Le programme Pyramide: un programme de prévention ciblée de l'affiliation aux gangs de rue s'adressant aux parents de préadolescents hébergés au Centre jeunesse de Montréal-Institut Universitaire

Rapport d'analyse d'intervention présenté à la Faculté des arts et sciences en vue de l'obtention du grade de Maîtrise ès sciences (M. Sc.) en psychoéducationLe projet d'intervention vise à prévenir l'affiliation aux gangs de rue chez les préadolescents hébergés en milieu substitut en améliorant les habiletés parentales de leurs parents: les renforcements positifs, la supervision, la discipline et la relation parent-préadolescent. Le programme de prévention développé s'inspire de l'Adolescent Transition Program (Claes & Poirier, 1995; Dishion & Kavanagh, 2003) et du Strengthening Families Program : For Parents and Youth 10-14 (Molgaard, Kumpfer & Fleming, 2007). Cinq parents participent au programme développé et implanté au Centre jeunesse de Montréal-Institut Universitaire, à l'unité l'Entreprise située sur le site du mont St-Antoine. Le devis de type expérimental utilisé est un protocole à cas unique de type A-B-A avec 14 points de mesure répétée. Les habiletés parentales sont évaluées chez trois parents, par les parents eux-mêmes et par leur préadolescent. Les résultats obtenus auprès des parents montrent, chez un parent, un accroissement léger des renforcements positifs des comportements adéquats de son préadolescent et une amélioration de la relation parent-préadolescent suite à l'introduction de l'intervention. Les résultats obtenus auprès des préadolescents montrent une amélioration de la discipline chez un parent suite à l'introduction de l'intervention; chez un autre parent, ils montrent une légère amélioration de la relation parent-préadolescent. Il y a incohérence entre les sources au niveau des améliorations observées

Requirement for PBAF in transcriptional repression and repair at DNA breaks in actively transcribed regions of chromatin

Actively transcribed regions of the genome are vulnerable to genomic instability. Recently, it was discovered that transcription is repressed in response to neighboring DNA double-strand breaks (DSBs). It is not known whether a failure to silence transcription flanking DSBs has any impact on DNA repair efficiency or whether chromatin remodelers contribute to the process. Here, we show that the PBAF remodeling complex is important for DSB-induced transcriptional silencing and promotes repair of a subset of DNA DSBs at early time points, which can be rescued by inhibiting transcription globally. An ATM phosphorylation site on BAF180, a PBAF subunit, is required for both processes. Furthermore, we find that subunits of the PRC1 and PRC2 polycomb group complexes are similarly required for DSB-induced silencing and promoting repair. Cancer-associated BAF180 mutants are unable to restore these functions, suggesting PBAF's role in repressing transcription near DSBs may contribute to its tumor suppressor activity

Structure-based design of a disulfide-linked oligomeric form of the simian virus 40 (SV40) large T antigen DNA-binding domain

With the aim of forming the ‘lock-washer’ conformation of the origin-binding domain of SV40 large T antigen in solution, using structure-based analysis an intermolecular disulfide bridge was engineered into the origin-binding domain to generate higher order oligomers in solution. The 1.7 Å resolution structure shows that the mutant forms a spiral in the crystal and has the de novo disulfide bond at the protein interface, although structural rearrangements at the interface are observed relative to the wild type

Histone H2A variants play a key role at DNA double-strand breaks during repair pathway choice

Histone post-translational modifications and variants play crucial roles in the adaptability of chromatin structure, facilitating rapid responses necessary for biological processes such as transcription, replication, and DNA damage signaling. Notably, DNA double-strand break (DSB) signaling heavily relies on these histone modifications, with signal amplification and the recruitment of specific DNA repair factors being dictated by them. Among the histones, H2A and its variants are central to this response, with phosphorylation of the variant H2A.X being the initial and most characteristic histone mark deposit upon DNA damage detection. Additional post-translational modifications of H2A and its variants contribute to the selective recruitment of DNA repair factors and influence the choice of DNA repair pathways. This review provides a summary of current knowledge regarding the roles of histone H2A post-translational modifications and variants in DSB signaling and repair, with a particular emphasis on modifications and variants that impact the choice of repair pathways. Additionally, the involvement of histone chaperones, chromatin modifiers, and remodelers in these processes is discussed

Integrating DNA damage repair with the cell cycle

Abstract DNA is labile and constantly subject to damage. In addition to external mutagens, DNA is continuously damaged by the aqueous environment, cellular metabolites and is prone to strand breakage during replication. Cell duplication is orchestrated by the cell division cycle and specific DNA structures are processed differently depending on where in the cell cycle they are detected. This is often because a specific structure is physiological in one context, for example during DNA replication, while indicating a potentially pathological event in another, such as interphase or mitosis. Thus, contextualising the biochemical entity with respect to cell cycle progression provides information necessary to appropriately regulate DNA processing activities. We review the links between DNA repair and cell cycle context, drawing together recent advances

