Exploiting the Gal4/UAS System as Plant Orthogonal Molecular Toolbox to Control Reporter Expression in Arabidopsis Protoplasts

The ability of protein domains to fold independently from the rest of the polypeptide is the principle governing the generation of fusion proteins with customized functions. A clear example is the split transcription factor system based on the yeast GAL4 protein and its cognate UAS enhancer. The rare occurrence of the UAS element in the transcriptionally sensitive regions of the Arabidopsis genome makes this transcription factor an ideal orthogonal platform to control reporter induction. Moreover, heterodimeric transcriptional complexes can be generated by exploiting posttranslational modifications hampering or promoting the interaction between GAL4-fused transcriptional partners, whenever this leads to the reconstitution of a fully functional GAL4 factor. The assembly of multiple engineered proteins into a synthetic transcriptional complex requires preliminary testing, before its components can be stably introduced into the plant genome. Mesophyll protoplast transformation represents a fast and reliable technique to test and optimize synthetic regulatory modules. Remarkable properties are the possibility to transform different combinations of plasmids (co-transformation) and the physiological resemblance of these isolated cells with the original tissue. Here we describe an extensive protocol to produce and exploit Arabidopsis mesophyll protoplasts to investigate the transcriptional output of GAL4/UAS-based complexes that are sensitive to posttranslational protein modifications

LANDSLIDE SUSCEPTIBILITY ASSESSMENT: SOIL MOISTURE MONITORING DATA PROCESSED BY AN AUTOMATIC PROCEDURE IN GIS FOR 3D DESCRIPTION OF THE SOIL SHEAR STRENGTH

Abstract. Slope stability is strongly influenced by soil hydraulic conditions. Considering rain-triggered shallow landslides, the stability can be markedly influenced by the propagation of the saturation front inside the unsaturated zone. Soil shear strength varies in the vadose zone depending on the type of soil and the variations of soil moisture. Monitoring of the unsaturated zone can be done by measuring volumetric water content using low-cost instrumentation, such as capacitive sensors that are easy to manage and provide data in near-real time. For a proper soil moisture assessment a laboratory soil-specific calibration of the sensors is recommended. Knowing the soil water content, the suction parameter can be estimated by a Water Retention Curve (WRC), and consequently the soil shear strength in unsaturated conditions is evaluated. Several models are already proposed for shallow landslide susceptibility evaluation, also in FOSS GIS environment. However, these models do not usually consider the soil shear strength in unsaturated conditions, even if it is crucial, especially in the case of shallow landslides. A procedure that allows the estimate of the soil shear strength starting from soil moisture monitoring data (from sensor networks or satellite-derived map) is here presented. Moreover, preliminary results relative to a case study (i.e. the landslide of Ceriana-Mainardo in Italy) are shown. The proposed procedure could be integrated into existing models for landslide susceptibility assessment and also for the emergency management

Research enrichment: evaluation of structured research in the curriculum for dental medicine students as part of the vertical and horizontal integration of biomedical training and discovery

<p>Abstract</p> <p>Background</p> <p>Research programs within medical and dental schools are important vehicles for biomedical and clinical discovery, serving as effective teaching and learning tools by providing situations in which predoctoral students develop problem-solving and critical-thinking skills. Although research programs at many medical and dental schools are well-established, they may not be well integrated into the predoctoral curriculum to effectively support the learning objectives for their students.</p> <p>Methods</p> <p>A series of structured seminars, incorporating faculty research, was designed for first-year dental students at the University of Nevada, Las Vegas, School of Dental Medicine to reinforce and support the concepts and skills taught in concurrent courses. A structured research enrichment period was also created to facilitate student engagement in active research using faculty and student curricular release time. Course evaluations and surveys were administered to gauge student perceptions of the curricular integration of research, the impact of these seminars on recruitment to the research program, and overall levels of student satisfaction with research enrichment.</p> <p>Results</p> <p>The analysis of course surveys revealed that students perceived the research-containing seminars effectively illustrated concepts, were logically sequenced, and were well-integrated into their curriculum. In addition, analysis of surveys revealed that the Integration Seminar courses motivated students to engage in research enrichment. Finally, this analysis provided evidence that students were very satisfied with their overall learning experience during research enrichment.</p> <p>Conclusion</p> <p>Curricular integration is one method of improving the teaching and learning of complicated and inter-related concepts, providing an opportunity to incorporate research training and objectives into traditionally separate didactic courses. Despite the benefits of curricular integration, finding the most appropriate points of integration, obtaining release time for curricular development and for research engagement, and funding predoctoral student research remain issues to be addressed in ways that reflect the character of the faculty and the goals of each institution.</p

