Long noncoding intronic RNAs are differentially expressed in primary and metastatic pancreatic cancer

<p>Abstract</p> <p>Background</p> <p>Pancreatic ductal adenocarcinoma (PDAC) is known by its aggressiveness and lack of effective therapeutic options. Thus, improvement in current knowledge of molecular changes associated with pancreatic cancer is urgently needed to explore novel venues of diagnostics and treatment of this dismal disease. While there is mounting evidence that long noncoding RNAs (lncRNAs) transcribed from intronic and intergenic regions of the human genome may play different roles in the regulation of gene expression in normal and cancer cells, their expression pattern and biological relevance in pancreatic cancer is currently unknown. In the present work we investigated the relative abundance of a collection of lncRNAs in patients' pancreatic tissue samples aiming at identifying gene expression profiles correlated to pancreatic cancer and metastasis.</p> <p>Methods</p> <p>Custom 3,355-element spotted cDNA microarray interrogating protein-coding genes and putative lncRNA were used to obtain expression profiles from 38 clinical samples of tumor and non-tumor pancreatic tissues. Bioinformatics analyses were performed to characterize structure and conservation of lncRNAs expressed in pancreatic tissues, as well as to identify expression signatures correlated to tissue histology. Strand-specific reverse transcription followed by PCR and qRT-PCR were employed to determine strandedness of lncRNAs and to validate microarray results, respectively.</p> <p>Results</p> <p>We show that subsets of intronic/intergenic lncRNAs are expressed across tumor and non-tumor pancreatic tissue samples. Enrichment of promoter-associated chromatin marks and over-representation of conserved DNA elements and stable secondary structure predictions suggest that these transcripts are generated from independent transcriptional units and that at least a fraction is under evolutionary selection, and thus potentially functional.</p> <p>Statistically significant expression signatures comprising protein-coding mRNAs and lncRNAs that correlate to PDAC or to pancreatic cancer metastasis were identified. Interestingly, <it>loci </it>harboring intronic lncRNAs differentially expressed in PDAC metastases were enriched in genes associated to the MAPK pathway. Orientation-specific RT-PCR documented that intronic transcripts are expressed in sense, antisense or both orientations relative to protein-coding mRNAs. Differential expression of a subset of intronic lncRNAs (<it>PPP3CB</it>, <it>MAP3K14 </it>and <it>DAPK1 loci</it>) in metastatic samples was confirmed by Real-Time PCR.</p> <p>Conclusion</p> <p>Our findings reveal sets of intronic lncRNAs expressed in pancreatic tissues whose abundance is correlated to PDAC or metastasis, thus pointing to the potential relevance of this class of transcripts in biological processes related to malignant transformation and metastasis in pancreatic cancer.</p

Análise de vizinhança de mapas conceituais a partir do uso de múltiplos conceitos obrigatórios

Os mapas conceituais (MCs) são úteis para representar o conhecimento e promover a aprendizagem significativa. O objetivo desse trabalho foi utilizar a análise de vizinhança (AViz) para avaliar a aprendizagem dos alunos. Oxigênio, hemácias, células e nutrientes foram conceitos obrigatórios (COs) que deveriam ser utilizados durante a elaboração de um MC para responder à seguinte pergunta: "como os nutrientes e o oxigênio chegam à célula?". Os resultados revelaram que proposições relacionando COs podem ser indicadores de uma adequada compreensão conceitual sobre o tema. Dos 36 MCs considerados, mais de 70% apresentaram 2 ou 3 proposições CO-CO. A AViz é uma maneira simples para identificar proposições com erros ou falta de clareza semântica, permitindo que os professores avaliem o nível de entendimento conceitual dos alunos durante o processo de aprendizagem

Análise de vizinhança de mapas conceituais a partir do uso de múltiplos conceitos obrigatórios

Concept maps (CMs) are useful to represent knowledge and promote meaningful learning. The aim of this study was to use Neighborhood Analysis (NeAn) to assess students' learning outcomes. Oxygen, erythrocyte, cells and nutrients were compulsory concepts (CCs) that must be used during the preparation of a CM to answer "how do nutrients and oxygen reach the cell?". The results showed that propositions involving CCs may be indicators of an adequate conceptual understanding about the topic. From all CMs considered in this study (n=36), more than 70% presented two or three CC-CC propositions. NeAn is a straightforward way to identify propositions with mistakes or lack of semantic, allowing teachers to assess students' level of conceptual understanding during the learning process.Os mapas conceituais (MCs) são úteis para representar o conhecimento e promover a aprendizagem significativa. O objetivo desse trabalho foi utilizar a análise de vizinhança (AViz) para avaliar a aprendizagem dos alunos. Oxigênio, hemácias, células e nutrientes foram conceitos obrigatórios (COs) que deveriam ser utilizados durante a elaboração de um MC para responder à seguinte pergunta: "como os nutrientes e o oxigênio chegam à célula?". Os resultados revelaram que proposições relacionando COs podem ser indicadores de uma adequada compreensão conceitual sobre o tema. Dos 36 MCs considerados, mais de 70% apresentaram 2 ou 3 proposições CO-CO. A AViz é uma maneira simples para identificar proposições com erros ou falta de clareza semântica, permitindo que os professores avaliem o nível de entendimento conceitual dos alunos durante o processo de aprendizagem

Cytogenetic and molecular analyses in troglobitic and epigean species of Pimelodella (Siluriformes: Heptapteridae) from Brazil

