El Premi Nobel de Química 2015: els mecanismes de reparació del DNA

La informació genètica es manté durant generacions en un ambient que pot malbaratar la seva integritat. El Premi Nobel de Química 2015 es va concedir als investigadors Tomas Lindahl, Paul Modrich i Aziz Sancar pel seus treballs en el descobriment dels mecanismes de reparació del DNA. En aquest article es detallen la importància i la repercussió dels descobriments realitzats pels tres investigadors guardonats.Genetic information is maintained for generations in an environment that can damage the integrity of DNA. The Nobel Prize in Chemistry 2015 has been awarded to researchers Tomas Lindahl, Paul Modrich and Aziz Sancar for their work in the discovery of the mechanisms of DNA repair. This article details the main discoveries made by the Nobel laureates

Monitoring denaturation behaviour and comparative stability of DNA triple helices using oligonucleotide–gold nanoparticle conjugates

Gold nanoparticle labels, combined with UV-visible optical absorption spectroscopic methods, are employed to probe the temperature-dependent solution properties of DNA triple helices. By using oligonucleotide-nanoparticle conjugates to characterize triplex denaturation, for the first time triplex to duplex melting transitions may be sensitively monitored, with minimal signal interference from duplex to single strand melting, for both parallel and antiparallel triple helices. Further, the comparative sequence-dependent stability of DNA triple helices may also be examined using this approach. Specifically, triplex to duplex melting transitions for triplexes formed using oligonucleotides that incorporate 8-aminoguanine derivatives were successfully monitored and stabilization of both parallel and antiparallel triplexes following 8-aminoguanine substitutions is demonstrated

Synthesis and properties of small interfering RNA duplexes carrying 5-ethyluridine residues

Oligoribonucleotides carrying 5-ethyluridine units were prepared using solid-phase phosphoramidite chemistry. The introduction of the tert-butyldimethylsilyl group at the 2'-OH position proceeded in good yield and very high 2'-regioselectivity. RNA duplexes carrying 5-ethyluridine either at the sense or the guide strands display RNAi activity comparable to or slightly better than that of unmodified RNA duplexes. Gene suppression experiments using luciferase targets in SH-SY5Y cells show that the ethyl group is generally well accepted at all positions although a small decrease in RNA interference activity is observed when one 5-ethylU residue is incorporated in the 3' overhangs

Resolution of a structural competition involving dimeric G-quadruplex and its C-rich complementary strand

The resolution of the dimeric intermolecular G-quadruplex/duplex competition of the telomeric DNA sequence 5′-TAG GGT TAG GGT-3′ and of its complementary 5′ ACC CTA ACC CTA-3′ is reported. To achieve this goal, melting experiments of both sequences and of the mixtures of these sequences were monitored by molecular absorption, molecular fluorescence and circular dichroism spectroscopies. Molecular fluorescence measurements were carried out using molecular beacons technology, in which the 5′-TAG GGT TAG GGT-3′ sequence was labelled with a fluorophore and a quencher at the ends of the strand. Mathematical analysis of experimental spectroscopic data was performed by means of multivariate curve resolution, allowing the calculation of concentration profiles and pure spectra of all resolved structures (dimeric antiparallel and parallel G-quadruplexes, Watson–Crick duplex and single strands) present in solution. Our results show that parallel G-quadruplex is more stable than antiparallel G-quadruplex. When the complementary C-rich strand is present, a mixture of both G-quadruplex structures and Watson–Crick duplex is observed, the duplex being the major species. In addition to melting temperatures, equilibrium constants for the parallel/antiparallel G-quadruplex equilibrium and for the G-quadruplex/duplex equilibrium were determined from the concentration profiles

Modified siRNAs for the study of the PAZ domain

Chemical modifications aimed at stabilizing the interaction between the 3'-end of siRNAs and the PAZ domain of RISC have been tested for their effect on RNAi activity. Such modifications contribute positively to the stability of siRNAs in human serum

Synthesis and G-Quadruplex-Binding Properties of Defined Acridine Oligomers

The synthesis of oligomers containing two or three acridine units linked through 2-aminoethylglycine using solid-phase methodology is described. Subsequent studies on cell viability showed that these compounds are not cytotoxic. Binding to several DNA structures was studied by competitive dialysis, which showed a clear affinity for DNA sequences that form G-quadruplexes and parallel triplexes. The fluorescence spectra of acridine oligomers were affected strongly upon binding to DNA. These spectral changes were used to calculate the binding constants (K). Log K were found to be in the order of 4–6

Influence of pH and a porphyrin ligand on the stability of a G-quadruplex structure within a duplex segment near the promoter region of the SMARCA4 gene

In a previous work, the formation of G-quadruplex structures in a 44-nucleotide long sequence found near the promoter region of the SMARCA4 gene was reported. The central 25 nucleotides were able to fold into an antiparallel G-quadruplex structure, the stability of which was pH-dependent. In the present work, the effect of the presence of lateral nucleotides and the complementary cytosine-rich strand on the stability of this G-quadruplex has been characterized. Moreover, the role of the model ligand TMPyP4 has been studied. Spectroscopic and separation techniques, as well as multivariate data analysis methods, have been used with these purposes. The results have shown that stability of the G-quadruplex as a function of pH or temperature is greatly reduced in the presence of the lateral nucleotides. The influence of the complementary strand does not prevent the formation of the G-quadruplex. Moreover, attempts to modulate the equilibria by an external ligand led us to determine the influence of the TMPyP4 porphyrin on these complex equilibria. This study could eventually help to understand the regulation of SMARCA4 expression

Stepwise synthesis of oligonucleotide-peptide conjugates containing guanidinium and lipophilic groups in their 3'-termini

Two different series of oligonucleotide-peptide conjugates have been efficiently synthesized by stepwise solid-phase synthesis. First, oligonucleotides and oligonucleotide phosphorothioates containing polar groups at the 3′-termini, such as amine and guanidinium groups were prepared. ODNs conjugates carrying several lysine residues were obtained directly from Fmoc deprotection whereas ODN conjugates with guanidinium groups were obtained by post-synthetic guanidinylation. The second family contains different urea moieties that were achieved by standard protocols. All products were fully characterized by reversed phase HPLC and MALDI-TOF mass spectrometry yielding satisfactory results. Oligonucleotide-phosphorothioate conjugates were evaluated as potential antisense oligonucleotides in the inhibition of the luciferase gene

Alginate Hydrogels as Scaffolds and Delivery Systems to Repair the Damaged Spinal Cord

Alginate (ALG) is a lineal hydrophilic polysaccharide present in brown algae cell walls, which turns into a gel state when hydrated. Gelation readily produces a series of three dimensional (3D) architectures like fibers, capillaries, and microspheres, used as biosensors and bio‐actuators in a plethora of biomedical applications like drug delivery and wound healing. Hydrogels have made a great impact on regenerative medicine and tissue engineering because they are able to mimic the mechanical properties of natural tissues due to their high water content. Recent advances in neurosciences have led to promising strategies for repairing and/or regenerating the damaged nervous system. Spinal cord injury (SCI) is particularly challenging, owing to its devastating medical, human, and social consequences. Although effective therapies to repair the damaged spinal cord (SC) are still lacking, multiple pharmacological, genetic, and cell‐based therapies are currently under study. In this framework, ALG hydrogels constitute a source of potential tools for the development of implants capable of promoting axonal growth and/or delivering cells or drugs at specific damaged sites, which may result in therapeutic strategies for SCI. In this mini‐review, the current state of the art of ALG applications in neural tissues for repairing the damaged spinal cord is discussed