Noradrenaline release from the locus coeruleus shapes stress-induced hippocampal gene expression.

Exposure to an acute stressor triggers a complex cascade of neurochemical events in the brain. However, deciphering their individual impact on stress-induced molecular changes remains a major challenge. Here, we combine RNA sequencing with selective pharmacological, chemogenetic, and optogenetic manipulations to isolate the contribution of the locus coeruleus-noradrenaline (LC-NA) system to the acute stress response in mice. We reveal that NA release during stress exposure regulates a large and reproducible set of genes in the dorsal and ventral hippocampus via ß-adrenergic receptors. For a smaller subset of these genes, we show that NA release triggered by LC stimulation is sufficient to mimic the stress-induced transcriptional response. We observe these effects in both sexes, and independent of the pattern and frequency of LC activation. Using a retrograde optogenetic approach, we demonstrate that hippocampus-projecting LC neurons directly regulate hippocampal gene expression. Overall, a highly selective set of astrocyte-enriched genes emerges as key targets of LC-NA activation, most prominently several subunits of protein phosphatase 1 (Ppp1r3c, Ppp1r3d, Ppp1r3g) and type II iodothyronine deiodinase (Dio2). These results highlight the importance of astrocytic energy metabolism and thyroid hormone signaling in LC-mediated hippocampal function and offer new molecular targets for understanding how NA impacts brain function in health and disease.

Direct R-Loop Visualization on Genomic DNA by Native Automated Electron Microscopy.

R-loop are physiologically present on genomic DNA of different organisms and play important roles in genome regulation. However, an increase in their abundance and/or size has been suggested to interfere with the DNA replication process, contributing to genome instability. Most available approaches to monitor R-loops are based on antibodies/enzymes that cannot effectively distinguish R-loops from DNA-RNA hybrids and assess R-loop size and frequency in a population of molecules. Electron microscopy has successfully allowed single-molecule visualization of DNA replication and repair intermediates, uncovering key architectural modifications in DNA, induced by genotoxic stress or by the associated cellular response. Here, we describe recent modifications of this visualization workflow to implement partial automation of image acquisition and analysis. Coupling this refined workflow with sample preparation procedures that protect R-loop stability allows for direct visualization of R-loop structures on genomic DNA, independently from probes. Combining single-molecule information and DNA content assessment, this approach provides direct estimations of R-loop frequency, size, and burden on genomic DNA.

Novel computational analysis of large transcriptome datasets identifies sets of genes distinguishing chronic obstructive pulmonary disease from healthy lung samples.

Chronic obstructive pulmonary disease (COPD) kills over three million people worldwide every year. Despite its high global impact, the knowledge about the underlying molecular mechanisms is still limited. In this study, we aimed to extend the available knowledge by identifying a small set of COPD-associated genes. We analysed different publicly available gene expression datasets containing whole lung tissue (WLT) and airway epithelium (AE) samples from over 400 human subjects for differentially expressed genes (DEGs). We reduced the resulting sets of 436 and 663 DEGs using a novel computational approach that utilises a random depth-first search to identify genes which improve the distinction between COPD patients and controls along the first principle component of the data. Our method identified small sets of 10 and 15 genes in the WLT and AE, respectively. These sets of genes significantly (p < 10-20) distinguish COPD patients from controls with high fidelity. The final sets revealed novel genes like cysteine rich protein 1 (CRIP1) or secretoglobin family 3A member 2 (SCGB3A2) that may underlie fundamental molecular mechanisms of COPD in these tissues.

Personalized medicine for patients with COPD: where are we?

Chronic airflow limitation is the common denominator of patients with chronic obstructive pulmonary disease (COPD). However, it is not possible to predict morbidity and mortality of individual patients based on the degree of lung function impairment, nor does the degree of airflow limitation allow guidance regarding therapies. Over the last decades, understanding of the factors contributing to the heterogeneity of disease trajectories, clinical presentation, and response to existing therapies has greatly advanced. Indeed, diagnostic assessment and treatment algorithms for COPD have become more personalized. In addition to the pulmonary abnormalities and inhaler therapies, extra-pulmonary features and comorbidities have been studied and are considered essential components of comprehensive disease management, including lifestyle interventions. Despite these advances, predicting and/or modifying the course of the disease remains currently impossible, and selection of patients with a beneficial response to specific interventions is unsatisfactory. Consequently, non-response to pharmacologic and non-pharmacologic treatments is common, and many patients have refractory symptoms. Thus, there is an ongoing urgency for a more targeted and holistic management of the disease, incorporating the basic principles of P4 medicine (predictive, preventive, personalized, and participatory). This review describes the current status and unmet needs regarding personalized medicine for patients with COPD. Also, it proposes a systems medicine approach, integrating genetic, environmental, (micro)biological, and clinical factors in experimental and computational models in order to decipher the multilevel complexity of COPD. Ultimately, the acquired insights will enable the development of clinical decision support systems and advance personalized medicine for patients with COPD.

ATR-Mediated Global Fork Slowing and Reversal Assist Fork Traverse and Prevent Chromosomal Breakage at DNA Interstrand Cross-Links.

Interstrand cross-links (ICLs) are toxic DNA lesions interfering with DNA metabolism that are induced by widely used anticancer drugs. They have long been considered absolute roadblocks for replication forks, implicating complex DNA repair processes at stalled or converging replication forks. Recent evidence challenged this view, proposing that single forks traverse ICLs by yet elusive mechanisms. Combining ICL immunolabeling and single-molecule approaches in human cells, we now show that ICL induction leads to global replication fork slowing, involving forks not directly challenged by ICLs. Active fork slowing is linked to rapid recruitment of RAD51 to replicating chromatin and to RAD51/ZRANB3-mediated fork reversal. This global modulation of fork speed and architecture requires ATR activation, promotes single-fork ICL traverse-here, directly visualized by electron microscopy-and prevents chromosomal breakage by untimely ICL processing. We propose that global fork slowing by remodeling provides more time for template repair and promotes bypass of residual lesions, limiting fork-associated processing.