Utilization of a deoxynucleoside diphosphate substrate by HIV reverse transcriptase

Background: Deoxynucleoside triphosphates (dNTPs) are the normal substrates for DNA sysnthesis is catalyzed by polymerases such as HIV-1 reverse transcriptase (RT). However, substantial amounts of deoxynucleoside diphosphates (dNDPs) are also present in the cell. Use of dNDPs in HIV-1 DNA sysnthesis could have significant implications for the efficacy of nucleoside RT inhibitors such as AZT which are first line therapeutics fro treatment of HIV infection. Our earlier work on HIV-1 reverse transcriptase (RT) suggested that the interaction between the γ phosphate of the incoming dNTP and RT residue K65 in the active site is not essential for dNTP insertion, implying that this polymerase may be able to insert dNPs in addition to dNTPs. Methodology/Principal Findings: We examined the ability of recombinant wild type (wt) and mutant RTs with substitutions at residue K65 to utilize a dNDP substrate in primer extension reactions. We found that wild type HIV-1 RT indeed catalyzes incorporation of dNDP substrates whereas RT with mutations of residue K645 were unable to catalyze this reaction. Wild type HIV-1 RT also catalyzed the reverse reaction, inorganic phosphate-dependent phosphorolysis. Nucleotide-mediated phosphorolytic removal of chain-terminating 3′-terminal nucleoside inhibitors such as AZT forms the basis of HIV-1 resistance to such drugs, and this removal is enhanced by thymidine analog mutations (TAMs). We found that both wt and TAM-containing RTs were able to catalyze Pi-mediated phosphorolysis of 3′-terminal AZT at physiological levels of Pi with an efficacy similar to that for ATP-dependent AZT-excision. Conclusion: We have identified two new catalytic function of HIV-1 RT, the use of dNDPs as substrates for DNA synthesis, and the use of Pi as substrate for phosphorolytic removal of primer 3′-terminal nucleotides. The ability to insert dNDPs has been documented for only one other DNA polymerase The RB69 DNA polymerase and the reverse reaction employing inorganic phosphate has not been documented for any DNA polymerase. Importantly, our results show that Pi-mediated phosphorolysis can contribute to AZT resistance and indicates that factors that influence HIV resistance to AZT are more complex than previously appreciated. © 2008 Garforth et al

Compacting XML Structures Using a Dynamic Labeling Scheme

Abstract. Due to the growing popularity of XML as a data exchange and storage format, the need to develop efficient techniques for stor-ing and querying XML documents has emerged. A common approach to achieve this is to use labeling techniques. However, their main prob-lem is that they either do not support updating XML data dynamically or impose huge storage requirements. On the other hand, with the ver-bosity and redundancy problem of XML, which can lead to increased cost for processing XML documents, compaction of XML documents has be-come an increasingly important research issue. In this paper, we propose an approach called CXDLS combining the strengths of both, labeling and compaction techniques. Our approach exploits repetitive consecu-tive subtrees and tags for compacting the structure of XML documents by taking advantage of the ORDPATH labeling scheme. In addition it stores the compacted structure and the data values separately. Using our proposed approach, it is possible to support efficient query and update processing on compacted XML documents and to reduce storage space dramatically. Results of a comprehensive performance study are provided to show the advantages of CXDLS.

ENVIRONMENTAL AND AGRIFOOD CULTURAL TRADITION PRESERVATION AS PART OF RURAL TOURISM. A SYSTEMATIC LITERATURE REVIEW IN ROMANIA

The terms rural tourism and agritourism can be considered synonymous, but there are differences even though they share certain identifying elements. Agritourism appeared as a solution to in-crease the income of rural households, by capitalizing on their economic potential, however the main activity remained agriculture. The main purpose of this research is to evaluate the situation of agritourism in Romania approached from different perspectives such as: rural development, traditional products, and landscape integrations. The methodology used in this study is a systematic literature review where the search process was carried out using keywords such as: "Agritourism in Romania" or "Rural development in Romania" or "Integration of the rural landscape in Romania". The research was conducted and filtered through the Prisma guidelines in 30 papers that were highlighted in the Results section. In concluding this research paper, we would like to say that there are many factors that should be considered to find a viable solution to help develop rural communities and simultaneously safeguarding the environment and using natural resources responsibly

