The scaffold protein SLX4/FANCP plays a conserved role in early steps of homologous recombination DNA repair

Both in yeast and mammals, the scaffold protein SLX4/FANCP has been implicated in late steps of homologous recombination DNA repair, delivering the structure specific nucleases MUS81, SLX1 and XPF/RAD1 onto DNA repair intermediates (such as joint molecules and 3\u2019 non homologous DNA flap). Working with the model organism S. cerevisiae, we showed that Slx4 competes with the 53BP1-ortholog Rad9 for DSB end binding, favoring DNA end resection and homologous recombination repair. To investigate a possible conservation of the pathway, we exploited the AsiSI restriction enzyme and Cas9-based systems to study SLX4 role in controlling DSB resection in U2OS human osteosarcoma cells and FANCP patient derived fibroblasts. We also analyzed homologous recombination DNA repair through standard GFP reporter cassette assays and immunofluorescence foci of specific factors. The obtained results indicate that down regulation of SLX4/FANCP limits DSB resection and repair, supporting an important conserved SLX4/FANCP role in early steps of homologous recombination DNA repair, independently of the nucleases MUS81 and XPF

The web-based simulation and information service for multi-hazard impact chains. Design document.

The overall objective of the PARATUS project and the platform is the co-development of a web-based simulation and information service for first and second responders and other stakeholders to evaluate the impact chains of multi-hazard events with particular emphasis on cross-border and cascading impacts. This deliverable provides a first impression of the platform and its components. A central theme in the PARATUS project is the co-development of the tools with stakeholders. The central stakeholders within the four applications case studies are therefore full project partners. They will be directly involved in the development of the platform. We foresee that the PARATUS Platform will have two major blocks: an information service that provides static information (or regularly updated information) and simulation service, which is a dynamic component where stakeholders can interactively work with the tools in the platform. The PARATUS will further make sure that documentation (e.g., software accompanying documentation) is also publicly available via the project website1 and other trusted repositories. The deliverable 4.1 was submitted to the European Commission on 31/07/2023 and is waiting for approval by the Research Executive Agency. Therefore, this current version may not represent the final version of the deliverable

A qPCR-based protocol to quantify DSB resection

The nucleolytic degradation of the 5\u2032-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

SLX4/FANCP controls checkpoint signalling and DNA resection at double-strand breaks

The DNA damage checkpoint pathway is activated in response to DNA lesions and replication stress to preserve genome integrity. However, hyper-activation of this surveillance system is detrimental to the cell, because it might prevent cell cycle re-start after repair, which may also lead to senescence. Using Saccharomyces cerevisiae as a model system, we recently showed that the scaffold proteins Slx4 and Rtt107 limit checkpoint signalling at a persistent double-strand DNA break (DSB) and at uncapped telomeres. We found that Slx4 is recruited within a few kilobases of an irreparable DSB, through the interaction with Rtt107 and the multi-BRCT domain scaffold Dpb11. In the absence of Slx4 or Rtt107, Rad9 binding near the irreparable DSB is increased, leading to robust checkpoint signalling and slower nucleolytic degradation of the 5\u2032 strand. We are currently investigating SLX4/FANCP role in DSB resection and checkpoint signalling in human cells. Our preliminary results suggest that SLX4 silencing leads to slower DSB resection in U2OS cell line stably expressing the inducible restriction enzyme AsiSI. Moreover, SLX4 may play a role in adaptation to the DNA damage checkpoint arrest in human cells, similarly to our results obtained in yeast. In fact, following ionizing radiation\u2013induced G2 checkpoint, a small percentage of U2OS cells enter mitosis (monitored as appearance of phosphorylated histone H3 foci) with \u3b3-H2AX foci, a marker for unrepaired DNA double-strand breaks. Exit from the G2 checkpoint is delayed depleting SLX4. Our study sheds new light on the molecular mechanism that coordinates the processing and repair of DSBs with DNA damage checkpoint signalling, preserving genome integrity. Since mutations in human SLX4 lead to Fanconi anemia, a genetic disorder associated with high checkpoint marker activation, which could be a cause of bone marrow failure, our results might get new insight into the pathogenesis of the disease

