Monitoring immediate-early gene expression through firefly luciferase imaging of HRS/J hairless mice

BACKGROUND: Gene promoters fused to the firefly luciferase gene (luc) are useful for examining gene regulation in live transgenic mice and they provide unique views of functioning organs. The dynamics of gene expression in cells and tissues expressing luciferase can be observed by imaging this enzyme's bioluminescent oxidation of luciferin. Neural pathways involved in specific behaviors have been identified by localizing expression of immediate-early genes such as c-fos. A transgenic mouse line with luc controlled by the human c-fos promoter (fos::luc) has enabled gene expression imaging in brain slice cultures. To optimize imaging of immediate-early gene expression throughout intact mice, the present study examined fos::luc mice and a second transgenic mouse containing luc controlled by the human cytomegalovirus immediate-early gene 1 promoter and enhancer (CMV::luc). Because skin pigments and hair can significantly scatter light from underlying structures, the two transgenic lines were crossed with a hairless albino mouse (HRS/J) to explore which deep structures could be imaged. Furthermore, live anesthetized mice were compared with overdosed mice. RESULTS: Bioluminescence imaging of anesthetized mice over several weeks corresponded with expression patterns in mice imaged rapidly after a lethal overdose. Both fos::luc and CMV::luc mice showed quantifiable bright bioluminescence in ear, nose, paws, and tail whether they were anesthetized or overdosed. CMV::luc and fos::luc neonates had bioluminescence patterns similar to those of adults, although intensity was significantly higher in neonates. CMV::luc mice crossed with HRS/J mice had high expression in bone, claws, head, pancreas, and skeletal muscle, but less in extremities than haired CMV::luc mice. Imaging of brain bioluminescence through the neonatal skull was also practical. By imaging luciferin autofluorescence it was clear that substrate distribution did not restrict bioluminescence imaging to capillaries after injection. Luciferin treatment and anesthesia during imaging did not adversely affect circadian rhythms in locomotor activity. CONCLUSIONS: Imaging of gene expression patterns with luciferase can be extended from studies of live animals to rapid imaging of mice following a pentobarbital overdose before significant effects from postmortem changes occurs. Bioluminescent transgenic mice crossed with HRS/J mice are valuable for examining gene expression in deep tissues

Scenarios of land use change and impacts on ecosystem services in the Brazilian Amazon.

Large-scale and high-intensity land use changes (LUC) are intrinsically related to the loss of biodiversity, and decreased integrity of natural systems that help maintain ecosystem services (ES). Landscape-scale patterns of land use (LU) can be correlated with different levels of ecosystem integrity (EI) and consequently with the potential ES provision. Once the relations between LU patterns and EI/ES are established it is possible to predict future environmental services provision considering different LUC scenarios. The objective of this work is to present a methodological approach and preliminary results of the prediction of future impacts on ES based on LUC scenarios for the Brazilian Amazon. The methodological approach used was to integrate: (i) Ecosystem Integrity Spatial Model (EISM) based on Bayesian probabilistic distribution of evidences using a Remote Sensing dataset. The validation was based on knowledge and field controls; (ii) Correlation of EISM and ES Models: (iia) Evapotranspiration Fluxes (ET) - MODIS/MOD16; (iib) Aboveground Carbon Stocks Spatial Model (WHRC); and (iii) Amazon legal region LUC-SSPs scenarios (Adapted from the IPCC-SSPs) - Clue Model (1 km2 pixel; projected for 2050). The preliminary results were promising and showed the decrease of ecosystem integrity related with each LUC scenario considered, allowing estimates of the impacts on the ecosystem services studied: water fluxes (ET) and aboveground carbon stocks. This work is part of ROBIN Project (EU- FP7Edict.ENV.2011.2.1.4-1)

Comparative Analysis of Distribution and Abundance of West Nile and Eastern Equine Encephalomyelitis Virus Vectors in Suffolk County, New York, Using Human Population Density and Land Use/Cover Data

