Exon expression arrays as a tool to identify new cancer genes

Background: Identification of genes that are causally implicated in oncogenesis is a major goal in cancer research. An estimated 10-20% of cancer-related gene mutations result in skipping of one or more exons in the encoded transcripts. Here we report on a strategy to screen in a global fashion for such exon-skipping events using PAttern based Correlation (PAC). The PAC algorithm has been used previously to identify differentially expressed splice variants between two predefined subgroups. As genetic changes in cancer are sample specific, we tested the ability of PAC to identify aberrantly expressed exons in single samples. Principal Findings: As a proof-of-principle, we tested the PAC strategy on human cancer samples of which the complete coding sequence of eight cancer genes had been screened for mutations. PAC detected all seven exon-skipping mutants among 12 cancer cell lines. PAC also identified exon-skipping mutants in clinical cancer specimens although detection was compromised due to heterogeneous (wild-type) transcript expression. PAC reduced the number candidate genes/exons for subsequent mutational analysis by two to three orders of magnitude and had a substantial true positive rate. Importantly, of 112 randomly selected outlier exons, sequence analysis identified two novel exon skipping events, two novel base changes and 21 previously reported base changes (SNPs). Conclusions: The ability of PAC to enrich for mutated transcripts and to identify known and novel genetic changes confirms its suitability as a strategy to identify candidate cancer genes

Eco-friendly one-pot synthesis of Prussian blue-embedded magnetic hydrogel beads for the removal of cesium from water

A simple one-step approach to fabricating Prussian blue-embedded magnetic hydrogel beads (PBMHBs) was fabricated for the effective magnetic removal of radioactive cesium (Cs-137) from water. Through the simple dropwise addition of a mixed aqueous solution of iron salts, commercial PB and polyvinyl alcohol (PVA) to an ammonium hydroxide (NH4OH) solution, the formation of hydrogel beads and the encapsulation of PB in beads were achieved in one pot through the gelation of PVA with in situ-formed iron oxide nanoparticles as the cross-linker. The obtained PB-MHBs, with 43.77 weight %of PB, were stable without releasing PB for up to 2 weeks and could be effectively separated from aqueous solutions by an external magnetic field, which is convenient for the large-scale treatment of Cs-contaminated water. Detailed Cs adsorption studies revealed that the adsorption isotherms and kinetics could be effectively described by the Langmuir isotherm model and the pseudo-second-order model, respectively. Most importantly, the PB-MHBs exhibited excellent selectivity for Cs-137 in (137)Cscontaminated simulated groundwater (55 Bq/g) with a high removal efficiency (>99.5%), and the effective removal of Cs-137 from real seawater by these PB-MHBs demonstrated the excellent potential of this material for practical application in the decontamination of Cs-137-contaminated seawate

GC content around splice sites affects splicing through pre-mRNA secondary structures

<p>Abstract</p> <p>Background</p> <p>Alternative splicing increases protein diversity by generating multiple transcript isoforms from a single gene through different combinations of exons or through different selections of splice sites. It has been reported that RNA secondary structures are involved in alternative splicing. Here we perform a genomic study of RNA secondary structures around splice sites in humans (<it>Homo sapiens</it>), mice (<it>Mus musculus</it>), fruit flies (<it>Drosophila melanogaster</it>), and nematodes (<it>Caenorhabditis elegans</it>) to further investigate this phenomenon.</p> <p>Results</p> <p>We observe that GC content around splice sites is closely associated with the splice site usage in multiple species. RNA secondary structure is the possible explanation, because the structural stability difference among alternative splice sites, constitutive splice sites, and skipped splice sites can be explained by the GC content difference. Alternative splice sites tend to be GC-enriched and exhibit more stable RNA secondary structures in all of the considered species. In humans and mice, splice sites of first exons and long exons tend to be GC-enriched and hence form more stable structures, indicating the special role of RNA secondary structures in promoter proximal splicing events and the splicing of long exons. In addition, GC-enriched exon-intron junctions tend to be overrepresented in tissue-specific alternative splice sites, indicating the functional consequence of the GC effect. Compared with regions far from splice sites and decoy splice sites, real splice sites are GC-enriched. We also found that the GC-content effect is much stronger than the nucleotide-order effect to form stable secondary structures.</p> <p>Conclusion</p> <p>All of these results indicate that GC content is related to splice site usage and it may mediate the splicing process through RNA secondary structures.</p