Prognostic Implications of the Complement Protein C1q in Gliomas

The contribution of the complement system in the pathophysiology of brain cancers has been recently considered in light of its well-known involvement in carcinogenesis. Complement system represents an important component of the inflammatory response, which acts as a functional bridge between the innate and adaptive immune response. C1q, the first recognition subcomponent of the complement classical pathway, has recently been shown to be involved in a range of pathophysiological functions that are not dependent on complement activation. C1q is expressed in the microenvironment of various types of human tumors, including melanoma, prostate, mesothelioma, and ovarian cancers, where it can exert a protective or a harmful effect on cancer progression. Despite local synthesis of C1q in the central nervous system, the involvement of C1q in glioma pathogenesis has been poorly investigated. We, therefore, performed a bioinformatics analysis, using Oncomine dataset and UALCAN database in order to assess whether the expression of the genes encoding for the three chains of C1q (C1qA, C1qB, and C1qC) could serve as a potential prognostic marker for gliomas. The obtained results were then validated using an independent glioma cohort from the Chinese Glioma Genome Atlas datasets. Our bioinformatics analysis, coupled with immunohistochemistry and fluorescence microscopy, appears to suggest a positive correlation between higher levels of C1q expression and unfavorable prognosis in a diverse grade of gliomas

THE CLINICOPATHOLOGICAL AND PROGNOSTIC SIGNIFICANCES OF C1Q EXPRESSION IN GLIOMAS: A BIOINFORMATICS ANALYSIS

Introduction. The complement system represents an important component of the inflammatory response and acts as a functional bridge between the innate and adaptive immune response. The contribution of the complement component C1q in the pathophysiology of brain cancers has been recently considered in light of its well-known involvement in carcinogenesis. Brain malignancies arise from cells of the CNS and are classified according to the tissue of phylogenetic origin. Gliomas represent the most common and aggressive form of brain tumours in adults. They derive from glial cells that help to support the functions of the other main brain cells type, the neurons (1). These are a heterogeneous group of diseases with multiple subtypes (1, 2). Glioblastoma multiforme (GBM) is the most common and fatal form of a primary brain tumour, accounting for approximately 60% of all glioma cases (3), whereas grade-II and -III gliomas are the second most common type of glioma in adults (~30%) (3). C1q molecule, together with other complement components, can be locally produced within the CNS by microglia and astrocytes, rendering it an attractive player in primary brain tumour development (4). The role of C1q in gliomas microenvironment is still poorly characterized and it is still quite puzzling whether it exerts a beneficial or a harmful activity for cancer progression. In the present study we performed a bioinformatics analysis aimed at investigating if C1q can serve as a potential prognostic marker for gliomas. Methods. The expression levels of C1qA, C1qB and C1qC genes in gliomas were analysed using Oncomine analysis. Available genomics data from The Cancer Genome Atlas project was used for Kaplan–Meier survival analysis to generate survival probability plots, using UALCAN analysis. Results. From the analysis performed on several data- sets using Oncomine, we showed a significantly higher mRNA expression levels for C1qA, C1qB and C1qC chains were detected in gliomas (different histotypes and grades) as compared to normal brain tissue (Fig. 1). We observed a positive correlation between the mRNA expression of C1qA, C1qB and C1qC mRNA poly- peptide chains and the unfavorable prognosis only in gliomas grade-II and -III, where the survival probability is indeed reduced (P &lt;0.05) (Fig. 2). No correlation was observed in glioblastoma multiforme (Fig. 2). By immu- nohistochemical approaches we detected a high depo- sition of C1q in the tumor microenvironment of both in grade-II and -III gliomas and in GBMs examined (Fig. 3a glioma, 3b glioblastoma multiforme; 20x Magnification). Moreover, in double immunocytochemical experiments we demonstrated that CD68 positive infiltrating cells are actively synthesizing C1q in the tumor micro-envi- ronment. CD68 expression is characteristic of tumor- associated macrophages, whose enrichment in glioma has been associated with poor prognosis (5). Conclusion. In our study C1q expression was significantly correlated with poor survival probability in gliomas grade-II and -III while this is not the case for GBM. These data altogether underline how complex, multifaceted and still poorly understood is the role C1q can exert on tumor progression, and how the very same molecule can differentially affect the outcome depending on the biological context it comes to act