The histone demethylase LSD1/KDM1A promotes the DNA damage response

Histone demethylation is known to regulate transcription, but its role in other processes is largely unknown. We report a role for the histone demethylase LSD1/KDM1A in the DNA damage response (DDR). We show that LSD1 is recruited directly to sites of DNA damage. H3K4 dimethylation, a major substrate for LSD1, is reduced at sites of DNA damage in an LSD1-dependent manner. The E3 ubiquitin ligase RNF168 physically interacts with LSD1 and we find this interaction to be important for LSD1 recruitment to DNA damage sites. Although loss of LSD1 did not affect the initial formation of pH2A.X foci, 53BP1 and BRCA1 complex recruitment were reduced upon LSD1 knockdown. Mechanistically, this was likely a result of compromised histone ubiquitylation preferentially in late S/G2. Consistent with a role in the DDR, knockdown of LSD1 resulted in moderate hypersensitivity to γ-irradiation and increased homologous recombination. Our findings uncover a direct role for LSD1 in the DDR and place LSD1 downstream of RNF168 in the DDR pathway

Two redundant ubiquitin-dependent pathways of BRCA1 localization to DNA damage sites

The tumor suppressor BRCA1 accumulates at sites of DNA damage in a ubiquitin-dependent manner. In this work, we revisit the role of RAP80 in promoting BRCA1 recruitment to damaged chromatin. We find that RAP80 acts redundantly with the BRCA1 RING domain to promote BRCA1 recruitment to DNA damage sites. We show that that RNF8 E3 ligase acts upstream of both the RAP80- and RING-dependent activities, whereas RNF168 acts uniquely upstream of the RING domain. BRCA1 RING mutations that do not impact BARD1 interaction, such as the E2 binding-deficient I26A mutation, render BRCA1 unable to accumulate at DNA damage sites in the absence of RAP80. Cells that combine BRCA1 I26A and mutations that disable the RAP80-BRCA1 interaction are hypersensitive to PARP inhibition and are unable to form RAD51 foci. Our results suggest that in the absence of RAP80, the BRCA1 E3 ligase activity is necessary for recognition of histone H2A Lys13/Lys15 ubiquitylation by BARD1, although we cannot rule out the possibility that the BRCA1 RING facilitates ubiquitylated nucleosome recognition in other ways.Genome Instability and Cance

Rif1 maintains telomeres and mediates DNA repair by encasing DNA ends

In yeast, Rif1 is part of the telosome, where it inhibits telomerase and checkpoint signaling at chromosome ends. In mammalian cells, Rif1 is not telomeric, but it suppresses DNA end resection at chromosomal breaks, promoting repair by nonhomologous end joining (NHEJ). Here, we describe crystal structures for the uncharacterized and conserved ∼125-kDa N-terminal domain of Rif1 from Saccharomyces cerevisiae (Rif1-NTD), revealing an α-helical fold shaped like a shepherd's crook. We identify a high-affinity DNA-binding site in the Rif1-NTD that fully encases DNA as a head-to-tail dimer. Engagement of the Rif1-NTD with telomeres proved essential for checkpoint control and telomere length regulation. Unexpectedly, Rif1-NTD also promoted NHEJ at DNA breaks in yeast, revealing a conserved role of Rif1 in DNA repair. We propose that tight associations between the Rif1-NTD and DNA gate access of processing factors to DNA ends, enabling Rif1 to mediate diverse telomere maintenance and DNA repair functions

Chemical Ubiquitination for Decrypting a Cellular Code

The modification of proteins with ubiquitin (Ub) is an important regulator of eukaryotic biology and deleterious perturbation of this process is widely linked to the onset of various diseases. The regulatory capacity of the Ub signal is high and, in part, arises from the capability of Ub to be enzymatically polymerised to form polyubiquitin (polyUb) chains of eight different linkage types. These distinct polyUb topologies can then be site-specifically conjugated to substrate proteins to elicit a number of cellular outcomes. Therefore, to further elucidate the biological significance of substrate ubiquitination, methodologies that allow the production of defined polyUb species, and substrate proteins that are site-specifically modified with them, are essential to progress our understanding. Many chemically inspired methods have recently emerged which fulfil many of the criteria necessary for achieving deeper insight into Ub biology. With a view to providing immediate impact in traditional biology research labs, the aim of this review is to provide an overview of the techniques that are available for preparing Ub conjugates and polyUb chains with focus on approaches that use recombinant protein building blocks. These approaches either produce a native isopeptide, or analogue thereof, that can be hydrolysable or non-hydrolysable by deubiquitinases. The most significant biological insights that have already been garnered using such approaches will also be summarized