Identification of Allele-Specific RNAi Effectors Targeting Genetic Forms of Parkinson's Disease

Parkinson's disease (PD) is a progressive neurological disorder affecting an estimated 5–10 million people worldwide. Recent evidence has implicated several genes that directly cause or increase susceptibility to PD. As well as advancing understanding of the genetic aetiology of PD these findings suggest new ways to modify the disease course, in some cases through genetic manipulation. Here we generated a ‘walk-through’ series of RNA Pol III-expressed shRNAs targeting both the α-synuclein A30P and LRRK2 G2019S PD-associated mutations. Allele-specific discrimination of the α-synuclein A30P mutation was achieved with alignments at position 10, 13 and 14 in two model systems, including a heterozygous model mimicking the disease setting, whilst 5′RACE was used to confirm stated alignments. Discrimination of the most common PD-linked LRRK2 G2019S mutation was assessed in hemizygous dual-luciferase assays and showed that alignment of the mutation opposite position 4 of the antisense species produced robust discrimination of alleles at all time points studied. Discrimination at this position was subsequently confirmed using siRNAs, where up to 10-fold discrimination was seen. The results suggest that RNAi-mediated silencing of PD-associated autosomal dominant genes could be a novel therapeutic approach for the treatment of the relevant clinical cases of PD in future

Adding to Hans Kuhn's thesis on the emergence of the genetic apparatus of the darwinian advantage to be neither too soluble, nor too insoluble, neither too solid, nor completely liquid

International audienceA scenario of the origin of the genetic apparatus is described where the surface and physico-chemical properties of lipid bilayers and multilayers of vesicles play a crucial role. Peptides, nucleic acids and lipids are 'collaborating' to bring about a first successful genetic apparatus. Lipidic vesicles acquire new properties through hosting nucleic acids that are transiently but covalently linked to lipophilic peptides. These peptides anchor the associated nucleic acids into the surface of lipidic vesicles. In the interior of such vesicles, within the lipidic bilayer, peptidyl transfers occur that are reminiscent of modern-day ribosomal peptidyl transfer reactions. One can expect that the growing peptides eventually acquire, stepwise and essentially arbitrarily, new functions different from anchoring nucleic acids, such as specific aminoacylation of nucleic acids, template-assisted nucleic acid synthesis, nucleotide deoxygenation and fatty acid synthesis

Zur präbiotischen synthese von nukleosiden und nukleotiden

International audienceVielleicht führt mehr als nur ein Weg zu RNA: Jüngste Entdeckungen bei der Synthese von Nukleosid- und Nukleotidvorstufen werden im weiteren Kontext der präbiotischen Systemchemie vorgestellt. Aus Gemischen von Butlerows Kohlenhydratvorstufen mit Traubes 5-Formylaminopyrimidinen wurden präbiotische Purin-Nukleoside erhalten, wohingegen beim Zusammenführen von 5-Phosphoribose, Barbitursäure und Melamin supramolekulare Fasern wuchsen

Total syntheses of a conformationally locked North-type methanocarba puromycin analogue and a dinucleotide derivative

International audienceAn original synthetic approach for the first synthesis of an enantiopure methanocarba puromycin (3'-alpha-aminoacylamino-3'-deoxyadenosine) analogue and its cytidine dinucleotide derivative is described. Each compound is conformationally locked in a North-type pucker and exhibits both a pseudoaxial hydroxy group and a pseudoequatorial aminoacyl group. The syntheses were accomplished from D-ribose in 18 and 19 steps, respectively, with key steps being a ring-closing metathesis, a Luche reduction, a Simmons-Smith cyclopropanation, a Mitsunobu coupling, a Mattocks bromoacetylation, a regioselective and a stereoselective nucleophilic substitution, a chemoselective phosphoramidite coupling and a Staudinger-Vilarrasa coupling. Both molecules are being tested for peptidyl transfer efficiency in ribosomes for comparison with the peptidyl transfer kinetics of natural puromycin and other natural and synthetic ribosomal A site substrates

New concept for quantification of similarity relates entropy and energy of objects : first and second law entangled group behavior of micro black holes expected

International audienceWhen the free energy of similar but distinct molecule-sized objects is plotted against the temperature at which their energy and entropy contributions cancel, a highly significant linear dependence results from which the degree of similarity between the distinctly different members within the group of objects can be quantified and a relationship between energy and entropy is derived. This energy-entropy relationship entirely reflects the mathematical structure of thermodynamic equations, is in this sense fundamental and therefore does probably not dependent on material nor scale. The energy-entropy relationship is likely to be of general interest in molecular biology, population biology, synthetic biology, biophysics, chemical thermodynamics, systems chemistry and physics, most notably in particle physics and cosmology. In physics we predict a consistent and perhaps testable way of classifying micro black holes, to be generated in future Large Hadron Collider experiments, by their gravitational energy and area entropy

Total syntheses of a conformationally locked North-type methanocarba puromycin analogue and a dinucleotide derivative

International audienceAn original synthetic approach for the first synthesis of an enantiopure methanocarba puromycin (3'-alpha-aminoacylamino-3'-deoxyadenosine) analogue and its cytidine dinucleotide derivative is described. Each compound is conformationally locked in a North-type pucker and exhibits both a pseudoaxial hydroxy group and a pseudoequatorial aminoacyl group. The syntheses were accomplished from D-ribose in 18 and 19 steps, respectively, with key steps being a ring-closing metathesis, a Luche reduction, a Simmons-Smith cyclopropanation, a Mitsunobu coupling, a Mattocks bromoacetylation, a regioselective and a stereoselective nucleophilic substitution, a chemoselective phosphoramidite coupling and a Staudinger-Vilarrasa coupling. Both molecules are being tested for peptidyl transfer efficiency in ribosomes for comparison with the peptidyl transfer kinetics of natural puromycin and other natural and synthetic ribosomal A site substrates

Synthesis of a deoxyxylopuromycin analogue

International audienceN6-Bis-demethylated deoxyxylopuromycin was synthesized over six steps in 56% overall yield. The key steps are Mitsunobu­ reaction with DPPA and a Staudinger-Vilarrasa coupling

The shortest synthetic route to puromycin analogues using a modified robins approach

International audienceWe are reporting on the utility of commercial vinyl isocyanate for a practical synthetic route from adenosine to N(6)-bis-demethylpuromycin in seven steps and 65% overall yield. A clean one-pot conversion of 3'-bromo-2'-carbamoyl derivative 8 to 3'-amino-3'-deoxyadenosine derivative 10 is the main feature of this synthetic pathway. This synthesis is the shortest synthetic route toward 3'-(aminoacylamido)deoxyadenosines to date