Project description:Alternative splicing of pre-mRNA generates protein diversity and has been linked to cancer progression and drug response. Exon microarray technology enables genome-wide quantication of expression levels for the majority of exons and facilitates the discovery of alternative splicing events. Analysis of exon array data is more challenging than gene expression data and there is a need for reliable quantication of exons and alternative spliced variants. We introduce a novel, computationally efficient methodology, MEAP, for exon array data preprocessing, analysis and visualization. We compared MEAP with other preprocessing methods, and validation of the results show that MEAP produces reliable quantication of exons and alternative spliced variants. Analysis of data from head and neck squamous cell carcinoma (HNSCC) cell lines revealed several variants associated with 11q13 amplication, which is a predictive marker of metastasis and decreased survival in HNSCC patients. Together these results demonstrate the utility of MEAP in suggesting novel experimentally testable predictions. Thus, in addition to novel methodology to process large-scale exon array data sets, our results provide several HNSCC candidate genes for further studies. We analyzed 15 samples using the Affymetrix Human Exon 1.0 ST platform, of which 7 samples have 11q13 amplification. Array data was preprocessed by using Multiple Exon Array Processing (MEAP).
Project description:Alternative splicing of pre-mRNA generates protein diversity and has been linked to cancer progression and drug response. Exon microarray technology enables genome-wide quantication of expression levels for the majority of exons and facilitates the discovery of alternative splicing events. Analysis of exon array data is more challenging than gene expression data and there is a need for reliable quantication of exons and alternative spliced variants. We introduce a novel, computationally efficient methodology, MEAP, for exon array data preprocessing, analysis and visualization. We compared MEAP with other preprocessing methods, and validation of the results show that MEAP produces reliable quantication of exons and alternative spliced variants. Analysis of data from head and neck squamous cell carcinoma (HNSCC) cell lines revealed several variants associated with 11q13 amplication, which is a predictive marker of metastasis and decreased survival in HNSCC patients. Together these results demonstrate the utility of MEAP in suggesting novel experimentally testable predictions. Thus, in addition to novel methodology to process large-scale exon array data sets, our results provide several HNSCC candidate genes for further studies.
Project description:We have sequenced miRNA libraries from human embryonic, neural and foetal mesenchymal stem cells. We report that the majority of miRNA genes encode mature isomers that vary in size by one or more bases at the 3’ and/or 5’ end of the miRNA. Northern blotting for individual miRNAs showed that the proportions of isomiRs expressed by a single miRNA gene often differ between cell and tissue types. IsomiRs were readily co-immunoprecipitated with Argonaute proteins in vivo and were active in luciferase assays, indicating that they are functional. Bioinformatics analysis predicts substantial differences in targeting between miRNAs with minor 5’ differences and in support of this we report that a 5’ isomiR-9-1 gained the ability to inhibit the expression of DNMT3B and NCAM2 but lost the ability to inhibit CDH1 in vitro. This result was confirmed by the use of isomiR-specific sponges. Our analysis of the miRGator database indicates that a small percentage of human miRNA genes express isomiRs as the dominant transcript in certain cell types and analysis of miRBase shows that 5’ isomiRs have replaced canonical miRNAs many times during evolution. This strongly indicates that isomiRs are of functional importance and have contributed to the evolution of miRNA genes
Project description:Gene expression profiling of immortalized human mesenchymal stem cells with hTERT/E6/E7 transfected MSCs. hTERT may change gene expression in MSCs. Goal was to determine the gene expressions of immortalized MSCs.