Chromatin epigenetics and nuclear lamina keep the nucleus in shape: Examples from natural and accelerated aging

As the repository of genetic information, the cell nucleus must protect DNA integrity from mechanical stresses. The nuclear lamina, which resides within the nuclear envelope (NE), is made up of lamins, intermediate filaments bound to DNA. The nuclear lamina provides the nucleus with the ability to deal with inward as well as outward mechanical stimuli. Chromatin, in turn, through its degrees of compaction, shares this role with the nuclear lamina, thus, ensuring the plasticity of the nucleus. Perturbation of chromatin condensation or the nuclear lamina has been linked to a plethora of biological conditions, that range from cancer and genetic diseases (laminopathies) to aging, both natural and accelerated, such as the case of Hutchinson-Gilford Progeria Syndrome (HGPS). From the experimental results accumulated so far on the topic, a direct link between variations of the epigenetic pattern and nuclear lamina structure would be suggested, however, it has never been clarified thoroughly. This relationship, instead, has a downstream important implication on nucleus shape, genome preservation, force sensing, and, ultimately, aging-related disease onset. With this review, we aim to collect recent studies on the importance of both nuclear lamina components and chromatin status in nuclear mechanics. We also aim to bring to light evidence of the link between DNA methylation and nuclear lamina in natural and accelerated aging

A Glimpse into Chromatin Organization and Nuclear Lamina Contribution in Neuronal Differentiation

During embryonic development, stem cells undergo the differentiation process so that they can specialize for different functions within the organism. Complex programs of gene transcription are crucial for this process to happen. Epigenetic modifications and the architecture of chromatin in the nucleus, through the formation of specific regions of active as well as inactive chromatin, allow the coordinated regulation of the genes for each cell fate. In this mini-review, we discuss the current knowledge regarding the regulation of three-dimensional chromatin structure during neuronal differentiation. We also focus on the role the nuclear lamina plays in neurogenesis to ensure the tethering of the chromatin to the nuclear envelop

Dicer prevents genome instability in response to replication stress

Dicer, an endoribonuclease best-known for its role in microRNA biogenesis and RNA interference pathway, has been shown to play a role in the DNA damage response and repair of double-stranded DNA breaks (DSBs) in mammalian cells. However, it remains unknown whether Dicer is also important to preserve genome integrity upon replication stress. To address this question, we focused our study on common fragile sites (CFSs), which are susceptible to breakage after replication stress. We show that inhibition of the Dicer pathway leads to an increase in CFS expression upon induction of replication stress and to an accumulation of 53BP1 nuclear bodies, indicating transmission of replication-associated damage. We also show that in absence of a functional Dicer or Drosha, the assembly into nuclear foci of the Fanconi anemia (FA) protein FANCD2 and of the replication and checkpoint factor TopBP1 in response to replication stress is impaired, and the activation of the S-phase checkpoint is defective. Based on these results, we propose that Dicer pre-vents genomic instability after replication stress, by allowing the proper recruitment to stalled forks of proteins that are necessary to maintain replication fork stability and activate the S-phase checkpoint, thus limiting cells from proceeding into mitosis with under-replicated DNA

Bypass of cell cycle arrest induced by transient DNMT1 post-transcriptional silencing triggers aneuploidy in human cells

<p>Abstract</p> <p>Background</p> <p>Aneuploidy has been acknowledged as a major source of genomic instability in cancer, and it is often considered the result of chromosome segregation errors including those caused by defects in genes controlling the mitotic spindle assembly, centrosome duplication and cell-cycle checkpoints. Aneuploidy and chromosomal instability has been also correlated with epigenetic alteration, however the molecular basis of this correlation is poorly understood.</p> <p>Results</p> <p>To address the functional connection existing between epigenetic changes and aneuploidy, we used RNA-interference to silence the <it>DNMT1 </it>gene, encoding for a highly conserved member of the DNA methyl-transferases. DNMT1 depletion slowed down proliferation of near-diploid human tumor cells (HCT116) and triggered G1 arrest in primary human fibroblasts (IMR90), by inducing p53 stabilization and, in turn, p21^waf1transactivation. Remarkably, p53 increase was not caused by DNA damage and was not observed after p14-ARF post-transcriptional silencing. Interestingly, DNMT1 silenced cells with p53 or p14-ARF depleted did not arrest in G1 but, instead, underwent DNA hypomethylation and became aneuploid.</p> <p>Conclusion</p> <p>Our results suggest that DNMT1 depletion triggers a p14ARF/p53 dependent cell cycle arrest to counteract the aneuploidy induced by changes in DNA methylation.</p

FANCD2 promotes mitotic rescue from transcription-mediated replication stress in SETX-deficient cancer cells