Health and Human Rights in Chin State, Western Burma: A Population-Based Assessment Using Multistaged Household Cluster Sampling

Sollom and colleagues report the findings from a household survey study carried out in Western Burma; they report a high prevalence of human rights violations such as forced labor, food theft, forced displacement, beatings, and ethnic persecution

Dim artificial light at night alters gene expression rhythms and growth in a key seagrass species (Posidonia oceanica)

Artificial light at night (ALAN) is a globally spreading anthropogenic stressor, affecting more than 20% of coastal habitats. The alteration of the natural light/darkness cycle is expected to impact the physiology of organisms by acting on the complex circuits termed as circadian rhythms. Our understanding of the impact of ALAN on marine organisms is lagging behind that of terrestrial ones, and effects on marine primary producers are almost unexplored. Here, we investigated the molecular and physiological response of the Mediterranean seagrass, Posidonia oceanica (L.) Delile, as model to evaluate the effect of ALAN on seagrass populations established in shallow waters, by taking advantage of a decreasing gradient of dim nocturnal light intensity (from P. oceanica, including that of shorter blue wavelengths, through the ELF3–LUX1–ZTL regulatory network, and suggested that the daily perturbation of internal clock orthologs in seagrass might have caused the recruitment of PoSEND33 and PoPSBS genes to mitigate the repercussions of a nocturnal stress on photosynthesis during the day. A long-lasting impairment of gene fluctuations in sites characterised by ALAN could explain the reduced growth of the seagrass leaves when these were transferred into controlled conditions and without lighting during the night. Our results highlight the potential contribution of ALAN to the global loss of seagrass meadows, posing questions about key interactions with a variety of other human-related stressors in urban areas, in order to develop more efficient strategies to globally preserve these coastal foundation species

Direct RNA Nanopore Sequencing of SARS-CoV-2 Extracted from Critical Material from Swabs

In consideration of the increasing prevalence of COVID-19 cases in several countries and the resulting demand for unbiased sequencing approaches, we performed a direct RNA sequencing (direct RNA seq.) experiment using critical oropharyngeal swab samples collected from Italian pa-tients infected with SARS-CoV-2 from the Palermo region in Sicily. Here, we identified the sequences SARS-CoV-2 directly in RNA extracted from critical samples using the Oxford Nanopore MinION technology without prior cDNA retrotranscription. Using an appropriate bioinformatics pipeline, we could identify mutations in the nucleocapsid (N) gene, which have been reported previously in studies conducted in other countries. In conclusion, to the best of our knowledge, the technique used in this study has not been used for SARS-CoV-2 detection previously owing to the difficulties in the extraction of RNA of sufficient quantity and quality from routine oropharyngeal swabs. Despite these limitations, this approach provides the advantages of true native RNA sequencing and does not include amplification steps that could introduce systematic errors. This study can provide novel information relevant to the current strategies adopted in SARS-CoV-2 next-generation sequencing