A local ATR-dependent checkpoint pathway is activated by a site-specific replication fork block in human cells.

When replication forks encounter DNA lesions that cause polymerase stalling, a checkpoint pathway is activated. The ATR-dependent intra-S checkpoint pathway mediates detection and processing of sites of replication fork stalling to maintain genomic integrity. Several factors involved in the global checkpoint pathway have been identified, but the response to a single replication fork barrier (RFB) is poorly understood.

A local ATR-dependent checkpoint pathway is activated by a site-specific replication fork block in human cells.

When replication forks encounter DNA lesions that cause polymerase stalling a checkpoint pathway is activated. The ATR-dependent intra-S checkpoint pathway mediates detection and processing of sites of replication fork stalling to maintain genomic integrity. Several factors involved in the global checkpoint pathway have been identified, but the response to a single replication fork barrier (RFB) is poorly understood.

Translesion polymerase eta both facilitates DNA replication and promotes increased human genetic variation at common fragile sites.

Common fragile sites (CFSs) are difficult-to-replicate genomic regions that form gaps and breaks on metaphase chromosomes under replication stress. They are hotspots for chromosomal instability in cancer. Repetitive sequences located at CFS loci are inefficiently copied by replicative DNA polymerase (Pol) delta.

TRF2 Mediates Replication Initiation within Human Telomeres to Prevent Telomere Dysfunction.

The telomeric shelterin protein telomeric repeat-binding factor 2 (TRF2) recruits origin recognition complex (ORC) proteins, the foundational building blocks of DNA replication origins, to telomeres. We seek to determine whether TRF2-recruited ORC proteins give rise to functional origins in telomere repeat tracts. We find that reduction of telomeric recruitment of ORC2 by expression of an ORC interaction-defective TRF2 mutant significantly reduces telomeric initiation events in human cells.

A qPCR-Based Protocol to Quantify DSB Resection.

The nucleolytic degradation of the 5'-ending strand of a Double-Strand DNA break (DSB) is necessary to initiate homologous recombination to correctly repair the break. This process is called DNA end resection and it is finely regulated to prevent genome rearrangements. Here, we describe a protocol to quantify DSB resection rate by qPCR, which could be applied to every organisms whenever the break site and its flanking region sequences are known.

TOPBP1 plays a conserved role in homologous recombination DNA repair through the coordinated recruitment of 53BP1.

Genome maintenance and cancer suppression require homologous recombination (HR) DNA repair. In yeast and mammals, the scaffold protein TOPBP1 has been implicated in HR, although its precise function and mechanism of action remain elusive. In this study, we show that yeast Dpb11 plays an antagonistic role in recombination control through regulated protein interactions.

Biogenesis and function of non-coding RNAs in muscle differentiation and in Duchenne muscular dystrophy.

It is now becoming largely accepted that the non-coding portion of the genome, rather than its coding counterpart, is likely to account for the greater complexity of higher eukaryotes. Moreover, non-coding RNAs have been demonstrated to participate in regulatory circuitries that are crucial for development and differentiation. Whereas the biogenesis and function of small non-coding RNAs, particularly miRNAs (microRNAs), has been extensively clarified in many eukaryotic systems, very little is known about the long non-coding counterpart of the transcriptome.

Exon 45 skipping through U1-snRNA antisense molecules recovers the Dys-nNOS pathway and muscle differentiation in human DMD myoblasts.

Exon skipping has been demonstrated to be a successful strategy for the gene therapy of Duchenne muscular dystrophy (DMD): the rational being to convert severe Duchenne forms into milder Becker ones. Here, we show the selection of U1 snRNA-antisense constructs able to confer effective rescue of dystrophin synthesis in a Δ44 Duchenne genetic background, through skipping of exon 45; moreover, we demonstrate that the resulting dystrophin is able to recover timing of myogenic marker expression, to relocalize neuronal nitric oxide synthase (nNOS) and to rescue expression of miRNAs previously shown to be sensitive to the Dystrophin-nNOS-HDAC2 pathway. Becker mutations display different phenotypes, likely depending on whether the shorter protein is able to reconstitute the wide range of wild-type functions.