bluuebunny/crossref_metadata_2025 huggingface dataset:
A new hybrid general iterative algorithm for common solutions of generalized mixed equilibrium problems and variational inclusions
Authors: Wattanawitoon Kriengsak , Jitpeera Thanyarat , Kumam Poom
Published: June 2012

In this article, we introduce a new general iterative method for finding a common element of the set of solutions generalized for mixed equilibrium problems, the set of solution for fixed point for nonexpansive mappings and the set of solutions for the variational inclusions for β
1, β
2-inverse-strongly monotone mappings in a real Hilbert space. We prove that the sequence converges strongly to a common element of the above sets under some suitable conditions. Our results improve and extend the corresponding results of Marino and Xu, Su et al., Tan and Chang and some authors.

Mathematics subject Classification: 46C05; 47H09; 47H10.
Transcriptome analysis of microRNAs in developing cerebral cortex of rat
Authors: Yao Mao-jin , Chen Gang , Zhao Ping-ping , Lu Ming-hua , Jian Jiang , Liu Mo-fang , Yuan Xiao-bing
Published: June 2012

Background

The morphogenesis of the cerebral cortex depends on the precise control of gene expression during development. Small non-coding RNAs, including microRNAs and other groups of small RNAs, play profound roles in various physiological and pathological processes via their regulation of gene expression. A systematic analysis of the expression profile of small non-coding RNAs in developing cortical tissues is important for clarifying the gene regulation networks mediating key developmental events during cortical morphogenesis.

Results

Global profiling of the small RNA transcriptome was carried out in rat cerebral cortex from E10 till P28 using next-generation sequencing technique. We found an extraordinary degree of developmental stage-specific expression of a large group of microRNAs. A group of novel microRNAs with functional hints were identified, and brain-enriched expression and Dicer-dependent production of high-abundant novel microRNAs were validated. Profound editing of known microRNAs at “seed” sequence and flanking sequence was observed, with much higher editing events detected at late postnatal stages than embryonic stages, suggesting the necessity of microRNA editing for the fine tuning of gene expression during the formation of complicated synaptic connections at postnatal stages.

Conclusion

Our analysis reveals extensive regulation of microRNAs during cortical development. The dataset described here will be a valuable resource for clarifying new regulatory mechanisms for cortical development and diseases and will greatly contribute to our understanding of the divergence, modification, and function of microRNAs.
Identification of metagenes and their Interactions through Large-scale Analysis of Arabidopsis Gene Expression Data
Authors: Wilson Tyler J , Lai Liming , Ban Yuguang , Ge Steven X
Published: June 2012

Background

Many plant genes have been identified through whole genome and deep transcriptome sequencing and other methods; yet our knowledge on the function of many of these genes remains limited. The integration and analysis of large gene-expression datasets gives researchers the ability to formalize hypotheses concerning the functionality and interaction between different groups of correlated genes.

Results

We applied the non-negative matrix factorization (NMF) algorithm to the AtGenExpress dataset which consists of 783 microarray samples (29 separate experimental series) conducted on the model plant Arabidopsis thaliana. We identified 15 metagenes, which are groups of genes with correlated expression. Functional roles of these metagenes are established by observing the enriched gene ontology (GO) categories using gene set enrichment analyses (GSEA). Activity levels of these metagenes in various experimental conditions are also analyzed to associate metagenes with stimuli/conditions. A metagene correlation network, constructed based on the results of NMF analysis, revealed many new interactions between the metagenes. Comparison of these metagenes with an earlier large-scale clustering analysis indicates many statistically significant overlaps.

Conclusions

This study identifies a network of correlated metagenes composed of Arabidopsis genes acting in a highly correlated fashion across a broad spectrum of experimental stimuli, which may shed some light on the function of many of the un-annotated genes.
Analyses of the sucrose synthase gene family in cotton: structure, phylogeny and expression patterns
Authors: Chen Aiqun , He Shae , Li Feifei , Li Zhao , Ding Mingquan , Liu Qingpo , Rong Junkang
Published: June 2012

Background

In plants, sucrose synthase (Sus) is widely considered as a key enzyme involved in sucrose metabolism. Several paralogous genes encoding different isozymes of Sus have been identified and characterized in multiple plant genomes, while limited information of Sus genes is available to date for cotton.

Results

Here, we report the molecular cloning, structural organization, phylogenetic evolution and expression profiles of seven Sus genes (GaSus1 to 7) identified from diploid fiber cotton (Gossypium arboreum). Comparisons between cDNA and genomic sequences revealed that the cotton GaSus genes were interrupted by multiple introns. Comparative screening of introns in homologous genes demonstrated that the number and position of Sus introns are highly conserved among Sus genes in cotton and other more distantly related plant species. Phylogenetic analysis showed that GaSus1, GaSus2, GaSus3, GaSus4 and GaSus5 could be clustered together into a dicot Sus group, while GaSus6 and GaSus7 were separated evenly into other two groups, with members from both dicot and monocot species. Expression profiles analyses of the seven Sus genes indicated that except GaSus2, of which the transcripts was undetectable in all tissues examined, and GaSus7, which was only expressed in stem and petal, the other five paralogues were differentially expressed in a wide ranges of tissues, and showed development-dependent expression profiles in cotton fiber cells.