Clinical profile and outcome of acute encephalitis syndrome (AES) patients treated in College of Medical Sciences-Teaching Hospital

Objective:&nbsp;Acute encephalitis syndrome is a cause of significant morbidity and mortality in Nepal. Although Japanese encephalitis virus (JEV) was thought to be a major cause for acute encephalitis syndrome, more non-Japanese encephalitis virus cases are reported. The outcome of patients with acute encephalitis syndrome is variable. Our study was designed to study the clinical profile and outcome of patients with acute encephalitis syndrome managed in tertiary care center in central Nepal. Methods:&nbsp;The record of patients admitted with diagnosis of acute encephalitis syndrome,from January 2010 to December 2010 in College of Medical Sciences-Teaching Hospital (CMS-TH) was reviewed. They were classified clinically as meningitis, encephalitis and meningoencephalitis. The clinical details and reports of the patients were recorded and analyzed. Results:&nbsp;Total of 85 cases of meningitis and encephalitis were identified. Mean age was 19.18 years. Fifty-six (65.9%) patients were males and 29 (34.1%) were females. Sixty (70.58%) patients had meningitis, 8 (9.41%) had encephalitis, and 17 (20.0%) had meningoencephalitis. JE serology was positive in 4 patients (4.7%). Seventy-two (84.7%) patients made full recovery and were discharged from hospital. Thirteen (15.3%) patients left against medical advice (LAMA). Conclusion:&nbsp;Acute encephalitis syndrome is still a major public health problem in Nepal. Few of these patients have Japanese Encephalitis. There is a trend towards improved outcome because of availability of improved health services. However, financial constraint remains a challenge in management of acute encephalitis syndrome. Journal of College of Medical Sciences-Nepal, 2013, Vol-9, No-2, 31-37 DOI: http://dx.doi.org/10.3126/jcmsn.v9i2.9685</p

Mode Sensitivity Analysis of Subwavelength Grating Slot Waveguides

In this paper, we investigate the mode sensitivity (S-mode) of subwavelength grating slot (SWGS) waveguides. S-mode is an important parameter in various waveguide-based photonic circuits such as sensors, modulators, and thermally-controlled devices. It is a measure of the sensitivity of the waveguide effective index towards the refractive index perturbations in the cladding medium. The SWGS waveguide exhibits high mode sensitivity, as it combines sensitivity enhancement features of both slot and subwavelength grating waveguides. Finite-difference time-domain simulations are performed for the analysis, design, and optimization of the hybrid structure. The SWGS waveguide is incorporated into a Mach-Zehnder interferometer and fabricated on a silicon-on-insulator platform for the experimental estimation of S-mode. The measured S-mode value of 79% is consistent with the theoretical prediction of 83%

Development of a Platform for the Generation, Visualisation and Quantification of Disaster Impact Chains

There is a lack of publicly available tools for first and second responders and local authorities to evaluate the impact chains of multi-hazard events, with cascading hazards that result in cascading impacts. Within the framework of the EU HORIZON Europe PARATUS project, we are developing an open and online, user-centered methodology and tool for systemic risk assessment, co-designed with stakeholders and addressing physical, socio-economic, and environmental aspects. It will consist of guidelines and lessons learned, links to datasets and tools for multi-hazard risk assessments that consider future scenarios and adaptation measures. We foresee that the PARATUS Platform will have two major blocks: an information service that provides static information (or regularly updated information) and a simulation service, which is a dynamic component where stakeholders can interactively work with the tools in the platform. Based on the analysis of impact chains for historical disaster events we will develop a WIKI with standardized impact chains. These form the basis for the simulation of future events, with particular emphasis on cross-border and cascading impacts. The simulation platform will consists of tools that are dynamic and interactive, and which support the stakeholders to define, analyze and quantify the impact of multi-hazard events, now and under different future scenarios, and to evaluate which risk reduction options would be most suitable