Five years of CDC light trap data from Suffolk County, NY, were analyzed to compare the applicability of human population density (HPD) and land use/cover (LUC) classification systems to describe mosquito abundance and to determine whether certain mosquito species of medical importance tend to be more common in urban (defined by HPD) or residential (defined by LUC) areas. Eleven study sites were categorized as urban or rural using U.S. Census Bureau data and by LUC types using geographic information systems (GISs). Abundance and percent composition of nine mosquito taxa, all known or potential vectors of arboviruses, were analyzed to determine spatial patterns. By HPD definitions, three mosquito species, Aedes canadensis (Theobald), Coquillettidia perturbans (Walker), and Culiseta melanura (Coquillett), differed significantly between habitat types, with higher abundance and percent composition in rural areas. Abundance and percent composition of these three species also increased with freshwater wetland, natural vegetation areas, or a combination when using LUC definitions. Additionally, two species, Ae. canadensis and Cs. melanura, were negatively affected by increased residential area. One species, Aedes vexans (Meigen), had higher percent composition in urban areas. Two medically important taxa, Culex spp. and Aedes triseriatus (Say), were proportionally more prevalent in residential areas by LUC classification, as was Aedes trivittatus (Coquillett). Although HPD classification was readily available and had some predictive value, LUC classification resulted in higher spatial resolution and better ability to develop location specific predictive models

Land Change Science and the STEPLand Framework : An Assessment of Its Progress

This contribution assesses a new term that is proposed to be established within Land Change Science: Spatio-TEmporal Patterns of Land ('STEPLand'). It refers to a specific workflow for analyzing land-use/land cover (LUC) patterns, identifying and modeling driving forces of LUC changes, assessing socio-environmental consequences, and contributing to defining future scenarios of land transformations. In this article, we define this framework based on a comprehensive meta-analysis of 250 selected articles published in international scientific journals from 2000 to 2019. The empirical results demonstrate that STEPLand is a consolidated protocol applied globally, and the large diversity of journals, disciplines, and countries involved shows that it is becoming ubiquitous. In this paper, the main characteristics of STEPLand are provided and discussed, demonstrating that the operational procedure can facilitate the interaction among researchers from different fields, and communication between researchers and policy makers

The MBD7 complex promotes expression of methylated transgenes without significantly altering their methylation status.

DNA methylation is associated with gene silencing in eukaryotic organisms. Although pathways controlling the establishment, maintenance and removal of DNA methylation are known, relatively little is understood about how DNA methylation influences gene expression. Here we identified a METHYL-CpG-BINDING DOMAIN 7 (MBD7) complex in Arabidopsis thaliana that suppresses the transcriptional silencing of two LUCIFERASE (LUC) reporters via a mechanism that is largely downstream of DNA methylation. Although mutations in components of the MBD7 complex resulted in modest increases in DNA methylation concomitant with decreased LUC expression, we found that these hyper-methylation and gene expression phenotypes can be genetically uncoupled. This finding, along with genome-wide profiling experiments showing minimal changes in DNA methylation upon disruption of the MBD7 complex, places the MBD7 complex amongst a small number of factors acting downstream of DNA methylation. This complex, however, is unique as it functions to suppress, rather than enforce, DNA methylation-mediated gene silencing

The Arabidopsis NADPH oxidases RbohD and RbohF display differential expression patterns and contributions during plant immunity

Plant NADPH oxidases, also known as respiratory burst oxidase homologues (RBOHs), produce reactive oxygen species (ROS) that perform a wide range of functions. RbohD and RbohF, two of the 10 Rboh genes present in Arabidopsis, are pleiotropic and mediate diverse physiological processes including the response to pathogens. We hypothesized that the spatio-temporal control of RbohD and RbohF gene expression might be critical in determining their multiplicity of functions. Transgenic Arabidopsis plants with RbohD and RbohF promoter fusions to β-glucuronidase and Luciferase reporter genes were generated. Analysis of these plants revealed a differential expression pattern for RbohD and RbohF throughout plant development and during immune responses. RbohD and RbohF gene expression was differentially modulated by pathogen-associated molecular patterns. Histochemical stains and in vivo expression analysis showed a correlation between the level of RbohD and RbohF promoter activity, H2O2 accumulation and the amount of cell death in response to the pathogenic bacterium Pseudomonas syringae pv. tomato DC3000 and the necrotrophic fungus Plectosphaerella cucumerina. A promoter-swap strategy revealed that the promoter region of RbohD was required to drive production of ROS by this gene in response to pathogens. Moreover, RbohD promoter was activated during Arabidopsis interaction with a non-virulent P. cucumerina isolate, and susceptibility tests with the double mutant rbohD rbohF uncovered a new function for these oxidases in basal resistance. Altogether, our results suggest that differential spatio-temporal expression of the Rboh genes contributes to fine-tune RBOH/NADPH oxidase-dependent ROS production and signaling in Arabidopsis immunity

