Search

The mammalian DNA replication timing (RT) program is crucial for the proper functioning and integrity of the genome. The best-known mechanism for controlling RT is the suppression of late origins of replication in heterochromatin by RIF1. Here, we report that in antigen-activated, hypermutating murine B lymphocytes, RIF1 binds predominantly to early-replicating active chromatin and promotes early replication, but plays a minor role in regulating replication origin activity, gene expression and genome organization in B cells. Furthermore, we find that RIF1 functions in a complementary and non-epistatic manner with minichromosome maintenance (MCM) proteins to establish early RT signatures genome-wide and, specifically, to ensure the early replication of highly transcribed genes. These findings reveal additional layers of regulation within the B cell RT program, driven by the coordinated activity of RIF1 and MCM proteins., (© 2023. The Author(s).)

The DNA double-strand break (DSB) repair factor 53BP1 has long been implicated in V(D)J and class switch recombination (CSR) of mammalian lymphocyte receptors. However, the dissection of the underlying molecular activities is hampered by a paucity of studies [V(D)J] and plurality of phenotypes (CSR) associated with 53BP1 deficiency. Here, we revisit the currently accepted roles of 53BP1 in antibody diversification in view of the recent identification of its downstream effectors in DSB protection and latest advances in genome architecture. We propose that, in addition to end protection, 53BP1-mediated end-tethering stabilization is essential for CSR. Furthermore, we support a pre-DSB role during V(D)J recombination. Our perspective underscores the importance of evaluating repair of DSBs in relation to their dynamic architectural contexts., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The appeal of combining polybenzimidazole (PBI) and graphene oxide (GO) for the manufacturing of membranes is increasingly growing, due to their versatility. Nevertheless, GO has always been used only as a filler in the PBI matrix. In such context, this work proposes the design of a simple, safe, and reproducible procedure to prepare self-assembling GO/PBI composite membranes characterized by GO-to-PBI (X:Y) mass ratios of 1:3, 1:2, 1:1, 2:1, and 3:1. SEM and XRD suggested a homogenous reciprocal dispersion of GO and PBI, which established an alternated stacked structure by mutual π-π interactions among the benzimidazole rings of PBI and the aromatic domains of GO. TGA indicated a remarkable thermal stability of the composites. From mechanical tests, improved tensile strengths but worsened maximum strains were observed with respect to pure PBI. The preliminary evaluation of the suitability of the GO/PBI X:Y composites as proton exchange membranes was executed via IEC determination and EIS. GO/PBI 2:1 (IEC: 0.42 meq g -1 ; proton conductivity at 100 °C: 0.0464 S cm -1 ) and GO/PBI 3:1 (IEC: 0.80 meq g -1 ; proton conductivity at 100 °C: 0.0451 S cm -1 ) provided equivalent or superior performances with respect to similar PBI-based state-of-the-art materials.

Critical illness myopathy (CIM) is an acquired, devastating, multifactorial muscle-wasting disease with incomplete recovery. The impact on hospital costs and permanent loss of quality of life is enormous. Incomplete recovery might imply that the function of muscle stem cells (MuSC) is impaired. We tested whether epigenetic alterations could be in part responsible. We characterized human muscle stem cells (MuSC) isolated from early CIM and analyzed epigenetic alterations (CIM n = 15, controls n = 21) by RNA-Seq, immunofluorescence, analysis of DNA repair, and ATAC-Seq. CIM-MuSC were transplanted into immunodeficient NOG mice to assess their regenerative potential. CIM-MuSC exhibited significant growth deficits, reduced ability to differentiate into myotubes, and impaired DNA repair. The chromatin structure was damaged, as characterized by alterations in mRNA of histone 1, depletion or dislocation of core proteins of nucleosome remodeling and deacetylase complex, and loosening of multiple nucleosome-spanning sites. Functionally, CIM-MuSC had a defect in building new muscle fibers. Further, MuSC obtained from the electrically stimulated muscle of CIM patients was very similar to control MuSC, indicating the impact of muscle contraction in the onset of CIM. CIM not only affects working skeletal muscle but has a lasting and severe epigenetic impact on MuSC.

Waste from electrical and electronic equipment (WEEE) is constantly increasing in quantity and becoming more and more heterogeneous as technology is rapidly advancing. The negative impacts it has on human and environment safety, and its richness in valuable rare earth elements (REEs), are accelerating the necessity of innovative methods for recycling and recovery processes. The aim of this work is to comprehend the adsorption and release mechanisms of two different solid sorbents, activated carbon (AC) and its pentaethylenehexamine (PEHA)-modified derivative (MAC), which were deemed adequate for the treatment of REEs deriving from WEEE. Experimental data from adsorption and release tests, performed on synthetic mono-ionic solutions of yttrium, neodymium, and lanthanum, were modelled via linear regression to understand the better prediction between the Langmuir and the Freundlich isotherms for each REE-sorbent couple. The parameters extrapolated from the mathematical modelling were useful to gain an a priori knowledge of the REEs-sorbents interactions. Intraparticle diffusion was the main adsorption mechanism for AC. PEHA contributed to adsorption by means of coordination on amino groups. Release was based on protons fostering both a cation exchange mechanism and protonation. The investigated materials confirmed their potential suitability to be employed in real processes on WEEE at the industrial level.

