Project description:More insights into the character differences between ChAT+ and ChAT- NK cells were obtained based on the gene-expression patterns. A clear delineation between ChAT+ NK cells and ChAT- NK cells was observed, with a total of 300 genes over-expressed and 941 genes under-expressed significantly in the ChAT+ subset. It will provide evidences for further investigation into their functional characters
Project description:Purpose: Choline acetyltransferase (ChAT)-expressing epithelial cells are found in the upper and lower airways. In the trachea, they are referred to as brush cells. In the mouse nose two distinct populations of ChAT-eGFP+ cells had been previously described: a population of rare solitary chemosensory cells (SCCs) in the respiratory mucosa and a more abundant population of microvillous cells (MVCs) in the olfactory epithelium. Besides ChAT expression, SCCs share the expression of bitter taste receptors and signaling machinery with tracheal brush cells as well as close association with CGPR+ nerve fibers and an elongated shape. MVCs do not express bitter taste receptors, are not clearly associated with nerves and are smaller in size than SCCs. We have previously reported the transcriptional profile of tracheal ChAT-expressing brush cells but the transcriptional profile of nasal chemosensory ChAT+ epithelial cells has not been reported. Methods: In this study, we isolated nasal ChAT-eGFP+ cells by FACS from naïve ChAT(BAC)-eGFP mice with knockin of eGFP within a BAC spanning the ChAT locus, marking brush cells in the epithelium and performed transcriptome profiling using low input RNA sequencing. We sorted two distinct subsets of ChAT-eGFP+ epithelial cells from the nasal mucosa based on FACS ice and granularity characteristic: ChAT-eGFP+ EpCAM+ FSC/SSChigh (representing 5% of all ChAT-eGFP+ cells) and FSC/SSClow (representing 95% of all ChAT-eGFP+ cells), respectively. We compared them to unfractionated ChAT-eGFP- EpCAM+ epithelial cells. Results: Both nasal ChAT-eGFP subsets shared the core transcriptional profile of chemosensory cells from the intestine, trachea, gallbladder and thymus including the expression of Il25, Pou2f3, Trpm5, Avil, Plcb2 and transcripts of eicosanoid biosynthetic enzymes suggesting that most ChAT-eGFP+ cells in the nose belong to the chemosensory/tuft/brush cel family. The two subsets of nasal ChAT-eGFP+ cells differed in expression of taste receptors and taste receptor signaling machinery. Conclusions: Our study represents the first detailed analysis of the transcriptome of nasal ChAT-eGFP+ cells (brush cells) and identifies two subsets of nasal brush cells that share a core transcriptional signature but differ in expression of bitter taste receptors.
Project description:The goal of this study was to look at genes that were affected by 69-kDa and/or 82-kDa ChAT proteins in IMR32 cells Experiment Overall Design: The gene expression changes of IMR32 cells stably expressing either 69-kDa or 82-kDa ChAT proteins were anaylzed and compared to control IMR32 wild type cells. 3 biological replicates were anaylzed per condition (69-kDa ChAT expressing cells, 82-kDa ChAT expressing cells, or wild type IMR32 cells) for a total of 9 samples altogether.
Project description:Using chromatin immunoprecipitation and next-generation sequencing (ChIP-seq), we assessed the effects of acute exposure to oligomeric amyloid-beta on 82-kDa ChAT and SATB1 genome association in human SH-SY5Y neural cells, finding that Aβ-exposure increased 82-kDa ChAT and SATB1 association with gene promoters, introns and matrix attachment regions. We found that both SATB1 and 82-kDa ChAT associate with synapse and cell stress related genes after amyloid-beta exposure.
Project description:Group 2 innate lymphoid cells (ILC2s) reside in multiple tissues including lymphoid organs and barrier surfaces, and secrete type 2 cytokines including interleukin (IL)-5, IL-9 and IL-13. These cells participate in multiple physiological processes including allergic inflammation, tissue repair, metabolic homeostasis and host defense against helminth infections. Recent studies indicate that neuropeptides can play an important role in regulating ILC2 responses, however, the mechanisms that underlie these processes in vivo remain incompletely defined. Here, we identify that activated ILC2s upregulate choline acetyltransferase (ChAT)—the enzyme responsible for the biosynthesis of acetylcholine (ACh)—following infection with the helminth parasite Nippostrongylus brasiliensis or treatment with alarmins or cytokines including IL-25, IL-33 and thymic stromal lymphopoietin (TSLP). ILC2s also express acetylcholine receptors (AChRs), and ACh administration promotes ILC2 cytokine production and elicits expulsion of helminth infection. In accordance with this, ChAT deficiency in ILC2s leads to defective ILC2 responses and impaired immunity against helminth infection. Together, these results reveal a previously unrecognized role of the ChAT-ACh pathway in promoting type 2 innate immunity to helminth infection.
Project description:The mouse neuroblastoma N18TG2 clone is unable to differentiate and defective for the enzymes of the biosynthesis of neurotransmitters. The forced expression of choline acetyltransferase (ChAT) in these cells results in the synthesis and release of acetylcholine (Ach) and hence in the expression of neurospecific features and markers. To understand how the expression of ChAT triggered neuronal differentiation, we studied the differences in genome-wide transcription profiles between the N18TG2 parental cells and its ChAT-expressing 2/4 derived clone. The engagement of the 2/4 cells in the neuronal developmental programme was confirmed by the increase of the expression level of several differentiation-related genes and by the reduction of the amount of transcripts of cell-cycle genes. At the same time, we observed a massive reorganisation of cytoskeletal proteins in terms of gene expression, with the accumulation of the nucleoskeletal lamina component Lamin A/C in differentiating cells. The increase of the Lmna transcripts induced by ChAT-espression in 2/4 cells was mimicked treating the parental N18TG2 cells with the acetylcholine-receptor agonist carbachol, thus demonstrating the direct role played by this receptor in neuron nuclei maturation. Conversely, a treatment of 2/4 cells with the muscarinic-receptor antagonist atropine resulted in the reduction of the amount of Lmna RNA. Finally, the hypothesis that Lmna gene product might play a crucial role in the ChAT-dependent molecular differentiation cascade was strongly supported by Lmna knock-down in 2/4 cells leading to the down-regulation of genes involved in differentiation and cytoskeleton formation and to the up-regulation of genes known to regulate self-renewal and stemness. The gene expression profile of the ChAT-expressing 2/4 clone was compared to that of the N18TG2 parental cells, with and without retinoic acid (RA) stimulation using Agilent microarray analysis.