Ezh2-dCas9 and KRAB-dCas9 enable engineering of epigenetic memory in a context-dependent manner.

BackgroundRewriting of the epigenome has risen as a promising alternative to gene editing for precision medicine. In nature, epigenetic silencing can result in complete attenuation of target gene expression over multiple mitotic divisions. However, persistent repression has been difficult to achieve in a predictable manner using targeted systems.ResultsHere, we report that persistent epigenetic memory required both a DNA methyltransferase (DNMT3A-dCas9) and a histone methyltransferase (Ezh2-dCas9 or KRAB-dCas9). We demonstrate that the histone methyltransferase requirement can be locus specific. Co-targeting Ezh2-dCas9, but not KRAB-dCas9, with DNMT3A-dCas9 and DNMT3L induced long-term HER2 repression over at least 50 days (approximately 57 cell divisions) and triggered an epigenetic switch to a heterochromatic environment. An increase in H3K27 trimethylation and DNA methylation was stably maintained and accompanied by a sustained loss of H3K27 acetylation. Interestingly, substitution of Ezh2-dCas9 with KRAB-dCas9 enabled long-term repression at some target genes (e.g., SNURF) but not at HER2, at which H3K9me3 and DNA methylation were transiently acquired and subsequently lost. Off-target DNA hypermethylation occurred at many individual CpG sites but rarely at multiple CpGs in a single promoter, consistent with no detectable effect on transcription at the off-target loci tested. Conversely, robust hypermethylation was observed at HER2. We further demonstrated that Ezh2-dCas9 required full-length DNMT3L for maximal activity and that co-targeting DNMT3L was sufficient for persistent repression by Ezh2-dCas9 or KRAB-dCas9.ConclusionsThese data demonstrate that targeting different combinations of histone and DNA methyltransferases is required to achieve maximal repression at different loci. Fine-tuning of targeting tools is a necessity to engineer epigenetic memory at any given locus in any given cell type

Unmanned Aircraft Systems for Archaeology Using Photogrammetry and LiDAR in Southwestern United States

Researchers can use small unmanned aircraft systems (sUAS), also known as drones, to make observations of historical sites, help interpret locations, and make new discoveries that may not be visible to the naked eye. A student team from Embry-Riddle Aeronautical University gathered data for historical site documentation in New Mexico using the DJI Phantom 4 Pro V2, DJI Mavic Pro 2, DJI M210 and DJI M600, and senseFly eBee. Utilizing these drones, student analysts were able to take the data gathered and create georectified orthomosaic images and 3D virtual objects. At Tularosa Canyon, at a site known as the Creekside Village, work aimed at imaging an amphitheater like structure (i.e., kiva) that dates back to 600 AD. The team used photogrammetry and LiDAR to determine the location of other manmade structures at the same location. Images were processed with Pix4Dmapper Pro. Team members generated LiDAR point clouds and post processed data in search of undiscovered features and structures

Exercise Is Medicine® on Campus during COVID-19: Necessary Adaptations and Continuing Importance

Exercise Is Medicine® on Campus (EIM-OC), created by the American College of Sports Medicine (ACSM), is an initiative connecting college medical providers to exercise providers to better promote physical activity (PA) among students. At the University of North Carolina at Chapel Hill (UNC), PA has been incorporated as a vital sign within the campus health medical system, in which the provider assesses the patient’s current PA. If needed, the provider refers the patient to the UNC EIM-OC team. The UNC EIM-OC initiative began in fall 2016, but it was not until spring 2020 that a fully operational system was in place, and UNC EIM-OC was awarded “gold status” by the ACSM. However, just as the system was running smoothly, the COVID-19 pandemic spread to the United States. COVID-19 and its associated social distancing restrictions have led to many changes in the EIM-OC process. With respect to EIM-OC, this commentary reports on (i) the continuing importance of the initiative during COVID-19, (ii) COVID-related adaptations to UNC EIM-OC, and (iii) recommendations for the upcoming school year