Conclusions

This is a comprehensive study of the Sus gene family in cotton plant. The results presented in this work provide new insights into the evolutionary conservation and sub-functional divergence of the cotton Sus gene family in response to cotton fiber growth and development.
Molecular interactions between the olive and the fruit fly Bactrocera oleae
Authors: Corrado Giandomenico , Alagna Fiammetta , Rocco Mariapina , Renzone Giovanni , Varricchio Paola , Coppola Valentina , Coppola Mariangela , Garonna Antonio , Baldoni Luciana , Scaloni Andrea , Rao Rosa
Published: June 2012

Background

The fruit fly Bactrocera oleae is the primary biotic stressor of cultivated olives, causing direct and indirect damages that significantly reduce both the yield and the quality of olive oil. To study the olive-B. oleae interaction, we conducted transcriptomic and proteomic investigations of the molecular response of the drupe. The identifications of genes and proteins involved in the fruit response were performed using a Suppression Subtractive Hybridisation technique and a combined bi-dimensional electrophoresis/nanoLC-ESI-LIT-MS/MS approach, respectively.

Results

We identified 196 ESTs and 26 protein spots as differentially expressed in olives with larval feeding tunnels. A bioinformatic analysis of the identified non-redundant EST and protein collection indicated that different molecular processes were affected, such as stress response, phytohormone signalling, transcriptional control and primary metabolism, and that a considerable proportion of the ESTs could not be classified. The altered expression of 20 transcripts was also analysed by real-time PCR, and the most striking differences were further confirmed in the fruit of a different olive variety. We also cloned the full-length coding sequences of two genes, Oe-chitinase I and Oe-PR27, and showed that these are wound-inducible genes and activated by B. oleae punctures.

Conclusions

This study represents the first report that reveals the molecular players and signalling pathways involved in the interaction between the olive fruit and its most damaging biotic stressor. Drupe response is complex, involving genes and proteins involved in photosynthesis as well as in the production of ROS, the activation of different stress response pathways and the production of compounds involved in direct defence against phytophagous larvae. Among the latter, trypsin inhibitors should play a major role in drupe resistance reaction.
The database of chromosome imbalance regions and genes resided in lung cancer from Asian and Caucasian identified by array-comparative genomic hybridization
Authors: Lo Fang-Yi , Chang Jer-Wei , Chang I-Shou , Chen Yann-Jang , Hsu Han-Shui , Huang Shiu-Feng Kathy , Tsai Fang-Yu , Jiang Shih Sheng , Kanteti Rajani , Nandi Suvobroto , Salgia Ravi , Wang Yi-Ching
Published: June 2012

Background

Cancer-related genes show racial differences. Therefore, identification and characterization of DNA copy number alteration regions in different racial groups helps to dissect the mechanism of tumorigenesis.

Methods

Array-comparative genomic hybridization (array-CGH) was analyzed for DNA copy number profile in 40 Asian and 20 Caucasian lung cancer patients. Three methods including MetaCore analysis for disease and pathway correlations, concordance analysis between array-CGH database and the expression array database, and literature search for copy number variation genes were performed to select novel lung cancer candidate genes. Four candidate oncogenes were validated for DNA copy number and mRNA and protein expression by quantitative polymerase chain reaction (qPCR), chromogenic in situ hybridization (CISH), reverse transcriptase-qPCR (RT-qPCR), and immunohistochemistry (IHC) in more patients.

Results

We identified 20 chromosomal imbalance regions harboring 459 genes for Caucasian and 17 regions containing 476 genes for Asian lung cancer patients. Seven common chromosomal imbalance regions harboring 117 genes, included gain on 3p13-14, 6p22.1, 9q21.13, 13q14.1, and 17p13.3; and loss on 3p22.2-22.3 and 13q13.3 were found both in Asian and Caucasian patients. Gene validation for four genes including ARHGAP19 (10q24.1) functioning in Rho activity control, FRAT2 (10q24.1) involved in Wnt signaling, PAFAH1B1 (17p13.3) functioning in motility control, and ZNF322A (6p22.1) involved in MAPK signaling was performed using qPCR and RT-qPCR. Mean gene dosage and mRNA expression level of the four candidate genes in tumor tissues were significantly higher than the corresponding normal tissues (P<0.001~P=0.06). In addition, CISH analysis of patients indicated that copy number amplification indeed occurred for ARHGAP19 and ZNF322A genes in lung cancer patients. IHC analysis of paraffin blocks from Asian Caucasian patients demonstrated that the frequency of PAFAH1B1 protein overexpression was 68% in Asian and 70% in Caucasian.

Conclusions

Our study provides an invaluable database revealing common and differential imbalance regions at specific chromosomes among Asian and Caucasian lung cancer patients. Four validation methods confirmed our database, which would help in further studies on the mechanism of lung tumorigenesis.
Detection of cervical lymph node metastasis in head and neck cancer patients with clinically N0 neck—a meta-analysis comparing different imaging modalities
Authors: Liao Li-Jen , Lo Wu-Chia , Hsu Wan-Lun , Wang Chi-Te , Lai Mei-Shu
Published: June 2012

Background

How to properly manage clinically negative neck of head and neck cancer patients is a controversial topic. Research is now directed toward finding a method sensitive enough to bring the risk of occult metastases below 20%. The aim of this review was to compare the diagnostic accuracy of different imaging modalities, including CT, MRI, PET and US, in clinically N0 head and neck cancer patients.

