Development of an efficient cis-trans-cis ribozyme cassette to inactivate plant genes

Summary Inactivation of a targeted gene is one of the main strategies used to understand their precise cellular role. In plants, apart from chemical or physical mutagenesis and random insertions of DNA elements followed by screening for a desired phenotype, the most common strategy to inhibit the expression of a given gene involves RNA silencing. This can be achieved either through antisense suppression, sense over-expression leading to co-suppression, or expression of double-stranded DNA constructs (dsRNA). The use of ribozymes to inhibit gene product accumulation has only been occasionally attempted, mainly because of the more complex genetic engineering procedure involved, although the specificity of ribozymes can be an important factor when targeting close members of a gene family. We report here the development of a new cis -acting ribozyme cassette for the production of RNAs with desired termini. Attention to many details has been brought in order to provide a powerful procedure for plant application. For example, ultrastable GNRA tetraloops were substituted for both loops II and III of cis -acting hammerhead sequences, thereby favouring folding into the catalytically active structure that results in the self-cleavage of all transcripts. We demonstrate the usefulness of this cassette by producing a ribozyme that cleaves in trans , originally embedded in the cis -acting self-cleaving cassette. The activity of the cistrans-cis construct, was demonstrated both in vitro and in vivo , in transgenic plants with the specific cleavage of an mRNA encoding a 2-oxo-glutarate-dependant dioxygenase predominantly expressed in pistils tissues and in leaves, from the wild potato Solanum chacoense

The impact of culture on neuropsychological performance: A global social cognition study across 12 countries

AbstractBackgroundDecades of researches aiming to unveil truths about human neuropsychology may have instead unveil facts appropriate to only a fraction of the world's population: those living in western educated rich democratic nations (Muthukrishna et al., 2020 Psych Sci). So far, most studies were conducted as if education and cultural assumptions on which neuropsychology is based were universals and applied everywhere in the world. The importance given to sociological or cultural factors is thus still relatively ignored. With the growth of international clinical studies on dementia, we believe that documenting the potential inter‐cultural differences at stake in a common neuropsychological assessment is an essential topic. This study thus aimed to explore these potential variations in two classical tasks used in neuropsychology that are composing the mini‐SEA (Bertoux et al., 2012 JNNP), i.e. a reduced version of the well‐known Ekman faces (FER), where one has to recognize facial emotions, and a modified version of the Faux Pas test (mFP), where one has to detect and explain social faux.MethodThe data of 573 control participants were collected through the Social Cognition & FTLD Network, an international consortium investigating social cognitive changes in dementia covering 3 continents (18 research centres in 12 countries). Impact of demographic factors and the effect of countries on performance (mini‐SEA, FER, mFP) were explored through linear mixed‐effects models.ResultAge, education and gender were found to significantly impact the performance of the mini‐SEA subtests. Significant and important variations across the countries were also retrieved, with England having the highest performance for all scores. When controlling for demographical factors, differences within countries explained between 14% (mFP) and 24% (FER) of the variance at the mini‐SEA. These variations were not explained by any economical or sociological metrics.ConclusionImportant variations of performance were observed across the 12 countries of the consortium, showing how cultural differences may critically impact neuropsychological performance in international studies

Canada-Québec

Taillon Patrick, Perreault Frédéric, Binette Amélie. Canada-Québec. In: Annuaire international de justice constitutionnelle, 36-2020, 2021. L'état d'exception, nouveau régime de droit commun des droits et libertés? Du terrorisme à l'urgence sanitaire- L'opinion publique aujourd'hui. Regards pluridisciplinaires. pp. 901-923

PACE4-Based Molecular Targeting of Prostate Cancer Using an Engineered 64Cu-Radiolabeled Peptide Inhibitor

The potential of PACE4 as a pharmacological target in prostate cancer has been demonstrated as this proprotein convertase is strongly overexpressed in human prostate cancer tissues and its inhibition, using molecular or pharmacological approaches, results in reduced cell proliferation and tumor progression in mouse tumor xenograft models. We developed a PACE4 high-affinity peptide inhibitor, namely, the multi-leucine (ML), and sought to determine whether this peptide could be exploited for the targeting of prostate cancer for diagnostic or molecular imaging purposes. We conjugated a bifunctional chelator 1,4,7-triazacyclononane-1,4,7- triacetic acid (NOTA) to the ML peptide for copper-64 (64Cu) labeling and positron emission tomography (PET)– based prostate cancer detection. Enzyme kinetic assays against recombinant PACE4 showed that the NOTA-modified ML peptide displays identical inhibitory properties compared to the unmodified peptide. In vivo biodistribution of the 64Cu/NOTA-ML peptide evaluated in athymic nude mice bearing xenografts of two human prostate carcinoma cell lines showed a rapid and high uptake in PACE4-expressing LNCaP tumor at an early time point and in PACE4-rich organs. Co-injection of unlabeled peptide confirmed that tumor uptake was target-specific. PACE4-negative tumors displayed no tracer uptake 15 minutes after injection, while the kidneys, demonstrated high uptake due to rapid renal clearance of the peptide. The present study supports the feasibility of using a 64Cu/NOTA-ML peptide for PACE4-targeted prostate cancer detection and PACE4 status determination by PET imaging but also provides evidence that ML inhibitor–based drugs would readily reach tumor sites under in vivo conditions for pharmacological intervention or targeted radiation therapy

Transcriptional pausing at the translation start site operates as a critical checkpoint for riboswitch regulation.

International audienceOn the basis of nascent transcript sequencing, it has been postulated but never demonstrated that transcriptional pausing at translation start sites is important for gene regulation. Here we show that the Escherichia coli thiamin pyrophosphate (TPP) thiC riboswitch contains a regulatory pause site in the translation initiation region that acts as a checkpoint for thiC expression. By biochemically probing nascent transcription complexes halted at defined positions, we find a narrow transcriptional window for metabolite binding, in which the downstream boundary is delimited by the checkpoint. We show that transcription complexes at the regulatory pause site favour the formation of a riboswitch intramolecular lock that strongly prevents TPP binding. In contrast, cotranscriptional metabolite binding increases RNA polymerase pausing and induces Rho-dependent transcription termination at the checkpoint. Early transcriptional pausing may provide a general mechanism, whereby transient transcriptional windows directly coordinate the sensing of environmental cues and bacterial mRNA regulation