An Artificial Sweating System for Sweat Sensor Testing Applications

This research proposes a completely automated, computer-controlled fluid mixing and dispensing system, which is suitable for testing sweat sensing devices, as an alternative to requiring human trials during the development phase of a sweat sensor device. An arm mold was designed and implemented with dragon skin and pores to simulate sweating action. The relay controlled mixing tanks allow for the different concentration of fluid solutions at various rates of fluid dispensing through pores. The onboard single board computer controls a dozen electronic relays and it switches and presents an easy to use graphical user interface to allow end users to conduct the experiments with ease and not require further programming. With the recent advances in sweat sensors, this platform offers a unique way of testing sensing devices during development, allowing for researchers to focus on their design parameters one at a time before actual validation through human trials are conducted. The current device can provide sweat rates from 1 µL/min to 500 µL/min. Furthermore, concentrations of 10 mM up to 200 mM of salt concentrations were able to be repeatedly produced. In an ANOVA test with salt concentrations varying from 40–60 mM, a p-value of 0.365 shows that the concentration does not have any effect on the flow rate. Similarly, a p-value of 0.329 and 0.167 for different relative humidity and temperature shows that the system does not present a statistical difference. Lastly, when the interactions among all the factors were considered, a p-value of 0.416 clearly presents that the system performance is insensitive to different factors, thus validating the system reliability

Rumen and Serum Metabolomes in Response to Endophyte-Infected Tall Fescue Seed and Isoflavone Supplementation in Beef Steers

Fescue toxicosis impacts beef cattle production via reductions in weight gain and muscle development. Isoflavone supplementation has displayed potential for mitigating these effects. The objective of the current study was to evaluate isoflavone supplementation with fescue seed consumption on rumen and serum metabolomes. Angus steers (n = 36) were allocated randomly in a 2 × 2 factorial arrangement of treatments including endophyte-infected (E+) or endophyte-free (E−) tall fescue seed, with (P+) or without (P−) isoflavones. Steers were provided a basal diet with fescue seed for 21 days, while isoflavones were orally administered daily. Following the trial, blood and rumen fluid were collected for metabolite analysis. Metabolites were extracted and then analyzed by UPLC-MS. The MAVEN program was implemented to identify metabolites for MetaboAnalyst 4.0 and SAS 9.4 statistical analysis. Seven differentially abundant metabolites were identified in serum by isoflavone treatment, and eleven metabolites in the rumen due to seed type (p \u3c 0.05). Pathways affected by treatments were related to amino acid and nucleic acid metabolism in both rumen fluid and serum (p \u3c 0.05). Therefore, metabolism was altered by fescue seed in the rumen; however, isoflavones altered metabolism systemically to potentially mitigate detrimental effects of seed and improve animal performance

Density‑dependent winter survival of immatures in an irruptive raptor with pulsed breeding

Highly mobile predators can show strong numerical responses to pulsed resources, sometimes resulting in irruptions where large numbers of young invade landscapes at a continental scale. High production of young in irruption years may have a strong influence on the population dynamics unless immature survival is reduced compared to non-irruption years. This could occur if subordinate individuals (mainly immatures) are forced into suboptimal habitats due to density-dependent effects in irruption years. To test whether irruptive individuals had lower survival than non-irruptive ones, we combined necropsy results (N = 365) with telemetry (N = 185) from more than 20 years to record timing and causes of mortality in snowy owls (Bubo scandiacus), which irrupt into eastern North America during winter following high breeding output caused by lemming peaks in the Arctic. Mortality was more than four times higher in irruption years than non-irruption years, but only for immatures, and occurred disproportionately in early winter for immatures, but not adults. Mortality was also higher in eastern North America, where owl abundance fluctuates considerably between years, compared to core winter regions of the Arctic and Prairies where populations are more stable. Most mortality was not due to starvation, but rather associated with human activity, especially vehicle collisions. We conclude that immature snowy owls that irrupt into eastern North America are limited by density-dependent factors, such as increased competition forcing individuals to occupy risky human-altered habitats. For highly mobile, irruptive animals, resource pulses may have a limited impact on population dynamics due to low subsequent survival of breeding output during the nonbreeding season.publishedVersio

Open science in archaeology

No abstract available