Project description:The mammary epithelial cell (MEC) system is a bi-layered ductal epithelial network consisting of luminal and basal cells, maintained by a lineage of stem and progenitor cell populations. Here, we used integrated single-cell transcriptomics and chromatin accessibility analysis to reconstruct the cell types of the mouse MEC system and their underlying gene regulatory features in an unbiased manner. We define differentiation states within the secretory type of luminal cells, which can be defined as a continuous spectrum of progenitor and mature secretory cells. By integrating single-cell transcriptomics and chromatin accessibility landscapes, we identified cis- and trans-regulatory elements that are differentially activated in the specific epithelial cell types and our newly defined luminal differentiation states. Our work provides an unprecedented resource to reveal cis/trans regulatory elements associated with MEC identity and differentiation that will serve as a valuable reference to determine how the chromatin accessibility landscape changes during breast cancer.
2020-09-14 | GSE157888 | GEO
Project description:Mircrobial community in MEC biogas upgrading system
Project description:The mammary epithelial cell (MEC) system is a bi-layered ductal epithelial network consisting of luminal and basal cells, maintained by a lineage of stem and progenitor cell populations. Here, we used integrated single-cell transcriptomics and chromatin accessibility analysis to reconstruct the cell types of the mouse MEC system and their underlying gene regulatory features in an unbiased manner. We define differentiation states within the secretory type of luminal cells, which can be defined as a continuous spectrum of progenitor and mature secretory cells. By integrating single-cell transcriptomics and chromatin accessibility landscapes, we identified cis- and trans-regulatory elements that are differentially activated in the specific epithelial cell types and our newly defined luminal differentiation states. Our work provides an unprecedented resource to reveal cis/trans regulatory elements associated with MEC identity and differentiation that will serve as a valuable reference to determine how the chromatin accessibility landscape changes during breast cancer.
Project description:Several methicillin resistance (SCCmec) clusters characteristic of hospital-associated methicillin-resistant Staphylococcus aureus (MRSA) strains harbor the psm-mec locus. In addition to encoding the cytolysin, phenol-soluble modulin (PSM) mec, this locus has been attributed gene regulatory functions. Here we employed genome-wide transcriptional profiling to define the regulatory function of the psm-mec locus. The immune evasion factor protein A emerged as the primary conserved and strongly regulated target of psm-mec, an effect we show is mediated by the psm-mec RNA. Furthermore, the psm-mec locus exerted regulatory effects that were more moderate in extent and possibly mediated by the PSM-mec peptide. For example, expression of PSM-mec limited expression of mecA, thereby decreasing methicillin resistance. Our study shows that the psm-mec locus has a rare dual regulatory RNA and encoded cytolysin function, both with the potential to enhance MRSA virulence. Furthermore, our findings reveal a specific mechanism underscoring the recently emerging concept that S. aureus strains balance pronounced virulence and high expression of antibiotic resistance.
Project description:Nociceptive neurons develop a complex dendritic arbor to sense noxious stimuli, which enables animals to react to environmental insults and perform self-protective behaviours. The genetic programs controlling neuronal dendritic morphogenesis are poorly understood. In C. elegans, the PVD sensory neuron generates a complex dendritic arbor that envelops the body of the animal. This nociceptive neuron enables study of dendrite formation in vivo. MEC-3 is a transcription factor expressed in the PVD neuron (Chatzigeorgiou et al., 2010; Li et al., 2011; Way and Chalfie, 1989). mec- 3 mutant PVD neurons fail to elaborate lateral branches and show defective function (Smith et al., 2010; Tsalik et al., 2003) (Husson et al., 2012). We used tiling arrays to identify genes regulated by the mec-3 transcription factor in PVD sensory neurons. We employ the mRNA-tagging method to isolate poly(A) RNA from the PVD and OLL neurons in wildtype and mec-3 mutants (3 replicates each) by expressing a 3X FLAG-tagged poly(A) binding protein PAB-1 in PVD and OLL under control of the ser-2prom3B promoter. The enriched poly(A) RNA is amplified using the NuGEN WT-Pico amplification system and applied to Affymetrix C. elegans tiling microarrays. Whole animal reference samples were obtained by isolating total RNA at the L2/L3 stages in wildtype and mec-3 mutants (2 replicates each) and processed as above.
Project description:Three-dimensional (3D) organoid models have been instrumental in understanding molecular mechanisms responsible for many cellular processes and diseases. However, established organic biomaterial scaffolds used for 3D hydrogel cultures, such as Matrigel, are biochemically complex and display significant batch variability, limiting reproducibility in experiments. Recently, there has been significant progress in the development of synthetic hydrogels for in vitro cell culture that are reproducible, mechanically tuneable, and biocompatible. Self-assembling peptide hydrogels (SAPHs) are synthetic biomaterials that can be engineered to be compatible with 3D cell culture. Here we investigate the ability of PeptiGel® SAPHs to model mammary epithelial cell (MEC) microenvironment in vitro. The positively charged PeptiGel®Alpha4 supported MEC viability, but did not promote formation of polarised acini. Modifying the stiffness of PeptiGel®Alpha4 stimulated changes in MEC viability and changes in protein expression associated with altered MEC function, but did not fully recapitulate the morphologies of MECs grown in Matrigel. To supply the appropriate biochemical signals for MEC organoids, we supplemented PeptiGels® with laminin. Laminin was found to require negatively charged PetiGel®Alpha7 for functionality, but was then able to provide appropriate signals for correct MEC polarisation and expression of characteristic proteins. Thus, optimisation of SAPH composition and mechanics allows tuning to support tissue-specific organoids.
Project description:Neural circuits in the medial entorhinal cortex (MEC) encode an animal’s position and orientation in space. Within the MEC spatial representations, including grid and directional firing fields, have a laminar and dorsoventral organization that corresponds to a similar topography of neuronal connectivity and cellular properties. Yet, in part due to the challenges of integrating anatomical data at the resolution of cortical layers and borders, we know little about the molecular components underlying this organization. To address this we develop a new computational pipeline for high-throughput analysis and comparison of in situ hybridization (ISH) images at laminar resolution. We apply this pipeline to ISH data for over 16,000 genes in the Allen Brain Atlas and validate our analysis with RNA sequencing of MEC tissue from adult mice. We find that differential gene expression delineates the borders of the MEC with neighboring brain structures and reveals its laminar and dorsoventral organization. Our analysis identifies ion channel-, cell adhesion- and synapse-related genes as candidates for functional differentiation of MEC layers and for encoding of spatial information at different scales along the dorsoventral axis of the MEC. Our results support the hypothesis that differences in gene expression contribute to functional specialization of superficial layers of the MEC and dorsoventral organization of the scale of spatial representations.