Project description:CD34+ CD117+ cells in human peripheral blood have mast cell-forming capacity. We have identified highly committed mast cell progenitors as Lin- CD34+ CD117 intermediate/high FcεRI+ cells. To explore the gene expression profile of the highly committed mast cell progenitors, we performed whole-transcriptome microarray analyses of the cells and compared them with mature basophils.
Project description:To investigate the mechanisms by which C/EBPa drives basophil differentiation and maintains basophil identity, we examined whether or not C/EBPa promotes basophil molecular programming and simultaneously represses mast cell molecular programming. We performed genome-wide gene expression profiling on basophils and mast cells and found that 6798 genes were shared by mast cells and basophils; 2033 genes were expressed 2-10 (log2 1-3.3)-fold higher in basophils (differentially expressed in basophils); and 413 genes were expressed greater than 10 (log2 3.3)-fold in basophils (highly expressed in basophils). On the other hand, we found 569 genes were expressed 2-10 (log2 -1 to -3.3) fold higher in mast cells and 171 genes were highly expressed in mast cells [greater than 10 fold (log2 -3.3)]. We treated purified basophils prepared from Cebpaf/f RosaYFP/creER mice and Cebpa+/+ RosaYFP/creER control mice with or without 4HT treatment for five days. Gene expression in the treated basophils was analyzed using microarray analysis. Overall, deletion of C/EBPa in basophils resulted in a reduction of mRNA expression for 248 genes and led to an increase in mRNA expression for 255 genes. The majority of the C/EBPa-regulated genes were either differentially or highly expressed in basophils or mast cells. In this study, we compared gene expression in basophils and mast cell and identified genes which specifically expressed in basophils and mast cells. By using Cebpa conditional knock out mice, we identified Cebpa regulated genes in basophils.
Project description:In this study, we explored gene expression profiles of CD34-CLEC12Ahi pre-basophils isolated from the bone marrow as well as CD34-CLEC12Alo mature basophils from the bone marrow and CD34+ basophil progenitors. We revealed that the gene expression of mature basophils isolated from the bone marrow and spleen resembled each other whereas pre-basophils and mature basophils showed distinct gene expression profiles.
Project description:E-cadherin is a calcium-dependent cell-cell adhesion molecule extensively studied for its involvement in tissue formation, epithelial cell behavior and suppression of cancer; however its expression in the hematopoietic system hasn't received much attention. Combining single-cell RNA sequencing analyses and immunophenotyping, we revealed expression of E-cadherin in the basophil and mast cell lineages. No defect in basophil and mast cell differentiation was observed in mice lacking E-cadherin in the hematopoietic system, supporting that E-cadherin is not required per se for basophil and mast cell differentiation. Yet, we evidenced that granulocyte-monocyte progenitors expressing high levels of E-cadherin have an enriched capacity to differentiate into basophils and mast cells. Importantly, E-cadherin expression is observed on committed progenitors prior to the expression of other reported markers of these lineages. We named those progenitors pro-BMPs (pro- basophil and mast cells progenitors). Using RNA-sequencing, we demonstrated the transcriptional priming of pro-BMPs to the basophil and mast cell lineages.
Project description:Basophils are the least common granulocytes which represent less than 1% of peripheral blood leukocytes. Recent development of analytical tools for basophils has enabled us to understand their critical roles in allergic reactions and protection from parasitic infections. Nevertheless, the differentiation trajectory of basophils remain unclear. To explore differentiation trajectory of basophils, we conducted highly sensitive single cell RNA sequencing (scRNA-seq) of bone marrow-derived basophils (BMBAs) and basophils under homeostatic conditions. As a result, we identified previously-unappreciated pre-basophils which are located down-stream of bi-potential basophil and mast cell common precursors and upstream of mature basophils along the ontogeny of basophlis.
Project description:To investigate the mechanisms by which C/EBPa drives basophil differentiation and maintains basophil identity, we examined whether or not C/EBPa promotes basophil molecular programming and simultaneously represses mast cell molecular programming. We performed genome-wide gene expression profiling on basophils and mast cells and found that 6798 genes were shared by mast cells and basophils; 2033 genes were expressed 2-10 (log2 1-3.3)-fold higher in basophils (differentially expressed in basophils); and 413 genes were expressed greater than 10 (log2 3.3)-fold in basophils (highly expressed in basophils). On the other hand, we found 569 genes were expressed 2-10 (log2 -1 to -3.3) fold higher in mast cells and 171 genes were highly expressed in mast cells [greater than 10 fold (log2 -3.3)]. We treated purified basophils prepared from Cebpaf/f RosaYFP/creER mice and Cebpa+/+ RosaYFP/creER control mice with or without 4HT treatment for five days. Gene expression in the treated basophils was analyzed using microarray analysis. Overall, deletion of C/EBPa in basophils resulted in a reduction of mRNA expression for 248 genes and led to an increase in mRNA expression for 255 genes. The majority of the C/EBPa-regulated genes were either differentially or highly expressed in basophils or mast cells.
Project description:Haematopoiesis is a multi-stage process that involves the differentiation of multipotent stem cells and progenitor cells into distinct mature cell lineages. Here we present Haemopedia, a comprehensive atlas of murine gene expression data that covers all the mature lineages in haematopoiesis, including rare cell populations such as eosinophils, mast cells, basophils and megakaryocytes and a large collection of progenitor and stem cells. We have used this dataset to identify gene sets with specifically enriched expression in each of the mature blood cell lineages. Many of the genes in these sets show conserved lineage-enriched expression patterns in human haematopoiesis. We also identify some genes with divergent expression patterns between mouse and human, highlighting species specific differences in blood cell production. To make analyses of Haemopedia and other blood cell transcriptional datasets easier, we have created an online web portal, Haemosphere, which provides simple tools to interrogate gene expression-based relationships between haematopoietic cell types and genes of interest. Total RNA was obtained from 54 haematopoetic cells types and 8 non-haematopoietic out groups.
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: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)