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:Affymetrix miRNA arrays were used to generate miRNA profiles of peripheral blood leukocytes and FACS sorted neutrophils, monocytes, B-cells, T-cells, CD4+ T-cells, and CD8+ T-cells, being the major leukocyte cell types in human. The study allowed for the determination of the miRNAs that were expressed in each leukocyte cell subtype. Two-way hierarchical clustering on the miRNAs and samples illustrated that miRNA expression profiles of B- and T-cells were very much alike, and that there there were a number of miRNAs which appeared to have an expression profile specific to certain leukocyte cell subtypes. This study will facilitate the identification of microRNAs associated with and contributing to single leukocyte cell subtypes. Peripheral blood leukocytes were extracted, and neutrophils, monocytes, B-cells, T-cells, CD4+ T-cells, and CD8+ T-cells were FACS sorted from total blood from a healthy subject. In order to determine the miRNA expression profile of these cells, RNA was extracted, labeled and hybridized on an Affymetrix miRNA array.
Project description:Affymetrix miRNA arrays were used to generate miRNA profiles of peripheral blood leukocytes and FACS sorted neutrophils, monocytes, B-cells, T-cells, CD4+ T-cells, and CD8+ T-cells, being the major leukocyte cell types in human. The study allowed for the determination of the miRNAs that were expressed in each leukocyte cell subtype. Two-way hierarchical clustering on the miRNAs and samples illustrated that miRNA expression profiles of B- and T-cells were very much alike, and that there there were a number of miRNAs which appeared to have an expression profile specific to certain leukocyte cell subtypes. This study will facilitate the identification of microRNAs associated with and contributing to single leukocyte cell subtypes.
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 Sequence library of miRNAs from a single sample of human foetal mesenchymal stem cells. Results tested and confirmed by northern blotting. Please note that only raw data files are available for the embryonic and neual samples and thus, directly submitted to SRA (SRX547311, SRX548700, respectively under SRP042115/PRJNA247767)
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.
Project description:Transcriptional profiling of human mesenchymal stem cells comparing normoxic MSCs cells with hypoxic MSCs cells. Hypoxia may inhibit senescence of MSCs during expansion. Goal was to determine the effects of hypoxia on global MSCs gene expression.
Project description:Gene methylation profiling of immortalized human mesenchymal stem cells comparing HPV E6/E7-transfected MSCs cells with human telomerase reverse transcriptase (hTERT)- and HPV E6/E7-transfected MSCs. hTERT may increase gene methylation in MSCs. Goal was to determine the effects of different transfected genes on global gene methylation in MSCs.
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. One-condition experment, gene expression of 3A6