Project description:Analysis of the RNA-seq data performed in IR vs NIR hematopoietic stem cells show the loss of the TNF_via_NFKB signature. We showed that the loss of this signature could be associated with H3K9me3 loss at specific retrotransposable elements . To validate this association, we tested if TNFa treatment before irradiation was able to prevent IR-effect on H3K9me3 loss at retrotransposable elements. For this purpose, we treated mice with TNFa 1h before irradiation (IR_TNF) and performed H3K9me3 cut&tag experiments on hematopoietic stem cells 1 month after irradiation and compared them to hematopoietic stem cells sorted from non irradiated mice (NIR) and from non-treated irradiated mice (IR).
Project description:Using UNC0638 and genetic assays to inhibit EHMT1/2 and derepress fetal hemoglobin in adult hematopoietic cells. ChIP-seq for 2 histone modifications and total H3 (for quantitative normalization) in biological triplicates of cord blood, adult bone marrow, and UNC0638-treated adult bone marrow.
Project description:Acute megakaryoblastic leukemia of Down syndrome (DS-AMKL) is a model of clonal evolution from a preleukemic transient myeloproliferative disorder requiring both a trisomy 21 (T21) and a GATA1s mutation to a leukemia driven by additional driver mutations. We modelled this leukemic evolution through stepwise gene editing of GATA1s, SMC3+/- and MPLW515K providing 20 different trisomy or disomy 21 iPSC clones. Cut&Tag against GATA1 was performed on CD41+CD42+ megakaryocytes obtained after 18 days of differentiation of the IPSC clones. 500,000 CD41+CD42+ MK sorted cells were used to analyze GATA1 and GATA1s chromatin occupancy using the CUT&Tag-IT Assay Kit (Active Motif) according to the manufacturer recommendations. Briefly, cells were bound to Concanavalin A-Coated Beads and incubated with primary anti-GATA1 antibody in buffer with Protease Inhibitor Cocktail and 5% digitonine overnight at 4°C under rotation. The Guinea Pig anti-rabbit secondary antibody was incubated in Dig-Wash buffer for 1 hour at RT under rotation. After 3 washes, the CUT&Tag-IT™ Assembled pA-Tn5 Transposomes (1:100) were added for 1 hour at RT under rotation and tagmentation was performed during 1 hour at 37°C. DNA was purified and libraries were generated by PCR. The final libraries were purified, pooled together in equal concentrations and subjected to paired-end sequencing (100 cycles: 2x50) in Novaseq-6000 sequencer (Illumina) at Gustave Roussy.
Project description:Contrasting H3K9Me3 binding site number and localisation between activated and quiescent hepatic stellate cells to identify regulation of gene repression.
Project description:We investigated whether exogenous supplies of different IL-2R agonists (IL-2wt, IL-2nα or IL-2α-bias) at the concentration of 3 mg/g body weight could rescue the transcriptional signatures of dysfunctional T cells in large tumors, or allow them to regain sensitivity to αPD-1 antibody. Remarkably, after transient stimulation by IL-2wt, large tumors fully restored the effector/activation signatures observed in small tumors, such as upregulation of effector genes (Gzma, Gzmb, Gzmk and Ifng), T cell activation/exhaustion genes (Ctla4, Pdcd1, Lag3 and Havcr2), inflammatory cytokine/cytokine receptors (Il2ra, Il2rb, Il2rg and Il21r), and chemokines (Cxcl9, Cxcl10 and Ccl19) (Fig.7d). Surprisingly, although IL-2α-bias had >10-fold weaker in vitro activity than IL-2wt, and could only activate the small fraction of CD25-upregulated CD8+Teff cells, the transcriptional profiles were indistinguishable between IL-2wt and IL-2α-bias treated large tumors. In stark contrast, IL-2nα treatment showed no meaningful transcriptional changes in large tumors compared to untreated control. These results once again demonstrate the critical role of CD25 in transmitting IL-2 signaling to reprogram the tumor immune microenvironment.
Project description:Sox2 is a master transcriptional regulator of embryonic development. Having found that Sox2 interacts with RNA-binding proteins, we designed an experiment to discover RNAs associated with Sox2 and other pluripotency factors. Briefly, we used RNA immunoprecipitation followed by high-throughput sequencing (RIP-seq) to sequence transcriptomes enriched for RNAs associated with Klf4, Nanog, Oct4, Sox2 and Suz12. For completeness, this submission includes all RIP-seq data from this study, although some of the data was only used for exploratory analyses. Such analysis indicated that it was important to include input samples for each of the the cell lines that were to be compared, to account for differences in gene expression (e.g. between J1-birA with and without bioSox2), and that overnight IP was more informative than 3 h IP. Therefore, the analysis described in the associated manuscript used samples from experiment batch 3 and 4 only (samples 11-18).
Project description:Cervical cancer (CC) remains a major cause of cancer-related mortality, particularly in regions with limited screening access, despite being highly preventable and treatable when detected early. MSX1, a homeobox transcription factor with dual roles as a tumor suppressor and oncogene, has an unclear role in CC pathogenesis. This study reveals that MSX1 acts as a tumor-promoting factor in CC, with de novo expression observed in precancerous lesions but absent in normal cervical epithelium. MSX1 enhances clonogenicity and migrationin cervical cancer cells, driven by epithelial-to-mesenchymal transition (EMT) induction. Mechanistically, MSX1 activates RHO/RAC/CDC42 cytoskeletal signaling pathways, with FOS—a downstream RHO effector—identified as a key mediator of CC aggressiveness. Targeting RHO signaling or FOS reverses MSX1-driven aggressive phenotypes, while proteasomal degradation of MSX1 reduces chemoresistance. These findings highlight MSX1’s critical role in CC progression and suggest its potential as a therapeutic target. The study underscores MSX1’s involvement in key oncogenic pathways, offering new insights for developing targeted therapies in cervical cancer.
Project description:Identification of open chromatin regions distinguishing primary human fibroblasts from abdomen, breast, and lung during proliferation or after 40 Gy X-irradiation, or after IL1-alpha or 1L1-beta treatments.