HFE variants and the expression of iron-related proteins in breast cancer-associated lymphocytes and macrophages

Disponível em: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5264664/The association of HFE (High Iron FE) major variants with breast cancer risk and behavior has been a matter of discussion for a long time. However, their impact on the expression of iron-related proteins in the breast cancer tissue has never been addressed. In the present study, hepcidin, ferroportin 1, transferrin receptor 1 (TfR1), and ferritin expressions, as well as tissue iron deposition were evaluated in a collection of samples from breast cancers patients and analyzed according to the patients’ HFE genotype. Within the group of patients with invasive carcinoma, those carrying the p.Cys282Tyr variant in heterozygosity presented a higher expression of hepcidin in lymphocytes and macrophages than wild-type or p.His63Asp carriers. An increased expression of TfR1 was also observed in all the cell types analyzed but only in p.Cys282Tyr/p.His63Asp compound heterozygous patients. A differential impact of the two HFE variants was further noticed with the observation of a significantly higher percentage of p.Cys282Tyr heterozygous patients presenting tissue iron deposition in comparison to p.His63Asp heterozygous. In the present cohort, no significant associations were found between HFE variants and classical clinicopathological markers of breast cancer behavior and prognosis. Although limited by a low sampling size, our results provide a new possible explanation for the previously reported impact of HFE major variants on breast cancer progression, i.e., not by influencing systemic iron homeostasis but rather by differentially modulating the local cellular expression of iron-related proteins and tissue iron deposition.OM is a recipient of the PhD grant SFRH/BD/2011/78184 from Fundação para a Ciência e Tecnologia (FCT). The authors also acknowledge financial support from ICBAS/AI&NSUMIB and by national funds through FCT and Ministério da Educação e Ciência (MEC) and when applicable co-funded by FEDER funds within the partnership agreement PT2020 related with the research unit number 4293.info:eu-repo/semantics/publishedVersio

Research Priorities for Neglected Infectious Diseases in Latin America and the Caribbean Region

Dujardin, J. C. et al. 5 p.-1 tab.Global priorities for research in neglected infectious diseases (NIDs) can be assessed in different ways, but it is important to realize that regional priorities may significantly differ one from another. The region of Latin America and the Caribbean (LAC) is—along with Africa and Asia—more affected by NIDs than other regions of the world. Some of the Latin American NIDs are common to other continents, while others are very specific or disproportionately affect the Latin American region [1– 3] (Table 1). Because of its huge ecological diversity, ongoing environmental changes, and massive migrations, LAC is also a catalyst for the (re-)emergence and spreading of NIDs, both inside and outside the subcontinent. Following a colloquium on NIDs in LAC held in Lima, Peru, between 12 and 14 November 2009, a thematic workshop was organized with the support of the European Commission (EC). It involved 29 scientists (16 from the Americas, two from the Democratic Republic of Congo and India, respectively, and nine from Europe) working on different NIDs and representing several research areas from basic to applied. This report summarizes the consensus comments of the expert group after oral and written consultation. It is envisaged that this document should stimulate a debate within the scientific community and serve as a recommendation for future actions by international or regional funding agencies in the area of NIDs in LACThis work was supported by the Directorate-General for Development Cooperation of the Belgian Government (framework agreement 03, project 95502) and the European CommissionPeer reviewe

Regulation of Retention of FosB Intron 4 by PTB

One effect of stressors such as chronic drug administration is that sequence within the terminal exon of the transcription factor FosB is recognized as intronic and removed by alternative splicing. This results in an open-reading-frame shift that produces a translation stop codon and ultimately a truncated protein, termed ΔFosB. In vitro splicing assays with control and mutated transcripts generated from a fosB mini-gene construct indicated a CU-rich sequence at the 3′ end of intron 4 (I4) plays an important role in regulating fosB pre-mRNA splicing due to its binding of polypyrimidine tract binding protein (PTB). PTB binding to this sequence is dependent upon phosphorylation by protein kinase A and is blocked if the CU-rich sequence is mutated to a U-rich region. When this mutated fosB minigene is expressed in HeLa cells, the splicing efficiency of its product is increased compared to wild type. Moreover, transient transfection of PTB-1 in HeLa cells decreased the splicing efficiency of a wild type fosB minigene transcript. Depletion of PTB from nuclear extracts facilitated U2AF65 binding to wild type sequence in vitro, suggesting these proteins function in a dynamic equilibrium to modulate fosB pre-mRNA alternative splicing. These results demonstrate for the first time that phosphorylated PTB promotes intron retention and thereby silences the splicing of fosB I4

