Endogenously induced DNA double strand breaks arise in heterochromatic DNA regions and require ataxia telangiectasia mutated and Artemis for their repair

Ataxia telangiectasia (ATM) mutated and Artemis, the proteins defective in ataxia telangiectasia and a class of Radiosensitive-Severe Combined Immunodeficiency (RS-SCID), respectively, function in the repair of DNA double strand breaks (DSBs), which arise in heterochromatic DNA (HC-DSBs) following exposure to ionizing radiation (IR). Here, we examine whether they have protective roles against oxidative damage induced and/or endogenously induced DSBs. We show that DSBs generated following acute exposure of G0/G1 cells to the oxidative damaging agent, tert-butyl hydroperoxide (TBH), are repaired with fast and slow components of similar magnitude to IR-induced DSBs and have a similar requirement for ATM and Artemis. Strikingly, DSBs accumulate in ATM−/− mouse embryo fibroblasts (MEFs) and in ATM or Artemis-defective human primary fibroblasts maintained for prolonged periods under confluence arrest. The accumulated DSBs localize to HC-DNA regions. Collectively, the results provide strong evidence that oxidatively induced DSBs arise in HC as well as euchromatic DNA and that Artemis and ATM function in their repair. Additionally, we show that Artemis functions downstream of ATM and is dispensable for HC-relaxation and for pKAP-1 foci formation. These findings are important for evaluating the impact of endogenously arising DNA DSBs in ATM and Artemis-deficient patients

Design and Optimization of a Compact Low-Cost Optical Particle Sizer

Thesis (Master's)--University of Washington, 2016-12Determining particulate matter (PM) concentrations in ambient air is of major importance in applications of aerosol research; personal exposure assessments, industrial particle monitoring, and air quality studies. Optical particle counters (OPCs) measure the elastic light scattering of individual particles and provide time and size-resolved PM number concentrations. They are common due to their simplicity and low-cost. However, many of them suffer from non-monotonic size dependence of scattered light intensity and its variability with changing the complex refractive index (CRI) of particles. This weakness is particularly common in portable low-cost OPCs. This contribution describes the process of designing, validating, and testing an OPC for size measurements of aerosols. The proposed device is characterized by four main principles; low sensitivity to variations in the CRI of particles, accurate sizing, compactness, and low-cost. The design utilizes small form factor low-cost components (total cost < $100) and measures less than 45 x 25 x 15mm (L, W, H) in size. An optimization methodology is defined and used to determine the optimal angular range for collection of scattered light. An adjustable experimental setup was used to validate the numerical findings and to test the performance of the optimized angular range in comparison to two equally sized angular ranges, commonly employed in OPCs. The experiments used six different spherical monodisperse particles of known size and CRI; PSL (n = 1.61), alumina (n = 1.78), and silica (n = 1.53); 2 and 4 μm in diameter. The PSL particles were used for calibration before the device was exposed to particles with different CRIs. The experimental response was in good agreement with the numerical calculations overall. The average sizing error was 6.87% for the optimal angular range, compared to 32.21% and 25.45% for the alternatives. The results show clearly that the optimal angular range is effective in eliminating the ambiguity that is commonly present when OPCs are used in the field. The findings were consistent across the two sizes and all CRIs