Project description:Epithelial cells possess remarkable plasticity, having the ability to become mesenchymal cells through alterations in adhesion and motility (epithelial-to-mesenchymal transition or EMT). Recent studies suggest that EMT endows differentiated epithelial cells with stem cell traits, posing the interesting question of how epithelial plasticity is properly restricted to ensure epithelial differentiation during tissue morphogenesis. Here we identify zinc-finger transcription factor Ovol2 as a key suppressor of EMT of mammary epithelial cells. Epithelia-specific deletion of Ovol2 completely arrests mammary ductal morphogenesis, and depletes epithelial stem/progenitor cell reservoirs. Further, Ovol2-deficient epithelial cells undergo EMT in vivo to become non-epithelial cell types, and that Ovol2 directly represses key EMT inducers such as Zeb1 and regulates stem/progenitor cell responsiveness to TGF-beta. We also provide evidence for a suppressive role of Ovol2 in breast cancer progression. Our findings underscore the critical importance of exquisitely regulating epithelial plasticity to balance stemness with epithelial differentiation in development and cancer. We report ChIPseq data illustrating Ovol2 genome-wide targets in mouse mammary epithelial cells, suggesting that Ovol2 regulates a plethora of genes associated with the EMT process. Immunoprecipitated samples from HC11 mouse mammary epithelial cells with antibodies against Ovol2 and control IgG respectively were used for ChIP-seq experiments.
Project description:Dermal fibroblasts from human, rhesus macaque, mouse and rat with and without dsRNA (poly I:C) stimulation (1ug/mL for 4 hours).<br>The innate immune response - the expression programme that is initiated once a pathogen is sensed - is known to be variable among responding cells, as well as to rapidly evolve in the course of mammal evolution. To study the transcriptional divergence and cell-to-cell variability of this response, we stimulated dermal fibroblast cells from two primates (human and macaque) and two rodents (mouse and rat) with dsRNA - a mimic of viral RNA that elicits a rapid innate immune response. Subsequently, we profiled the response using bulk RNA-seq, scRNA-seq and ChIP-seq across the four species and across different time points.
Project description:Natural epigenetic variation provides a source for the generation of phenotypic diversity, but to understand its contribution to phenotypic diversity, its interaction with genetic variation requires further investigation. MethylC-seq from naturally-occurring Arabidopsis accessions
Project description:We used micro-dissection with FACS sorting techniques to isolate single cells from the metanephric mesenchyme of the E11.5 developing kidney. A subset of these single cell populations is analysed individually via Fluidigm single cell analysis. This analysis will determine the transcriptional profile of each cell type, identify compartment specific transcripts, compartment specific transcript isoforms and cell-type specific long-noncoding RNAs. In addition the unbiased nature of RNA-SEQ will potentially identify novel transcripts that have not been annotated in the database. Kidneys are harvested from Tg(Crym-EGFP)GF82Gsat mice. Single cells are extracted from E11.5 metanephric mesenchyme using manual micro-dissection techniques. A subset of these cells is analyzed individually via Fluidigm single cell analysis. The long term goal is to generate a transcriptional atlas of the developing kidney.
Project description:Opioid analgesics are frequently prescribed in the United States and worldwide. However, serious side effects such as addiction, immunosuppression and gastrointestinal symptoms limit long term use. In the current study using a chronic morphine-murine model a longitudinal approach was undertaken to investigate the role of morphine modulation of gut microbiome as a mechanism contributing to the negative consequences associated with opioids use. The results revealed a significant shift in the gut microbiome and metabolome within 24 hours following morphine treatment when compared to placebo. Morphine induced gut microbial dysbiosis exhibited distinct characteristic signatures profiles including significant increase in communities associated with pathogenic function, decrease in communities associated with stress tolerance. Collectively, these results reveal opioids-induced distinct alteration of gut microbiome, may contribute to opioids-induced pathogenesis. Therapeutics directed at these targets may prolong the efficacy long term opioid use with fewer side effects.