Multivariate Analysis and Visualization of Splicing Correlations in Single-Gene Transcriptomes

BACKGROUND: RNA metabolism, through 'combinatorial splicing', can generate enormous structural diversity in the proteome. Alternative domains may interact, however, with unpredictable phenotypic consequences, necessitating integrated RNA-level regulation of molecular composition. Splicing correlations within transcripts of single genes provide valuable clues to functional relationships among molecular domains as well as genomic targets for higher-order splicing regulation. RESULTS: We present tools to visualize complex splicing patterns in full-length cDNA libraries. Developmental changes in pair-wise correlations are presented vectorially in 'clock plots' and linkage grids. Higher-order correlations are assessed statistically through Monte Carlo analysis of a log-linear model with an empirical-Bayes estimate of the true probabilities of observed and unobserved splice forms. Log-linear coefficients are visualized in a 'spliceprint,' a signature of splice correlations in the transcriptome. We present two novel metrics: the linkage change index, which measures the directional change in pair-wise correlation with tissue differentiation, and the accuracy index, a very simple goodness-of-fit metric that is more sensitive than the integrated squared error when applied to sparsely populated tables, and unlike chi-square, does not diverge at low variance. Considerable attention is given to sparse contingency tables, which are inherent to single-gene libraries. CONCLUSION: Patterns of splicing correlations are revealed, which span a broad range of interaction order and change in development. The methods have a broad scope of applicability, beyond the single gene – including, for example, multiple gene interactions in the complete transcriptome

In Vitro and In Vivo Efficacy of Ether Lipid Edelfosine against Leishmania spp. and SbV-Resistant Parasites

Leishmaniasis represents a major international health problem, has a high morbidity and mortality rate, and is classified as an emerging and uncontrolled disease by the World Health Organization. The migration of population from endemic to nonendemic areas, and tourist activities in endemic regions are spreading the disease to new areas. Unfortunately, treatment of leishmaniasis is far from satisfactory, with only a few drugs available that show significant side-effects. Here, we show in vitro and in vivo evidence for the antileishmanial activity of the ether phospholipid edelfosine, being effective against a wide number of Leishmania spp. causing cutaneous, mucocutaneous and visceral leishmaniasis. Our experimental mouse and hamster models demonstrated not only a significant antileishmanial activity of edelfosine oral administration against different wild-type Leishmania spp., but also against parasites resistant to pentavalent antimonials, which constitute the first line of treatment worldwide. In addition, edelfosine exerted a higher antileishmanial activity and a lower proneness to generate drug resistance than miltefosine, the first drug against leishmaniasis that can be administered orally. These data, together with our previous findings, showing an anti-inflammatory action and a very low toxicity profile, suggest that edelfosine is a promising orally administered drug for leishmaniasis, thus warranting clinical evaluation

Cellulose nanofiber backboned Prussian blue nanoparticles as powerful adsorbents for the selective elimination of radioactive cesium

On 11 March 2011, the day of the unforgettable disaster of the 9 magnitude Tohoku earthquake and quickly followed by the devastating Tsunami, a damageable amount of radionuclides had dispersed from the Fukushima Daiichi’s damaged nuclear reactors. Decontamination of the dispersed radionuclides from seawater and soil, due to the huge amounts of coexisting ions with competitive functionalities, has been the topmost difficulty. Ferric hexacyanoferrate, also known as Prussian blue (PB), has been the most powerful material for selectively trapping the radioactive cesium ions; its high tendency to form stable colloids in water, however, has made PB to be impossible for the open-field radioactive cesium decontamination applications. A nano/nano combinatorial approach, as is described in this study, has provided an ultimate solution to this intrinsic colloid formation difficulty of PB. Cellulose nanofibers (CNF) were used to immobilize PB via the creation of CNF-backboned PB. The CNF-backboned PB (CNF/PB) was found to be highly tolerant to water and moreover, it gave a 139 mg/g capability and a million (106) order of magnitude distribution coefficient (Kd) for absorbing of the radioactive cesium ion. Field studies on soil and seawater decontaminations in Fukushima gave satisfactory results, demonstrating high capabilities of CNF/PB for practical applications.National Science Foundation (U.S.) (DMR-1507806