Methods

For this systematic review and meta-analysis, PubMed and the Cochrane Database were searched for relevant original articles published up to May 2011. Inclusion criteria were as follows: articles were reported in English; CT, MRI, PET or US were performed to identify cervical metastases in clinically N0 head and neck squamous cell carcinoma; and data were sufficient for the calculation of true-positive or false-negative values. A bivariate random effect model was used to obtain pooled sensitivity and specificity. The positive and negative test probability of neck metastasis was generated based on Bayesian theory and collected data for different pre-test possibilities.

Results

Of the 168 identified relevant articles, 7 studies fulfilled all inclusion criteria for CT, 6 studies for MRI, 11 studies for PET and 8 studies for US. There was no difference in sensitivity and specificity among these imaging modalities, except CT was superior to US in specificity. The pooled estimates for sensitivity were 52% (95% confidence interval [CI], 39% ~ 65%), 65% (34 ~ 87%) 66% (47 ~ 80%), and 66% (45 ~ 77%), on a per-neck basis for CT, MRI, PET and US, respectively. The pooled estimates for specificity were 93% (87% ~ 97%), 81% (64 ~ 91%), 87% (77 ~ 93%), and 78% (71 ~ 83%) for CT, MRI, PET and US, respectively. With pre-examination nodal metastasis probabilities set at 10%, 20% and 30%, the post-exam probabilities of positive nodal metastasis rates were 47%, 66% and 77% for CT; 27%, 46% and 59% for MRI; 36%, 56% and 69% for PET; and 25%, 42% and 56% for US, respectively. Negative nodal metastasis probabilities were 95%, 89% and 82% for CT; 95%, 90% and 84% for MRI; 96%, 91% and 86% for PET; and 95%, 90% and 84% for US, respectively.

Conclusions

Modern imaging modalities offer similar diagnostic accuracy to define and diagnose clinically N0 neck. Minimizing morbidity and avoiding elective neck dissection is acceptable in some select cases.
Li-Fraumeni-like syndrome associated with a large BRCA1 intragenic deletion
Authors: Silva Amanda Gonçalves , Ewald Ingrid Petroni , Sapienza Marina , Pinheiro Manuela , Peixoto Ana , de Nóbrega Amanda França , Carraro Dirce M , Teixeira Manuel R , Ashton-Prolla Patricia , Achatz Maria Isabel W , Rosenberg Carla , Krepischi Ana C V
Published: June 2012

Background

Li-Fraumeni (LFS) and Li-Fraumeni-like (LFL) syndromes are associated to germline TP53 mutations, and are characterized by the development of central nervous system tumors, sarcomas, adrenocortical carcinomas, and other early-onset tumors. Due to the high frequency of breast cancer in LFS/LFL families, these syndromes clinically overlap with hereditary breast cancer (HBC). Germline point mutations in BRCA1, BRCA2, and TP53 genes are associated with high risk of breast cancer. Large rearrangements involving these genes are also implicated in the HBC phenotype.

Methods

We have screened DNA copy number changes by MLPA on BRCA1, BRCA2, and TP53 genes in 23 breast cancer patients with a clinical diagnosis consistent with LFS/LFL; most of these families also met the clinical criteria for other HBC syndromes.

Results

We found no DNA copy number alterations in the BRCA2 and TP53 genes, but we detected in one patient a 36.4 Kb BRCA1 microdeletion, confirmed and further mapped by array-CGH, encompassing exons 9–19. Breakpoints sequencing analysis suggests that this rearrangement was mediated by flanking Alu sequences.

Conclusion

This is the first description of a germline intragenic BRCA1 deletion in a breast cancer patient with a family history consistent with both LFL and HBC syndromes. Our results show that large rearrangements in these known cancer predisposition genes occur, but are not a frequent cause of cancer susceptibility.
Prognostic factors of successful tympanoplasty in pediatric patients: a cohort study
Authors: Boronat-Echeverría Nuria Esperanza , Reyes-García Esmeralda , Sevilla-Delgado Yolanda , Aguirre-Mariscal Héctor , Mejía-Aranguré Juan Manuel
Published: June 2012

Background

Tympanoplasty in children is a current and controversial theme. The success of tympanoplasty traditionally has been measured only by the post-operative integrity of the graft. Yet, there are other variables that may be used to determine success. The objectives of the present work were to analyze which factors are predictive of successful tympanoplasty in pediatric patients and to construct and validate a prognostic index that could be used as a tool to predict the success of tympanoplasty in children.

Methods

Setting. Department of Pediatric Otorhinolaryngology, tertiary-care hospital, Mexico City. Patients. Forty-eight patients, who were older that five years of age, had persistent perforation of the tympanic membrane, and had undergone tympanoplasty (January 2005–June 2008), were followed for a year. Main Outcome Measures. The factors tested for their value as predictors were the following: age at time of surgery, state of contralateral ear, previous adenoidectomy, cause of perforation, size of perforation, infection at the time of surgery, state of mucosa, age at first occurrence of perforation, presence of craniofacial dysmorphia, and surgical technique. These factors were compared with the criterion, success, which was defined as attaining three positive outcomes: 1) integrity of the implant or membrane; 2) minimum of 10-dB gain in the auditory threshold or, in the case of normal hearing, conservation of same; and 3) air-filled space in the middle ear. The best model was obtained through logistic regression analysis; the model was validated.

Results

The most balanced prediction model was that in which the three success criteria were included, with age, surgical technique, and infection at surgery being excluded as variables. The additional 12 pediatric cases used in the validation had a probability of success >0.425 (best cut-off level); two patients (17%) had poor evolution.