Project description:Opioids such as morphine have many beneficial properties as analgesics, however, opioids may induce multiple adverse gastrointestinal symptoms. We have recently demonstrated that morphine treatment results in significant disruption in gut barrier function leading to increased translocation of gut commensal bacteria. However, it is unclear how opioids modulate the gut homeostasis. By using a mouse model of morphine treatment, we studied effects of morphine treatment on gut microbiome. We characterized phylogenetic profiles of gut microbes, and found a significant shift in the gut microbiome and increase of pathogenic bacteria following morphine treatment when compared to placebo. In the present study, wild type mice (C57BL/6J) were implanted with placebo, morphine pellets subcutaneously. Fecal matter were taken for bacterial 16s rDNA sequencing analysis at day 3 post treatment. A scatter plot based on an unweighted UniFrac distance matrics obtained from the sequences at OTU level with 97% similarity showed a distinct clustering of the community composition between the morphine and placebo treated groups. By using the chao1 index to evaluate alpha diversity (that is diversity within a group) and using unweighted UniFrac distance to evaluate beta diversity (that is diversity between groups, comparing microbial community based on compositional structures), we found that morphine treatment results in a significant decrease in alpha diversity and shift in fecal microbiome at day 3 post treatment compared to placebo treatment. Taxonomical analysis showed that morphine treatment results in a significant increase of potential pathogenic bacteria. Our study shed light on effects of morphine on the gut microbiome, and its role in the gut homeostasis.
Project description:We used micro-dissection with FACS sorting techniques to isolate renal vesicle single cell types from post natal (P4) kidneys. A subset of these single cell populations is analysed individually via Fluidigm single cell analysis. This analysis will determine the transcriptional profile of each cell type, identify compartment specific transcripts, compartment specific transcript isoforms and cell-type specific long-noncoding RNAs. In addition the unbiased nature of RNA-SEQ will potentially identify novel transcripts that have not been annotated in the database. Kidneys are harvested from Tg(Crym-EGFP)GF82Gsat mice. Single cells are extracted from P4 renal vesicles using micro-dissection with FACS sorting techniques. A subset of these cells is analyzed individually via Fluidigm single cell analysis. The long term goal is to generate a transcriptional atlas of the developing kidney.
Project description:We used micro-dissection and trypsinization techniques to isolate single cells from the E12.5 total kidney. A subset of these single cell populations is analysed individually via Fluidigm single cell analysis. This analysis will determine the transcriptional profile of each cell type, identify compartment specific transcripts, compartment specific transcript isoforms and cell-type specific long-noncoding RNAs. In addition the unbiased nature of RNA-SEQ will potentially identify novel transcripts that have not been annotated in the database. E12.5 kidneys are dissected; the kidneys are made into a single cell suspension via trypsinization. A subset of these cells is analysed individually via Fluidigm C1 single cell analysis. The long term goal is to generate a single cell resolution transcriptional atlas of the developing kidney.
Project description:Epithelial cells possess remarkable plasticity, having the ability to become mesenchymal cells through alterations in adhesion and motility (epithelial-to-mesenchymal transition or EMT). However, it is still unknown whether and how epithelial plasticity is kept in check in epithelial cells during development. Here we show that restricting the EMT of mammary epithelial cells by transcription factor Ovol2 is required for proper morphogenesis and regeneration. Deletion of Ovol2 blocks mammary ductal morphogenesis, depletes stem/progenitor cell reservoirs, and leads epithelial cells to undergo EMT in vivo to become non-epithelial cell types. Ovol2 directly represses myriad EMT inducers and its absence switches response to TGF-beta from growth arrest to EMT. Furthermore, forced expression of the repressor isoform of Ovol2 is able to reprogram metastatic breast cancer cells from a mesenchymal to an epithelial state. Our findings underscore the critical importance of exquisitely regulating epithelial plasticity in development and cancer. Refer to individual Series