Project description:To understand gene expression signatures between wild-type and Satb1-deficient cells To examine genes regulated by Satb1 expression, sets of microarrays were conducted with Lin- c-kitHi Sca-1+ Flt3- HSC-enriched cells, Lin- c-kitHi Sca-1+ Flt3+LMPP-enriched cells, and Lin- c-kitLo Sca-1Lo IL7Rα+ CLP-enriched cells derived from E18.5 FL of Satb1-null mice or their WT littermates.
Project description:Hematopoietic stem cells (HSCs) are now recognized as a heterogeneous population in self-renewing and differentiation capabilities. However, fundamental mechanisms governing the heterogeneity remain uncertain. We here show that special AT-rich sequence-binding protein 1 (SATB1), a global chromatin organizer, is involved in the mechanisms. Analyzing hematological lineage-restricted SATB1 knock out mice proved that SATB1 is indispensable for both self-renewal and normal differentiation of adult HSCs. Using SATB1/Tomato knock-in mice, we subdivided HSCs according to SATB1 intensity. Culture experiments and RNA-sequencing revealed essential differences between SATB1- and SATB1+ HSCs regarding lineage potential.
Project description:CD4+ and CD8+ double-positive (DP) thymocytes are at a critical stage during the T cell development in thymus. DP cells rearrange the T cell receptor gene Tcra to generate T cell receptors with TCR. Then DP cells differentiate into CD4 or CD8 single-positive (SP) thymocytes, Regulatory T cells, or invariant nature kill T cells (iNKT) according to the TCR signal. Chromatin organizer SATB1 is highly expressed in DP cells and plays an essential role in regulating Tcra rearrangement and differentiation of DP cells. Here we explored the mechanism of SATB1 orchestrating gene expression in DP cells. Single-cell RNA sequencing assay of SATB1-deficient thymocytes showed that the cell identity of DP thymocytes was changed, and the genes specifically highly expressed in DP cells were down-regulated. ChIP-seq and ATAC-seq data showed the similar tendency. The super-enhancers regulate the expressions of the DP-specific genes, and the SATB1 deficiency reduced the super-enhancer activity. Hi-C data showed that interactions in super-enhancers and between super-enhancers and promoters decreased in SATB1 deficient thymocytes. We further explored the regulation mechanism of two SATB1-regulating genes, ETS2 and Bcl6, in DP cells and found that the knockout of the super-enhancers of these two genes impaired the development of DP cells. Our research reveals that SATB1 globally regulates super-enhancers of DP cells and promotes the establishment of DP cell identity, which helps understand the role of SATB1 in thymocyte development.
Project description:To understand gene expression signatures between wild-type and SFRP5-overexpressing cells To examine genes regulated by SFRP5 expression, sets of microarrays were conducted with Lin- c-kitHi Sca-1+ cells and Lin- c-kitLo Sca-1Lo IL7Rα+ CLP-enriched cells derived from adult bone marrow of SFRP5-transgenic mice or their WT littermates.
Project description:Our data show Satb1 deficiency leads to alterations in DNA cytosine methylation and a commitment-primed epigenetic state in HSCs. Examination of DNA cytosine methylation in wild type HSC and differentiation-committed progenitors as well as in wild type HSC and HSC lacking Satb1 (n=2 each).
Project description:Gene expression analysis on purified murine hematopoietic stem cells (HSCs) deficient for Special AT-rich sequence-binding protein 1 (Satb1) compared to wild-type HSCs. Analysis of gene expression of bone marrow-derived CD45.2+ CD150+cKit+Sca-1+CD4-CD8a-CD19-B220-Gr-1- HSCs from congenic recipient animals transplanted >20 weeks with either wild-type or Satb1-deficient hematopoeitic cells.
Project description:Hut78 cell line was used as Sezary syndrome cell model. Comparative transcriptome profiles of SATB1 transduced Hut78 cells (Hut78-SATB1) relative to empty MIG vector transduced control Hut78 cells (Hut78-MIG) were analyzed. The primary goal is to establish a list of genes with differential expression between SATB1 transduced Hut78 cells and control Hut78 cells to identify the gene expression regulation effect of SATB1 expression in Sezary cells. Two color experiment, 3 biological replicates (3 unique cell clones) with Hut78 cells, each compared with a distinct individual clone from cells transduced with empty MIG vector.
Project description:T-cell receptor (TCR) signaling by MHC class-I and -II induces thymocytes to acquire cytotoxic and helper fates via induction of Runx3 or ThPOK transcription factors, respectively. The mechanisms by which TCR signaling is translated into transcriptional programs for each cell fate remain elusive. We show that a genome organizer, Satb1, activates genes for lineage-specifying factors, including ThPOK and Runx3, in post-selection thymocytes. Indeed, Satb1-deficient thymocytes are partially redirected to inappropriate T lineages after MHC selection. Although Satb1 is dispensable for maintaining ThPOK in CD4+ T cells, it is necessary for restraining expression of the Treg factor FoxP3. Collectively, our findings demonstrate that Satb1 shapes the primary T-cell pool by initially directing lineage-specific transcriptional programs and subsequently balancing effector versus regulatory subsets via FoxP3 repression.
Project description:Hut78 cell line was used as Sezary syndrome cell model. Comparative transcriptome profiles of SATB1 transduced Hut78 cells (Hut78-SATB1) relative to empty MIG vector transduced control Hut78 cells (Hut78-MIG) were analyzed. The primary goal is to establish a list of genes with differential expression between SATB1 transduced Hut78 cells and control Hut78 cells to identify the gene expression regulation effect of SATB1 expression in Sezary cells.