Samples from seven different locations of the genus Pimelodella were genetically examined, two caves (exclusively subterranean, upper Tocantins River and São Francisco River) and five epigean (from upper Paraná River basin). Cytogenetic analyses revealed the same diploid number (2n=46) for all species besides similarities in both number and location of nucleolar organizer regions and C bands. FISH with 5S rDNA probes and CMA3 staining indicated significant differences among the studied species. Application of PCR-RFLP in ATPase 6 and 8 mitochondrial genes allowed building a minimum evolution phenogram identifying the close evolutionary relationship among groups. Both chromosomal and molecular data were useful to infer the relationships among studied Pimelodella species.<br>Amostras de sete diferentes localidades do gênero Pimelodella foram geneticamente analisadas, duas cavernícolas (exclusivamente subterrâneas, alto rio Tocantins e rio São Francisco) e cinco epígeas (provenientes da bacia do alto Paraná). Análises citogenéticas revelaram o mesmo número diploide (2n=46) para todas as espécies, além de similaridades no número e localização das regiões organizadoras de nucléolo e bandas C. FISH com sondas de rDNA 5S e marcação com CMA3 indicaram diferenças significativas entre as espécies estudadas. A aplicação da técnica de PCR-RFLP nos genes mitocondriais ATPase 6 e 8 permitiu a construção de um fenograma de evolução mínima identificando uma estreita relação evolutiva entre as espécies estudadas

Cytogenetic and molecular analyses in troglobitic and epigean species of Pimelodella (Siluriformes: Heptapteridae) from Brazil

Samples from seven different locations of the genus Pimelodella were genetically examined, two caves (exclusively subterranean, upper Tocantins River and São Francisco River) and five epigean (from upper Paraná River basin). Cytogenetic analyses revealed the same diploid number (2n=46) for all species besides similarities in both number and location of nucleolar organizer regions and C bands. FISH with 5S rDNA probes and CMA3 staining indicated significant differences among the studied species. Application of PCR-RFLP in ATPase 6 and 8 mitochondrial genes allowed building a minimum evolution phenogram identifying the close evolutionary relationship among groups. Both chromosomal and molecular data were useful to infer the relationships among studied Pimelodella species

<i>Schistosoma mansoni</i> Egg, Adult Male and Female Comparative Gene Expression Analysis and Identification of Novel Genes by RNA-Seq

<div><p>Background</p><p>Schistosomiasis is one of the most prevalent parasitic diseases worldwide and is a public health problem. <i>Schistosoma mansoni</i> is the most widespread species responsible for schistosomiasis in the Americas, Middle East and Africa. Adult female worms (mated to males) release eggs in the hepatic portal vasculature and are the principal cause of morbidity. Comparative separate transcriptomes of female and male adult worms were previously assessed with using microarrays and Serial Analysis of Gene Expression (SAGE), thus limiting the possibility of finding novel genes. Moreover, the egg transcriptome was analyzed only once with limited bacterially cloned cDNA libraries.</p><p>Methodology/Principal findings</p><p>To compare the gene expression of <i>S</i>. <i>mansoni</i> eggs, females, and males, we performed RNA-Seq on these three parasite forms using 454/Roche technology and reconstructed the transcriptome using Trinity <i>de novo</i> assembly. The resulting contigs were mapped to the genome and were cross-referenced with predicted Smp genes and H3K4me3 ChIP-Seq public data. For the first time, we obtained separate, unbiased gene expression profiles for <i>S</i>. <i>mansoni</i> eggs and female and male adult worms, identifying enriched biological processes and specific enriched functions for each of the three parasite forms. Transcripts with no match to predicted genes were analyzed for their protein-coding potential and the presence of an encoded conserved protein domain. A set of 232 novel protein-coding genes with putative functions related to reproduction, metabolism, and cell biogenesis was detected, which contributes to the understanding of parasite biology.</p><p>Conclusions/Significance</p><p>Large-scale RNA-Seq analysis using <i>de novo</i> assembly associated with genome-wide information for histone marks in the vicinity of gene models constitutes a new approach to transcriptome analysis that has not yet been explored in schistosomes. Importantly, all data have been consolidated into a UCSC Genome Browser search- and download-tool (<a href="http://schistosoma.usp.br/" target="_blank">http://schistosoma.usp.br/</a>). This database provides new ways to explore the schistosome genome and transcriptome and will facilitate molecular research on this important parasite.</p></div

Contigs from the RNA-Seq data improve the Smp gene predictions, including the rearrangement of gene structure and the addition of new 5′-ends.

<p>The figure shows a Chromosome ZW 40-kb locus (Chr_ZW:2,310,000–2,350,000) harboring Smp_142960, Smp_177650 and Smp_188430 (blue). Contigs (red) from our RNA-Seq assembly align to Smp genes, showing their new architecture. The new contig c807_g1_i1 (red boxes on the right-hand side) merges Smp_177650 and Smp_188430 with the final exons from Smp_142960 and has a new 5′-end exon that overlaps the histone H3K4me3 peak (green). The new contig c3145_g1_i1 (red boxes on the left-hand side) matches the first two exons of Smp_142960 and adds a new 5′-end exon that corrects the TSS of the gene and overlaps the histone H3K4me3 peak (green). This locus can be seen at <a href="http://schistosoma.usp.br/" target="_blank">http://schistosoma.usp.br/</a> by entering the chromosome coordinates, the Smp number or the contig number in the “<i>enter position or search terms</i>” field at the top of the browser.</p