reality, the level of voter participation in the election mayor of Pekanbaru in 2017 is still lacking. Percentage of voters\u27 participation in sub district Tenayan Raya only reached 43.72%, which means there are still many voters who did not participate that reached 56.28%. This study aims to explain the effect of political awareness on political participation. This study uses quantitative descriptive by spreading questionnaires to 100 respondents in four urban villages in Tenayan Raya sub district, then the data is analyzed by using logistic regression. Based on the result of research, it can be concluded that there is a significant influence between political awareness on political participation because the value of p value Chi-Square 13,105 > Chi-Square table 3,841. In the Pseudo R Square table of 16.4% political awareness contribution to political participation, in other words 83.6% other factors from outside the model that explain the dependent variable. Based on the analysis of political awareness data of Tenayan Raya sub-district is low down to 68%, while the political participation of Tenayan Raya community in the election of mayor of Pekanbaru is also categorized as low by 94%
Small-Molecule Inhibition of HIV pre-mRNA Splicing as a Novel Antiretroviral Therapy to Overcome Drug Resistance
The development of multidrug-resistant viruses compromises antiretroviral therapy efficacy and limits therapeutic options. Therefore, it is an ongoing task to identify new targets for antiretroviral therapy and to develop new drugs. Here, we show that an indole derivative (IDC16) that interferes with exonic splicing enhancer activity of the SR protein splicing factor SF2/ASF suppresses the production of key viral proteins, thereby compromising subsequent synthesis of full-length HIV-1 pre-mRNA and assembly of infectious particles. IDC16 inhibits replication of macrophage- and T cell–tropic laboratory strains, clinical isolates, and strains with high-level resistance to inhibitors of viral protease and reverse transcriptase. Importantly, drug treatment of primary blood cells did not alter splicing profiles of endogenous genes involved in cell cycle transition and apoptosis. Thus, human splicing factors represent novel and promising drug targets for the development of antiretroviral therapies, particularly for the inhibition of multidrug-resistant viruses
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