The appropriateness of the localised uniform conditioning technique for high-nugget Birimian-style gold deposits

A dissertation submitted in fulfilment of the requirements for the degree of Masters of Science in Engineering to the Faculty of Engineering and the Built Environment, University of the Witwatersrand, 2017The localized uniform conditioning (LUC) technique converts conventional Uniform Conditioning (UC) grade-tonnage curves into single grade values attached to each smallest mining unit (SMU). This is achieved by ranking the SMUs within a panel in increasing order of their grade based on the local grade patterns predicted by direct kriging of the SMUs. However, the quality of this localization process will depend heavily on the validity of the predicted grade patterns. A study was undertaken to determine how valid the predicted grade patterns of a typical Birimian-style gold deposit (with high nugget effect and strong short-range variability) might be expected to be. The direct SMU kriging rankings (based on sparse data) were compared with the grade control model ranking (based on close-spaced data and the best available estimate of the deposit). The results showed a satisfactory correlation and relationship between these rankings. It was concluded that the application of the LUC technique is still useful and appropriate for this style of deposit.XL201

A study of urban heat island of Banda Aceh City, Indonesia based on land use/cover changes and land surface temperature

This article described the spatial and temporal of land surface temperature (LST) patterns in Banda Aceh City, Indonesia, in the context of urban heat island (UHI) phenomenon. Landsat imaginary in 1998 and 2018 was used in this study, which represents the conditions before and after the tsunami. Geographic Information System (GIS) and Remote Sensing (RS) technique were used for data analysis. The 1998 and 2018 LUC maps were derived from remote sensing satellite images using a supervised classification method (maximum likelihood). Both LUC maps contained five categories, namely built-up area, vegetation, water body, vacant land, and wet land. The 1998 LUC map had a kappa coefficient 0.91, while the 2018 LUC map had 0.84. It was found that the built-up area increased by 100%, while the vegetation category fell 50%. The overall mean LST in the study area increased 5.90C between 1998 and 2018, with the highest mean increase in the built-up area category. The study recommends that LST should be taken into consideration in urban planning process to realize sustainable urban development. It also emphasizes the importance of optimizing the availability of green open space to reduce UHI effects and helps in improving the quality of the urban environment.

Sexual Differentiation of Circadian Clock Function in the Adrenal Gland

Sex differences in glucocorticoid production are associated with increased responsiveness of the adrenal gland in females. However, the adrenal-intrinsic mechanisms that establish sexual dimorphic function remain ill defined. Glucocorticoid production is gated at the molecular level by the circadian clock, which may contribute to sexual dimorphic adrenal function. Here we examine sex differences in the adrenal gland using an optical reporter of circadian clock function. Adrenal glands were cultured from male and female Period2::Luciferase (PER2::LUC) mice to assess clock function in vitro in real time. We confirm that there is a pronounced sex difference in the intrinsic capacity to sustain PER2::LUC rhythms in vitro, with higher amplitude rhythms in adrenal glands collected from males than from females. Changes in adrenal PER2::LUC rhythms over the reproductive life span implicate T as an important factor in driving sex differences in adrenal clock function. By directly manipulating hormone levels in adult mice in vivo, we demonstrate that T increases the amplitude of PER2::LUC rhythms in adrenal glands of both male and female mice. In contrast, we find little evidence that ovarian hormones modify adrenal clock function. Lastly, we find that T in vitro can increase the amplitude of PER2::LUC rhythms in male adrenals but not female adrenals, which suggests the existence of sex differences in the mechanisms of T action in vivo. Collectively these results reveal that activational effects of T alter circadian timekeeping in the adrenal gland, which may have implications for sex differences in stress reactivity and stress-related disorders