Replication stress (RS) is a leading cause of genome instability and cancer development. A substantial source of endogenous RS originates from the encounter between the transcription and replication machineries operating on the same DNA template. This occurs predominantly under specific contexts, such as oncogene activation, metabolic stress, or a deficiency in proteins that specifically act to prevent or resolve those transcription-replication conflicts (TRCs). One such protein is Senataxin (SETX), an RNA:DNA helicase involved in resolution of TRCs and R-loops. Here we identify a synthetic lethal interaction between SETX and proteins of the Fanconi anemia (FA) pathway. Depletion of SETX induces spontaneous under-replication and chromosome fragility due to active transcription and R-loops that persist in mitosis. These fragile loci are targeted by the Fanconi anemia protein, FANCD2, to facilitate the resolution of under-replicated DNA, thus preventing chromosome mis-segregation and allowing cells to proliferate. Mechanistically, we show that FANCD2 promotes mitotic DNA synthesis that is dependent on XPF and MUS81 endonucleases. Importantly, co-depleting FANCD2 together with SETX impairs cancer cell proliferation, without significantly affecting non-cancerous cells. Therefore, we uncovered a synthetic lethality between SETX and FA proteins for tolerance of transcription-mediated RS that may be exploited for cancer therapy

Site-Specific RNA Editing of Stop Mutations in the CFTR mRNA of Human Bronchial Cultured Cells

It is reported that about 10% of cystic fibrosis (CF) patients worldwide have nonsense (stop) mutations in the CFTR gene, which cause the premature termination of CFTR protein synthesis, leading to a truncated and non-functional protein. To address this issue, we investigated the possibility of rescuing the CFTR nonsense mutation (UGA) by sequence-specific RNA editing in CFTR mutant CFF-16HBEge, W1282X, and G542X human bronchial cells. We used two different base editor tools that take advantage of ADAR enzymes (adenosine deaminase acting on RNA) to edit adenosine to inosine (A-to-I) within the mRNA: the REPAIRv2 (RNA Editing for Programmable A to I Replacement, version 2) and the minixABE (A to I Base Editor). Immunofluorescence experiments show that both approaches were able to recover the CFTR protein in the CFTR mutant cells. In addition, RT-qPCR confirmed the rescue of the CFTR full transcript. These findings suggest that site-specific RNA editing may efficiently correct the UGA premature stop codon in the CFTR transcript in CFF-16HBEge, W1282X, and G542X cells. Thus, this approach, which is safer than acting directly on the mutated DNA, opens up new therapeutic possibilities for CF patients with nonsense mutations

Aneuploid IMR90 cells induced by depletion of pRB, DNMT1 and MAD2 show a common gene expression signature.

Chromosome segregation defects lead to aneuploidy which is a major feature of solid tumors. How diploid cells face chromosome mis-segregation and how aneuploidy is tolerated in tumor cells are not completely defined yet. Thus, an important goal of cancer genetics is to identify gene networks that underlie aneuploidy and are involved in its tolerance. To this aim, we induced aneuploidy in IMR90 human primary cells by depleting pRB, DNMT1 and MAD2 and analyzed their gene expression profiles by microarray analysis. Bioinformatic analysis revealed a common gene expression profile of IMR90 cells that became aneuploid. Gene Set Enrichment Analysis (GSEA) also revealed gene-sets/pathways that are shared by aneuploid IMR90 cells that may be exploited for novel therapeutic approaches in cancer. Furthermore, Protein-Protein Interaction (PPI) network analysis identified TOP2A and KIF4A as hub genes that may be important for aneuploidy establishment

Autoeficacia en las prácticas pedagógicas de docentes en formación durante la pandemia

The objective of this article is to analyze the teaching self-efficacy in pedagogy students from two universities in southern Chile who were carrying out intermediate and professional practice in the context of the COVID-19 pandemic. The research was approached through the quantitative paradigm and had a cross-sectional character with a descriptive scope. A non-probabilistic sample of 150 teachers in training was selected. The results show that students feel capable of teaching all students in adverse contexts. It is concluded that, given the health emergency, teachers in training increased teaching self-efficacy, with virtual teaching being a time of intense professional learning.El presente artículo tiene como objetivo analizar la autoeficacia docente en estudiantes de pedagogía de dos universidades del sur de Chile que se encontraban realizando la práctica intermedia y profesional en contexto de pandemia por Covid-19. La investigación se abordó mediante el paradigma cuantitativo y tuvo un carácter transversal con un alcance descriptivo. Se seleccionó una muestra no probabilística de 150 docentes en formación. Los resultados evidencian que los participantes se sienten capaces de enseñar a los estudiantes en contextos adversos. Se concluye que, dada la situación de emergencia sanitaria, los profesores en formación incrementaron la autoeficacia docente, siendo la enseñanza en modalidad virtual un tiempo de intenso aprendizaje profesional