Conclusions

This is the first study that validated a predictive index of the result of tympanoplasty in children. This index predicted 81% of the successful outcomes.
Prevalence of molecular markers of Plasmodium falciparum drug resistance in Dakar, Senegal
Authors: Wurtz Nathalie , Fall Bécaye , Pascual Aurélie , Diawara Silmane , Sow Kowry , Baret Eric , Diatta Bakary , Fall Khadidiatou B , Mbaye Pape S , Fall Fatou , Diémé Yaya , Rogier Christophe , Bercion Raymond , Briolant Sébastien , Wade Boubacar , Pradines Bruno
Published: June 2012

Background

As a result of the widespread resistance to chloroquine and sulphadoxine-pyrimethamine, artemisinin-based combination therapy (ACT) (including artemether-lumefantrine and artesunate-amodiaquine) has been recommended as a first-line anti-malarial regimen in Senegal since 2006. Intermittent preventive treatments with anti-malarial drugs based on sulphadoxine-pyrimethamine are also given to children or pregnant women once per month during the transmission season. Since 2006, there have been very few reports on the susceptibility of Plasmodium falciparum to anti-malarial drugs. To estimate the prevalence of resistance to several anti-malarial drugs since the introduction of the widespread use of ACT, the presence of molecular markers associated with resistance to chloroquine and sulphadoxine-pyrimethamine was assessed in local isolates at the military hospital of Dakar.

Methods

The prevalence of genetic polymorphisms in genes associated with anti-malarial drug resistance, i.e., Pfcrt, Pfdhfr, Pfdhps and Pfmdr1, and the copy number of Pfmdr1 were evaluated for a panel of 174 isolates collected from patients recruited at the military hospital of Dakar from 14 October 2009 to 19 January 2010.

Results

The Pfcrt 76T mutation was identified in 37.2% of the samples. The Pfmdr1 86Y and 184F mutations were found in 16.6% and 67.6% of the tested samples, respectively. Twenty-eight of the 29 isolates with the 86Y mutation were also mutated at codon 184. Only one isolate (0.6%) had two copies of Pfmdr1. The Pfdhfr 108N/T, 51I and 59R mutations were identified in 82.4%, 83.5% and 74.1% of the samples, respectively. The double mutant (108N and 51I) was detected in 83.5% of the isolates, and the triple mutant (108N, 51I and 59R) was detected in 75.3%. The Pfdhps 437G, 436F/A and 613S mutations were found in 40.2%, 35.1% and 1.8% of the samples, respectively. There was no double mutant (437G and 540E) or no quintuple mutant (Pfdhfr 108N, 51I and 59R and Pfdhps 437G and 540E). The prevalence of the quadruple mutant (Pfdhfr 108N, 51I and 59R and Pfdhps 437G) was 36.5%.

Conclusions

Since 2004, the prevalence of chloroquine resistance had decreased. The prevalence of isolates with high-level pyrimethamine resistance is 83.5%. The prevalence of isolates resistant to sulphadoxine is 40.2%. However, no quintuple mutant (Pfdhfr 108N, 51I and 59R and Pfdhps 437G and 540E), which is associated with a high level of sulphadoxine-pyrimethamine resistance, has been identified to date. The resistance to amodiaquine remains moderate.
The RASSF gene family members RASSF5, RASSF6 and RASSF7 show frequent DNA methylation in neuroblastoma
Authors: Djos Anna , Martinsson Tommy , Kogner Per , Carén Helena
Published: June 2012

Background

Hypermethylation of promotor CpG islands is a common mechanism that inactivates tumor suppressor genes in cancer. Genes belonging to theRASSFgene family have frequently been reported as epigenetically silenced by promotor methylation in human cancers. Two members of this gene family,RASSF1AandRASSF5Ahave been reported as methylated in neuroblastoma. Data from our previously performed genome-wide DNA methylation array analysis indicated that other members of theRASSFgene family are targeted by DNA methylation in neuroblastoma.

Results

In the current study, we found that several of theRASSFfamily genes (RASSF2,RASSF4,RASSF5,RASSF6,RASSF7, andRASSF10) to various degrees were methylated in neuroblastoma cell lines and primary tumors. In addition, several of theRASSFfamily genes showed low or absent mRNA expression in neuroblastoma cell lines.RASSF5andRASSF6were to various degrees methylated in a large portion of neuroblastoma tumors andRASSF7was heavily methylated in most tumors. Further, CpG methylation sites in the CpG islands of someRASSFfamily members could be used to significantly discriminate between biological subgroups of neuroblastoma tumors. For example,RASSF5methylation highly correlated toMYCNamplification and INRG stage M. Furthermore, high methylation ofRASSF6was correlated to unfavorable outcome, 1p deletion andMYCNamplification in our tumor material.