RIF1 is a multifunctional protein that plays key roles in the regulation of DNA processing. During repair of DNA double-strand breaks (DSBs), RIF1 functions in the 53BP1-Shieldin pathway that inhibits resection of DNA ends to modulate the cellular decision on which repair pathway to engage. Under conditions of replication stress, RIF1 protects nascent DNA at stalled replication forks from degradation by the DNA2 nuclease. How these RIF1 activities are regulated at the post-translational level has not yet been elucidated. Here, we identified a cluster of conserved ATM/ATR consensus SQ motifs within the intrinsically disordered region (IDR) of mouse RIF1 that are phosphorylated in proliferating B lymphocytes. We found that phosphorylation of the conserved IDR SQ cluster is dispensable for the inhibition of DSB resection by RIF1, but is essential to counteract DNA2-dependent degradation of nascent DNA at stalled replication forks. Therefore, our study identifies a key molecular feature that enables the genome-protective function of RIF1 during DNA replication stress., Competing Interests: SB, JA, TS, DS, WZ, OP, SH, AR, DR, PM, BC, AD, MD No competing interests declared, MA is affiliated with Tacalyx GmbH. The author has no financial interests to declare, JG is affiliated with Adrestia Therapeutics Ltd. The author has no financial interests to declare, (© 2022, Balasubramanian et al.)

This work deals with the development of graphene oxide (GO)-based self-assembling membranes as possible innovative proton conductors to be used in polymer electrolyte membrane fuel cells (PEMFCs). Nowadays, the most adopted electrolyte is Chemours' Nafion; however, it reveals significant deficiencies such as strong dehydration at high temperature and low humidity, which drastically reduces its proton conductivity. The presence of oxygenated moieties in the GO framework makes it suitable for functionalization, which is required to enhance the promising, but insufficient, proton-carrying features of GO. In this study, sulfonic acid groups (-SO 3 H) that should favor proton transport were introduced in the membrane structure via a reaction between GO and concentrated sulfuric acid. Six acid-to-GO molar ratios were adopted in the synthesis procedure, giving rise to final products with different sulfonation degrees. All the prepared samples were characterized by means of TGA, ATR-FTIR and Raman spectroscopy, temperature-dependent XRD, SEM and EDX, which pointed out morphological and microstructural changes resulting from the functionalization stage, confirming its effectiveness. Regarding functional features, electrochemical impedance spectroscopy (EIS) as well as measurements of ion exchange capacity (IEC) were carried out to describe the behavior of the various samples, with pristine GO and commercial Nafion ® 212 used as reference. EIS tests were performed at five different temperatures (20, 40, 60, 80 and 100 °C) under high (95%) and medium (42%) relative humidity conditions. Compared to both GO and Nafion ® 212, the sulfonated specimens demonstrate an increase in the number of ion-carrying groups, as proved by both IEC and EIS tests, which reveal the enhanced proton conductivity of these novel membranes. Specifically, an acid-to-GO molar ratio of 10 produces a six-fold improvement of IEC (4.23 meq g -1 ) with respect to pure GO (0.76 meq g -1 ), while a maximum eight-fold improvement (5.72 meq g -1 ) is achieved in SGO-15.

Leigh syndrome (LS) is a severe manifestation of mitochondrial disease in children and is currently incurable. The lack of effective models hampers our understanding of the mechanisms underlying the neuronal pathology of LS. Using patient-derived induced pluripotent stem cells and CRISPR/Cas9 engineering, we developed a human model of LS caused by mutations in the complex IV assembly gene SURF1. Single-cell RNA-sequencing and multi-omics analysis revealed compromised neuronal morphogenesis in mutant neural cultures and brain organoids. The defects emerged at the level of neural progenitor cells (NPCs), which retained a glycolytic proliferative state that failed to instruct neuronal morphogenesis. LS NPCs carrying mutations in the complex I gene NDUFS4 recapitulated morphogenesis defects. SURF1 gene augmentation and PGC1A induction via bezafibrate treatment supported the metabolic programming of LS NPCs, leading to restored neuronal morphogenesis. Our findings provide mechanistic insights and suggest potential interventional strategies for a rare mitochondrial disease.

Immunoglobulin (Ig) class switch recombination (CSR) is the process occurring in mature B cells that diversifies the effector component of antibody responses. CSR is initiated by the activity of the B cell-specific enzyme activation-induced cytidine deaminase (AID), which leads to the formation of programmed DNA double-strand breaks (DSBs) at the Ig heavy chain (Igh) locus. Mature B cells use a multilayered and complex regulatory framework to ensure that AID-induced DNA breaks are channeled into productive repair reactions leading to CSR, and to avoid aberrant repair events causing lymphomagenic chromosomal translocations. Here, we review the DNA repair pathways acting on AID-induced DSBs and their functional interplay, with a particular focus on the latest developments in their molecular composition and mechanistic regulation., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The establishment of protective humoral immunity is dependent on the ability of mature B cells to undergo antibody gene diversification while adjusting to the physiological stressors induced by activation with the antigen. Mature B cells diversify their antibody genes by class switch recombination (CSR) and somatic hypermutation (SHM), which are both dependent on efficient induction of activation-induced cytidine deaminase (AID). Here, we identified PDGFA-associated protein 1 (Pdap1) as an essential regulator of cellular homeostasis in mature B cells. Pdap1 deficiency leads to sustained expression of the integrated stress response (ISR) effector activating transcription factor 4 (Atf4) and induction of the ISR transcriptional program, increased cell death, and defective AID expression. As a consequence, loss of Pdap1 reduces germinal center B cell formation and impairs CSR and SHM. Thus, Pdap1 protects mature B cells against chronic ISR activation and ensures efficient antibody diversification by promoting their survival and optimal function., Competing Interests: Disclosures: The authors declare no competing interests exist., (© 2020 Delgado-Benito et al.)