龍動と地震

REPORT 6 訪問・派遣研究員によるレポー

Comprehensive comparison of three commercial human whole-exome capture platforms

BackgroundExome sequencing, which allows the global analysis of protein coding sequences in the human genome, has become an effective and affordable approach to detecting causative genetic mutations in diseases. Currently, there are several commercial human exome capture platforms; however, the relative performances of these have not been characterized sufficiently to know which is best for a particular study.ResultsWe comprehensively compared three platforms: NimbleGen's Sequence Capture Array and SeqCap EZ, and Agilent's SureSelect. We assessed their performance in a variety of ways, including number of genes covered and capture efficacy. Differences that may impact on the choice of platform were that Agilent SureSelect covered approximately 1,100 more genes, while NimbleGen provided better flanking sequence capture. Although all three platforms achieved similar capture specificity of targeted regions, the NimbleGen platforms showed better uniformity of coverage and greater genotype sensitivity at 30- to 100-fold sequencing depth. All three platforms showed similar power in exome SNP calling, including medically relevant SNPs. Compared with genotyping and whole-genome sequencing data, the three platforms achieved a similar accuracy of genotype assignment and SNP detection. Importantly, all three platforms showed similar levels of reproducibility, GC bias and reference allele bias.ConclusionsWe demonstrate key differences between the three platforms, particularly advantages of solutions over array capture and the importance of a large gene target set

Predicting human microRNA precursors based on an optimized feature subset generated by GA–SVM

AbstractMicroRNAs (miRNAs) are non-coding RNAs that play important roles in post-transcriptional regulation. Identification of miRNAs is crucial to understanding their biological mechanism. Recently, machine-learning approaches have been employed to predict miRNA precursors (pre-miRNAs). However, features used are divergent and consequently induce different performance. Thus, feature selection is critical for pre-miRNA prediction. We generated an optimized feature subset including 13 features using a hybrid of genetic algorithm and support vector machine (GA–SVM). Based on SVM, the classification performance of the optimized feature subset is much higher than that of the two feature sets used in microPred and miPred by five-fold cross-validation. Finally, we constructed the classifier miR-SF to predict the most recently identified human pre-miRNAs in miRBase (version 16). Compared with microPred and miPred, miR-SF achieved much higher classification performance. Accuracies were 93.97%, 86.21% and 64.66% for miR-SF, microPred and miPred, respectively. Thus, miR-SF is effective for identifying pre-miRNAs

Association between CORIN promoter methylation and stroke: Results from two independent samples of Chinese adults

ObjectiveAs the physical activator of natriuretic peptides, corin has been associated with stroke, but the underlying mechanism is not very clear. Here, we examined whether the CORIN promoter’s methylation, an epigenetic DNA modification, was associated with the risk of stroke in two independent samples.MethodsA total of 1771 participants including 853 stroke cases and 918 healthy controls were included as a discovery sample and 2,498 community members with 10 years of follow-up were included as a replication sample. DNA methylation of the CORIN promoter was quantified by target bisulfite sequencing in both samples. We first examined the single CpG association, followed by a gene-based analysis of the joint association between multiple CpG methylation and stroke, adjusting for conventional risk factors.ResultsThe single CpG association analysis found that hypermethylation at all of the 9 CpG sites assayed was significantly associated with lower odds of prevalent stroke in the discovery sample (all p < 0.05), and three of them located at Chr4:47840038 (HR = 0.74, p = 0.015), Chr4:47839941 (HR = 0.80, p = 0.047), and Chr4:47839933 (HR = 0.82, p = 0.050) were also significantly associated with incident stroke in the replication sample. The gene-based association analysis found that DNA methylation of the 9 CpG sites at the CORIN promoter was jointly associated with stroke in both samples (all p < 0.05).ConclusionDNA methylation levels of the CORIN gene promoter were lower in stroke patients and predicted a higher risk of incident stroke in Chinese adults. The underlying causality warranted further investigation

Loss of COPZ1 induces NCOA4 mediated autophagy and ferroptosis in glioblastoma cell lines

Dysregulated iron metabolism is a hallmark of many cancers, including glioblastoma (GBM). However, its role in tumor progression remains unclear. Herein, we identified coatomer protein complex subunit zeta 1 (COPZ1) as a therapeutic target candidate which significantly dysregulated iron metabolism in GBM cells. Overexpression of COPZ1 was associated with increasing tumor grade and poor prognosis in glioma patients based on analysis of expression data from the publicly available database The Cancer Genome Atlas (P < 0.001). Protein levels of COPZ1 were significantly increased in GBM compared to non-neoplastic brain tissue samples in immunohistochemistry and western blot analysis. SiRNA knockdown of COPZ1 suppressed proliferation of U87MG, U251 and P3#GBM in vitro. Stable expression of a COPZ1 shRNA construct in U87MG inhibited tumor growth in vivo by ~60% relative to controls at day 21 after implantation (P < 0.001). Kaplan–Meier analysis of the survival data demonstrated that the overall survival of tumor bearing animals increased from 20.8 days (control) to 27.8 days (knockdown, P < 0.05). COPZ1 knockdown also led to the increase in nuclear receptor coactivator 4 (NCOA4), resulting in the degradation of ferritin, and a subsequent increase in the intracellular levels of ferrous iron and ultimately ferroptosis. These data demonstrate that COPZ1 is a critical mediator in iron metabolism. The COPZ1/NCOA4/FTH1 axis is therefore a novel therapeutic target for the treatment of human GBM.publishedVersio

A Systematic Analysis on DNA Methylation and the Expression of Both mRNA and microRNA in Bladder Cancer

Background: DNA methylation aberration and microRNA (miRNA) deregulation have been observed in many types of cancers. A systematic study of methylome and transcriptome in bladder urothelial carcinoma has never been reported. Methodology/Principal Findings: The DNA methylation was profiled by modified methylation-specific digital karyotyping (MMSDK) and the expression of mRNAs and miRNAs was analyzed by digital gene expression (DGE) sequencing in tumors and matched normal adjacent tissues obtained from 9 bladder urothelial carcinoma patients. We found that a set of significantly enriched pathways disrupted in bladder urothelial carcinoma primarily related to "neurogenesis" and "cell differentiation" by integrated analysis of -omics data. Furthermore, we identified an intriguing collection of cancer-related genes that were deregulated at the levels of DNA methylation and mRNA expression, and we validated several of these genes (HIC1, SLIT2, RASAL1, and KRT17) by Bisulfite Sequencing PCR and Reverse Transcription qPCR in a panel of 33 bladder cancer samples. Conclusions/Significance: We characterized the profiles between methylome and transcriptome in bladder urothelial carcinoma, identified a set of significantly enriched key pathways, and screened four aberrantly methylated and expressed genes. Conclusively, our findings shed light on a new avenue for basic bladder cancer research