In conclusion

This study shows that several genes belonging to theRASSFgene family are methylated in neuroblastoma. The genesRASSF5,RASSF6andRASSF7stand out as the most promising candidate genes for further investigations in neuroblastoma.
Role of glycogen synthase kinase-3 beta in the inflammatory response caused by bacterial pathogens
Authors: Cortés-Vieyra Ricarda , Bravo-Patiño Alejandro , Valdez-Alarcón Juan J , Juárez Marcos Cajero , Finlay B Brett , Baizabal-Aguirre Víctor M
Published: June 2012

Glycogen synthase kinase 3β (GSK3β) plays a fundamental role during the inflammatory response induced by bacteria. Depending on the pathogen and its virulence factors, the type of cell and probably the context in which the interaction between host cells and bacteria takes place, GSK3β may promote or inhibit inflammation. The goal of this review is to discuss recent findings on the role of the inhibition or activation of GSK3β and its modulation of the inflammatory signaling in monocytes/macrophages and epithelial cells at the transcriptional level, mainly through the regulation of nuclear factor-kappaB (NF-κB) activity. Also included is a brief overview on the importance of GSK3 in non-inflammatory processes during bacterial infection.
Comparative proteomic analysis of transition of saccharomyces cerevisiae from glucose-deficient medium to glucose-rich medium
Authors: Giardina Bennett J , Stanley Bruce A , Chiang Hui-Ling
Published: June 2012

Background

When glucose is added to Saccharomyces cerevisiae grown in non-fermentable carbon sources, genes encoding ribosomal, cell-cycle, and glycolytic proteins are induced. By contrast, genes involved in mitochondrial functions, gluconeogenesis, and the utilization of other carbon sources are repressed. Glucose also causes the activation of the plasma membrane ATPase and the inactivation of gluconeogenic enzymes and mitochondrial enzymes. The goals of this study were to use the iTRAQ-labeling mass spectrometry technique to identify proteins whose relative levels change in response to glucose re-feeding and to correlate changes in protein abundance with changes in transcription and enzymatic activities. We used an experimental condition that causes the degradation of gluconeogenic enzymes when glucose starved cells are replenished with glucose. Identification of these enzymes as being down-regulated by glucose served as an internal control. Furthermore, we sought to identify new proteins that were either up-regulated or down-regulated by glucose.

Results

We have identified new and known proteins that change their relative levels in cells that were transferred from medium containing low glucose to medium containing high glucose. Up-regulated proteins included ribosomal subunits, proteins involved in protein translation, and the plasma membrane ATPase. Down-regulated proteins included small heat shock proteins, mitochondrial proteins, glycolytic enzymes, and gluconeogenic enzymes. Ach1p is involved in acetate metabolism and is also down-regulated by glucose.

Conclusions

We have identified known proteins that have previously been reported to be regulated by glucose as well as new glucose-regulated proteins. Up-regulation of ribosomal proteins and proteins involved in translation may lead to an increase in protein synthesis and in nutrient uptake. Down-regulation of glycolytic enzymes, gluconeogenic enzymes, and mitochondrial proteins may result in changes in glycolysis, gluconeogenesis, and mitochondrial functions. These changes may be beneficial for glucose-starved cells to adapt to the addition of glucose.
EQ-5D™-derived utility values for different levels of migraine severity from a UK sample of migraineurs
Authors: Stafford Megan R , Hareendran Asha , Ng-Mak Daisy S , Insinga Ralph P , Xu Ruifeng , Stull Donald E
Published: June 2012

Background

To estimate utility values for different levels of migraine pain severity from a United Kingdom (UK) sample of migraineurs.

Methods

One hundred and six migraineurs completed the EQ-5D to evaluate their health status for mild, moderate and severe levels of migraine pain severity for a recent migraine attack, and for current health defined as health status within seven days post-migraine attack. Statistical tests were used to evaluate differences in mean utility scores by migraine severity.

Results

Utility scores for each health state were significantly different from 1.0 (no problems on any EQ-5D dimension) (p < 0.0001) and one another (p < 0.0001). The lowest mean utility, − 0.20 (95% confidence interval [CI]: -0.27 – -0.13), was for severe migraine pain. The smallest difference in mean utility was between mild and moderate migraine pain (0.13) and the largest difference in mean utility was between current health (without migraine) and severe migraine pain (1.07).

Conclusions

Results indicate that all levels of migraine pain are associated with significantly reduced utility values. As severity worsened, utility decreased and severe migraine pain was considered a health state worse than death. Results can be used in cost-utility models examining the relative economic value of therapeutic strategies for migraine in the UK.
Role of SAMHD1 nuclear localization in restriction of HIV-1 and SIVmac
Authors: Brandariz-Nuñez Alberto , Valle-Casuso Jose Carlos , White Tommy E , Laguette Nadine , Benkirane Monsef , Brojatsch Jurgen , Diaz-Griffero Felipe
Published: June 2012

Background

SAMHD1 is a nuclear protein that blocks lentiviral infection before reverse transcription in macrophages and dendritic cells. The viral accessory protein Vpx overcomes the SAMHD1-mediated lentiviral block by inducing its proteasomal degradation.

Results

Here, we identified the nuclear localization signal (NLS) of SAMHD1, and studied its contribution to restriction of HIV-1 and SIVmac. By studying the cellular distribution of different SAMHD1 variants, we mapped the nuclear localization of SAMHD1 to residues11KRPR14. Mutagenesis of these residues changed the cellular distribution of SAMHD1 from the nucleus to the cytoplasm. SAMHD1 mutants that lost nuclear localization restricted HIV-1 and SIV as potently as the wild type protein. Interestingly, SAMHD1 mutants that localized to the cytoplasm were not degraded by nuclear Vpx alleles. Therefore, nuclear Vpx alleles require nuclear localization of SAMHD1 in order to induce its degradation. In agreement, SIVmac viruses encoding Vpx did not overcome the restriction imposed by the cytoplasmic variants of SAMHD1.