The Repeatability Experiment of SIGMOD 2008

SIGMOD 2008 was the first database conference that offered to test submitters' programs against their data to verify the experiments published. This paper discusses the rationale for this effort, the community's reaction, our experiences, and advice for future similar efforts

Co-expression network of neural-differentiation genes shows specific pattern in schizophrenia

Background: Schizophrenia is a neurodevelopmental disorder with genetic and environmental factors contributing to its pathogenesis, although the mechanism is unknown due to the difficulties in accessing diseased tissue during human neurodevelopment. The aim of this study was to find neuronal differentiation genes disrupted in schizophrenia and to evaluate those genes in post-mortem brain tissues from schizophrenia cases and controls. Methods: We analyzed differentially expressed genes (DEG), copy number variation (CNV) and differential methylation in human induced pluripotent stem cells (hiPSC) derived from fibroblasts from one control and one schizophrenia patient and further differentiated into neuron (NPC). Expression of the DEG were analyzed with microarrays of post-mortem brain tissue (frontal cortex) cohort of 29 schizophrenia cases and 30 controls. A Weighted Gene Co-expression Network Analysis (WGCNA) using the DEG was used to detect clusters of co-expressed genes that werenon-conserved between adult cases and controls brain samples. Results: We identified methylation alterations potentially involved with neuronal differentiation in schizophrenia, which displayed an over-representation of genes related to chromatin remodeling complex (adjP = 0.04). We found 228 DEG associated with neuronal differentiation. These genes were involved with metabolic processes, signal transduction, nervous system development, regulation of neurogenesis and neuronal differentiation. Between adult brain samples from cases and controls there were 233 DEG, with only four genes overlapping with the 228 DEG, probably because we compared single cell to tissue bulks and more importantly, the cells were at different stages of development. The comparison of the co-expressed network of the 228 genes in adult brain samples between cases and controls revealed a less conserved module enriched for genes associated with oxidative stress and negative regulation of cell differentiation. Conclusion: This study supports the relevance of using cellular approaches to dissect molecular aspects of neurogenesis with impact in the schizophrenic brain. We showed that, although generated by different approaches, both sets of DEG associated to schizophrenia were involved with neocortical development. The results add to the hypothesis that critical metabolic changes may be occurring during early neurodevelopment influencing faulty development of the brain and potentially contributing to further vulnerability to the illness.We thank the patients, doctors and nurses involved with sample collection and the Stanley Medical Research Institute. This research was supported by either Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq #17/2008) and Fundação Carlos Chagas Filho de Amparo a Pesquisa do Estado do Rio de Janeiro (FAPERJ). MM (CNPq 304429/2014-7), ACT (FAPESP 2014/00041-1), LL (CAPES 10682/13-9) HV (CAPES) and BP (PPSUS 137270) were supported by their fellowshipsinfo:eu-repo/semantics/publishedVersio

SATCON2: Community Engagement Working Group Report

The SATCON2 Community Engagement Working Group aimed to engage a broad and diverse swath of stakeholders in dark skies and near-Earth space who are impacted by large mega-constellations of tens of thousands of low-Earth orbit (LEO) satellites, beyond professional astronomy alone. The working group consisted of 22 members across 23 time zones including professional and amateur astronomers, members of sovereign Indigenous/First Nations communities, dark-sky advocates, planetarium professionals, and environmental/ecological non-governmental organizations. We set out to work together towards a new and effective conceptual, ethical, legal, and regulatory framework for the protection and sustainability of space and the night sky as a global cultural, natural and scientific commons. Community Engagement Working Group members invested thousands of volunteer hours in working group meetings, listening sessions with impacted constituencies, numerous conversations, developing, conducting and analyzing surveys, and finalizing our results and recommendations

Shotgun Mass Spectrometry Workflow Combining IEF and LC-MALDI-TOF/TOF

We present a high throughput shotgun mass spectrometry workflow using a bidimensional peptide fractionation procedure consisting of isoelectric focusing and RP-HPLC prior to mass spectrometric analysis, with the aim of optimizing peptide separation and protein identification. As part of the workflow we used the ‘Isotope-Coded Protein Labeling’ (ICPL) method for accurate relative quantitation of protein expression. Such workflow was successfully applied to a comparative proteome analysis of schizophrenia versus healthy control brain tissues and can be an alternative to proteome researches