Conclusions

We mapped the NLS of SAMHD1 to residues11KRPR14and studied the contribution of SAMHD1 nuclear localization to restriction of HIV-1 and SIV. These experiments demonstrate that cytoplasmic variants of SAMHD1 potently block lentiviral infection and are resistant to Vpx-mediated degradation. The nuclear Vpx alleles studied here are only capable of degrading a nuclearly localized SAMHD1 suggesting that Vpx-mediated degradation of SAMHD1 is initiated in the nucleus.
Loop-mediated isothermal amplification for the detection of goose circovirus
Authors: Woźniakowski Grzegorz , Kozdruń Wojciech , Samorek-Salamonowicz Elżbieta
Published: June 2012

Background

Goose circovirus (GCV) presents an immunosuppressive problem in production of geese. The infection’s clinical symptoms include growth retardation or feathering disorders but the infection process may remain non-symptomatic what makes the infected birds more susceptible for secondary viral, bacterial and fungal infections. Diagnosis of GCV infection is made by histopathological examination, dot blot hybridization, polymerase chain reaction (PCR) and real-time PCR. However these techniques require application of thermocyclers and qualified staff which may be cost-consuming for some diagnostic units. The aim of this study was to develop loop-mediated isothermal amplification assay (LAMP) as a simple method of GCV detection.

Results

The presented study has shown LAMP as a rapid tool of detecting DNA of goose circovirus (GCV) as soon in 30 min time. The method used three sets of primers: two outer primers (F3 and B3), two inner primers (FIP and BIP) and two loop primers (FL and BL) to accelerate the reaction. The optimum reaction temperature and the time were 61°C for 30 min, respectively. The results were analysed using SYBR Green dye and GelRedTM solutions. Thirty-eight isolates of GCV collected from geese flocks in Poland were examined. For comparison, real-time polymerase chain reaction with F3 and B3 primers and SYBR Green dye was conducted. The obtained results have shown GCV-LAMP as a sensitive, rapid and specific assay and alternative for PCR-based methods.

Conclusions

The developed technique due to its simplicity may be applied by any veterinary laboratory or even mobile diagnostics units for the routine detection of GCV.
Rapid Detection of high-level oncogene amplifications in ultrasonic surgical aspirations of brain tumors
Authors: Truong Long N , Patil Shashikant , Martin Sherry S , LeBlanc Jay F , Nanda Anil , Nordberg Mary L , Beckner Marie E
Published: June 2012

Background

Genomic tumor information, such as identification of amplified oncogenes, can be used to plan treatment. The two sources of a brain tumor that are commonly available include formalin-fixed, paraffin-embedded (FFPE) sections from the small diagnostic biopsy and the ultrasonic surgical aspiration that contains the bulk of the tumor. In research centers, frozen tissue of a brain tumor may also be available. This study compared ultrasonic surgical aspiration and FFPE specimens from the same brain tumors for retrieval of DNA and molecular assessment of amplified oncogenes.

Methods

Surgical aspirations were centrifuged to separate erythrocytes from the tumor cells that predominantly formed large, overlying buffy coats. These were sampled to harvest nuclear pellets for DNA purification. Four glioblastomas, 2 lung carcinoma metastases, and an ependymoma were tested. An inexpensive PCR technique, multiplex ligation-dependent probe amplification (MLPA), quantified 79 oncogenes using 3 kits. Copy number (CN) results were normalized to DNA from non-neoplastic brain (NB) in calculated ratios, [tumor DNA]/[NB DNA]. Bland-Altman and Spearman rank correlative comparisons were determined. Regression analysis identified outliers.

Results

Purification of DNA from ultrasonic surgical aspirations was rapid (<3 days) versus FFPE (weeks) and yielded greater amounts in 6 of 7 tumors. Gene amplifications up to 15-fold corresponded closely between ultrasonic aspiration and FFPE assays in Bland-Altman analysis. Correlation coefficients ranged from 0.71 to 0.99 using 3 kit assays per tumor. Although normalized CN ratios greater than 2.0 were more numerous in FFPE specimens, some were found only in the ultrasonic surgical aspirations, consistent with tumor heterogeneity. Additionally, CN ratios revealed 9 high-level (≥ 6.0) gene amplifications in FFPE of which 8 were also detected in the ultrasonic aspirations at increased levels. The ultrasonic aspiration levels of these amplified genes were also greater than 6.0 CN ratio, except in one case (3.53 CN ratio). Ten of 17 mid-level (≥3.0 & <6.0 CN ratio) amplifications detected in FFPE were also detected as being increased (≥ 2.0 CN ratio) in the aspirations.

Conclusions

Buffy coats of centrifuged ultrasonic aspirations contained abundant tumor cells whose DNA permitted rapid, multiplex detection of high-level oncogene amplifications that were confirmed in FFPE.

Virtual slides

http://www.diagnosticpathology.diagnomx.eu/vs/1883718801686466
Distinct perinatal features of the hyperpolarization-activated non-selective cation current Ih in the rat cortical plate
Authors: Battefeld Arne , Rocha Nino , Stadler Konstantin , Bräuer Anja U , Strauss Ulf
Published: June 2012

Background

During neocortical development, multiple voltage- and ligand-gated ion channels are differentially expressed in neurons thereby shaping their intrinsic electrical properties. One of these voltage-gated ion channels, the hyperpolarization-activated cyclic nucleotide-gated (HCN) channel and its current I
h, is an important regulator of neuronal excitability. Thus far, studies on an early I
h appearance in rodent neocortex are missing or conflicting. Therefore, we focused our study on perinatal neocortical I
h and its properties.

Results

In the perinatal rat neocortex we observed a rapid increase in the number of neurons exhibiting I
h. Perinatal I
h had unique properties: first, a pronounced cAMP sensitivity resulting in a marked shift of the voltage sufficient for half-maximum activation of the current towards depolarized voltages and second, an up to 10 times slower deactivation at physiological membrane potentials when compared to the one at postnatal day 30. The combination of these features was sufficient to suppress membrane resonance in our in silico and in vitro experiments. Although all four HCN subunits were present on the mRNA level we only detected HCN4, HCN3 and HCN1 on the protein level at P0. HCN1 protein at P0, however, appeared incompletely processed. At P30 glycosilated HCN1 and HCN2 dominated. By in silico simulations and heterologous co-expression experiments of a ‘slow’ and a ‘fast’ I
h conducting HCN channel subunit in HEK293 cells, we mimicked most characteristics of the native current, pointing to a functional combination of subunit homo- or heteromeres.

Conclusion

Taken together, these data indicate a HCN subunit shift initiated in the first 24 hours after birth and implicate a prominent perinatal role of the phylogenetically older HCN3 and/or HCN4 subunits in the developing neocortex.
Combinatorial network of transcriptional regulation and microRNA regulation in human cancer
Authors: Yu Hui , Tu Kang , Wang Yi-Jie , Mao Jun-Zhe , Xie Lu , Li Yuan-Yuan , Li Yi-Xue
Published: June 2012

Background

Both transcriptional control and microRNA (miRNA) control are critical regulatory mechanisms for cells to direct their destinies. At present, the combinatorial regulatory network composed of transcriptional regulations and post-transcriptional regulations is often constructed through a forward engineering strategy that is based solely on searching of transcriptional factor binding sites or miRNA seed regions in the putative target sequences. If the reverse engineering strategy is integrated with the forward engineering strategy, a more accurate and more specific combinatorial regulatory network will be obtained.

Results

In this work, utilizing both sequence-matching information and parallel expression datasets of miRNAs and mRNAs, we integrated forward engineering with reverse engineering strategies and as a result built a hypothetical combinatorial gene regulatory network in human cancer. The credibility of the regulatory relationships in the network was validated by random permutation procedures and supported by authoritative experimental evidence-based databases. The global and local architecture properties of the combinatorial regulatory network were explored, and the most important tumor-regulating miRNAs and TFs were highlighted from a topological point of view.

Conclusions

By integrating the forward engineering and reverse engineering strategies, we manage to sketch a genome-scale combinatorial gene regulatory network in human cancer, which includes transcriptional regulations and miRNA regulations, allowing systematic study of cancer gene regulation. Our work establishes a pipeline that can be extended to reveal conditional combinatorial regulatory landscapes correlating to specific cellular contexts.
New finite-size correction for local alignment score distributions
Authors: Park Yonil , Sheetlin Sergey , Ma Ning , Madden Thomas L , Spouge John L
Published: June 2012

Background

Local alignment programs often calculate the probability that a match occurred by chance. The calculation of this probability may require a “finite-size” correction to the lengths of the sequences, as an alignment that starts near the end of either sequence may run out of sequence before achieving a significant score.

Findings

We present an improved finite-size correction that considers the distribution of sequence lengths rather than simply the corresponding means. This approach improves sensitivity and avoids substituting an ad hoc length for short sequences that can underestimate the significance of a match. We use a test set derived from ASTRAL to show improved ROC scores, especially for shorter sequences.

Conclusions

The new finite-size correction improves the calculation of probabilities for a local alignment. It is now used in the BLAST+ package and at the NCBI BLAST web site (http://blast.ncbi.nlm.nih.gov).
C5a and its receptors in human anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis
Authors: Yuan Jun , Gou Shen-Ju , Huang Jing , Hao Jian , Chen Min , Zhao Ming-Hui
Published: June 2012

Introduction

The complement system is crucial for the development of antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). In particular, C5a plays a central role. In this study, plasma and urinary levels of C5a as well as renal C5a receptors (CD88 and C5L2) expression were investigated in patients with AAV.

Methods

Twenty-four patients with AAV in the active phase, 19 patients with AAV in the remission phase, and 20 patients with lupus nephritis (LN) were included. Plasma and urinary levels of C5a were measured with enzyme-linked immunosorbent assay (ELISA). The staining of CD88 and C5L2 in renal specimens was detected with immunohistochemistry.

Results

The level of plasma C5a was significantly higher in patients with AAV in the active phase than that in patients in remission, that in patients with LN, and that in normal controls. The urinary C5a level was significantly higher in patients with AAV in the active phase than that in patients in remission and that in normal controls, but not significantly different between patients with active AAV and patients with LN. The mean optical density of CD88 staining in the tubulointerstitium was significantly lower in AAV patients than that in normal controls (0.0052 ± 0.0011 versus 0.029 ± 0.0042; P = 0.005). The mean optical density of C5L2 in glomeruli was significantly higher in AAV patients than that in normal controls (0.013 ± 0.0027 versus 0.0032 ± 0.0006; P < 0.001). The mean optical density of CD88 staining closely correlated with the initial eGFR (r = 0.835; P < 0.001) in AAV patients. Double-labeling immunofluorescence assay suggested that CD88 did not express on neutrophils, monocytes, or macrophages, but C5L2 expressed on neutrophils (or monocytes) and macrophages.

Conclusion

The elevated plasma and urinary C5a levels indicated complement activation in human AAV. The level of renal CD88 expression could reflect the disease severity of ANCA-associated glomerulonephritis. CD88 expression was downregulated, and C5L2 was upregulated in ANCA-associated glomerulonephritis.
The genomic landscape shaped by selection on transposable elements across 18 mouse strains
Authors: Nellåker Christoffer , Keane Thomas M , Yalcin Binnaz , Wong Kim , Agam Avigail , Belgard T Grant , Flint Jonathan , Adams David J , Frankel Wayne N , Ponting Chris P
Published: June 2012

Background

Transposable element (TE)-derived sequence dominates the landscape of mammalian genomes and can modulate gene function by dysregulating transcription and translation. Our current knowledge of TEs in laboratory mouse strains is limited primarily to those present in the C57BL/6J reference genome, with most mouse TEs being drawn from three distinct classes, namely short interspersed nuclear elements (SINEs), long interspersed nuclear elements (LINEs) and the endogenous retrovirus (ERV) superfamily. Despite their high prevalence, the different genomic and gene properties controlling whether TEs are preferentially purged from, or are retained by, genetic drift or positive selection in mammalian genomes remain poorly defined.

Results

Using whole genome sequencing data from 13 classical laboratory and 4 wild-derived mouse inbred strains, we developed a comprehensive catalogue of 103,798 polymorphic TE variants. We employ this extensive data set to characterize TE variants across the Mus lineage, and to infer neutral and selective processes that have acted over 2 million years. Our results indicate that the majority of TE variants are introduced though the male germline and that only a minority of TE variants exert detectable changes in gene expression. However, among genes with differential expression across the strains there are twice as many TE variants identified as being putative causal variants as expected.

Conclusions

Most TE variants that cause gene expression changes appear to be purged rapidly by purifying selection. Our findings demonstrate that past TE insertions have often been highly deleterious, and help to prioritize TE variants according to their likely contribution to gene expression or phenotype variation.
Apple miRNAs and tasiRNAs with novel regulatory networks
Authors: Xia Rui , Zhu Hong , An Yong-qiang , Beers Eric P , Liu Zongrang
Published: June 2012

Background

MicroRNAs (miRNAs) and their regulatory functions have been extensively characterized in model species but whether apple has evolved similar or unique regulatory features remains unknown.

Results

We performed deep small RNA-seq and identified 23 conserved, 10 less-conserved and 42 apple-specific miRNAs or families with distinct expression patterns. The identified miRNAs target 118 genes representing a wide range of enzymatic and regulatory activities. Apple also conserves two TAS gene families with similar but unique trans-acting small interfering RNA (tasiRNA) biogenesis profiles and target specificities. Importantly, we found that miR159, miR828 and miR858 can collectively target up to 81 MYB genes potentially involved in diverse aspects of plant growth and development. These miRNA target sites are differentially conserved among MYBs, which is largely influenced by the location and conservation of the encoded amino acid residues in MYB factors. Finally, we found that 10 of the 19 miR828-targeted MYBs undergo small interfering RNA (siRNA) biogenesis at the 3' cleaved, highly divergent transcript regions, generating over 100 sequence-distinct siRNAs that potentially target over 70 diverse genes as confirmed by degradome analysis.

Conclusions

Our work identified and characterized apple miRNAs, their expression patterns, targets and regulatory functions. We also discovered that three miRNAs and the ensuing siRNAs exploit both conserved and divergent sequence features of MYB genes to initiate distinct regulatory networks targeting a multitude of genes inside and outside the MYB family.
Monocarboxylate Transporter 10 Functions as a Thyroid Hormone Transporter in Chondrocytes
Authors: Abe Sanae , Namba Noriyuki , Abe Makoto , Fujiwara Makoto , Aikawa Tomonao , Kogo Mikihiko , Ozono Keiichi
Published: June 2012

Thyroid hormone is essential for normal proliferation and differentiation of chondrocytes. Thus, untreated congenital hypothyroidism is marked by severe short stature. The monocarboxylate transporter 8 (MCT8) is a highly specific transporter for thyroid hormone. The hallmarks of Allan-Herndon-Dudley syndrome, caused by MCT8 mutations, are severe psychomotor retardation and elevated T3 levels. However, growth is mostly normal. We therefore hypothesized that growth plate chondrocytes use transporters other than MCT8 for thyroid hormone uptake. Extensive analysis of thyroid hormone transporter mRNA expression in mouse chondrogenic ATDC5 cells revealed that monocarboxylate transporter 10 (Mct10) was most abundantly expressed among the transporters known to be highly specific for thyroid hormone, namely Mct8, Mct10, and organic anion transporter 1c1. Expression levels of Mct10 mRNA diminished with chondrocyte differentiation in these cells. Accordingly, Mct10 mRNA was expressed most abundantly in the growth plate resting zone chondrocytes in vivo. Small interfering RNA-mediated knockdown of Mct10 mRNA in ATDC5 cells decreased [125I]T3 uptake up to 44% compared with negative control (P < 0.05). Moreover, silencing Mct10 mRNA expression abolished the known effects of T3, i.e. suppression of proliferation and enhancement of differentiation, in ATDC5 cells. These results suggest that Mct10 functions as a thyroid hormone transporter in chondrocytes and can explain at least in part why Allan-Herndon-Dudley syndrome patients do not exhibit significant growth impairment.