Transcriptional regulation in the absence of inositol trisphosphate receptor signaling
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ABSTRACT:
PROVIDER: PRJNA1193071 | ENA |
REPOSITORIES: ENA
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Project description:The activation of IP3 receptor (IP3R) Ca2+ channels generates agonist-mediated Ca2+ signals that are critical for the regulation of a wide range of biological processes. It is therefore surprising that CRISPR induced loss of all three IP3R isoforms (TKO) in HEK293 and HeLa cell lines yields cells that can survive, grow and divide, albeit more slowly than wild-type cells. In an effort to understand the adaptive mechanisms involved, we have examined the activity of key Ca2+ dependent transcription factors (NFAT, CREB and AP-1) and signaling pathways using luciferase-reporter assays, phosphoprotein immunoblots and whole genome transcriptomic studies. In addition, the diacylglycerol arm of the signaling pathway was investigated with protein kinase C (PKC) inhibitors and siRNA knockdown. The data showed that agonist-mediated NFAT activation was lost but CREB activation was maintained in IP3R TKO cells. Under base-line conditions transcriptome analysis indicated the differential expression of 828 and 311 genes in IP3R TKO HEK293 or HeLa cells, respectively, with only 18 genes being in common. Three main adaptations in TKO cells were identified in this study: 1) increased basal activity of NFAT, CREB and AP-1; 2) an increased reliance on Ca2+- insensitive PKC isoforms; and 3) increased production of reactive oxygen species and upregulation of antioxidant defense enzymes. We suggest that whereas wild-type cells rely on a Ca2+ and DAG signal to respond to stimuli, the TKO cells utilize the adaptations to allow key signaling pathways (e.g. PKC, Ras/MAPK, CREB) to transition to the activated state using a DAG signal alone.
2024-12-03 | GSE283304 | GEO
Project description:Elevated inositol trisphosphate receptor (IP3R) levels have been previously reported in skeletal muscle myotubes derived from patients with ryanodine receptor 1 (RyR1) mutation related core myopathies. However, the functional relevance and the relationship of IP3R mediated Ca2+ signalling with the pathophysiology of the disease is unclear. It has also been suggested that mitochondrial dysfunction underlies the development of central and diffuse multi-mini-cores, devoid of mitochondrial activity, a key pathological consequence of RyR1 mutations. Here we used muscle biopsies of central core and multi-minicore disease patients with RyR1 mutations, as well as cellular and in vivo mouse models of the disease to characterise whole genome and mitochondrial gene expression to assess if remodeling of skeletal muscle following loss of functional RyR1 mediates bioenergetic adaptation.
2020-09-15 | GSE103854 | GEO
Project description:Elevated inositol trisphosphate receptor (IP3R) levels have been previously reported in skeletal muscle myotubes derived from patients with ryanodine receptor 1 (RyR1) mutation related core myopathies. However, the functional relevance and the relationship of IP3R mediated Ca2+ signalling with the pathophysiology of the disease is unclear. It has also been suggested that mitochondrial dysfunction underlies the development of central and diffuse multi-mini-cores, devoid of mitochondrial activity, a key pathological consequence of RyR1 mutations. Here we used muscle biopsies of central core and multi-minicore disease patients with RyR1 mutations, as well as cellular and in vivo mouse models of the disease to characterise whole genome and mitochondrial gene expression to assess if remodeling of skeletal muscle following loss of functional RyR1 mediates bioenergetic adaptation.
2020-09-15 | GSE103853 | GEO
Project description:Inositol 1,4,5-trisphosphate 3-kinase A (IP3K-A) is a molecule enriched in the brain and neurons that regulates intracellular calcium levels via signaling through the inositol trisphosphate receptor. In the present study, we found that IP3K-A expression is highly enriched in the central nucleus of the amygdala (CeA), which plays a pivotal role in the processing and expression of emotional phenotypes in mammals. We used microarray to identify differentially expressed genes in the amydala of wild type (WT) and IP3K-A KO mice.
2016-01-21 | GSE77001 | GEO
Project description:Intracellular signaling is conserved in eukaryotes to allow for response to extracellular signals and to regulate development and cellular functions. In fungi, inositol phosphate signaling has been shown to be involved in growth, sexual reproduction, and metabolic adaptation lacking, however, reports of mushroom forming fungi so far. In Schizophyllum commune, an inositol monophosphatase has been found up-regulated during sexual development. The enzyme is crucial for inositol cycling where it catalyzes the last step of inositol phosphate metabolism restoring the inositol pool from the monophosphorylated inositol monophosphate. We overexpressed the gene in this model basidiomycete and verified its involvement in cell wall integrity and intracellular trafficking. Strong phenotypes in mushroom formation and cell metabolism were evidenced by proteome analyses. In addition, altered inositol signaling was shown to be involved in tolerance towards cesium and zinc, and increased metal tolerance towards cadmium, associated with induced expression of kinases and repression of phosphatases within the inositol cycle. The presence of the heavy metals Sr, Cs, Cd, and Zn lowered intracellular calcium levels. We could develop a model integrating inositol signaling in the known signal transduction pathways governed by Ras, G-protein coupled receptors, cAMP and elucidate different roles in development.
2021-07-01 | PXD026465 | Pride
Project description:Contactin-associated protein-like 2 (Caspr2) is a neurexin-like protein that has been associated with numerous neurological conditions. However, the mechanisms underlying Caspr2 function in the central nervous system remain incompletely understood. Here, we report on a functional role for Caspr2 in the developing cerebellum. Loss of Caspr2 impairs Purkinje cell dendritic development, alters cell signaling and results in motor coordination deficits. Caspr2 is highly enriched at synaptic specializations in the cerebellum. Using a proteomic approach, we identify type 1 inositol 1,4,5-trisphosphate receptor (IP3R1) as a specific synaptic interaction partner of the Caspr2 extracellular domain (ECD) in the molecular layer (ML) of the developing cerebellum. The interaction of Caspr2 ECD with IP3R1 inhibits IP3R1-mediated changes in cellular morphology. Together, our work defines a mechanism by which Caspr2 controls the development and function of the cerebellum, and advances our understanding of how Caspr2 dysfunction might lead to specific brain disorders.
2020-07-13 | PXD018972 | Pride
Project description:Inositol Trisphosphate Receptor Mediated Ca2+ Signalling Stimulates Mitochondrial Function and Gene Expression in Core Myopathy Patients
| PRJNA407299 | ENA
Project description:Inositol 1,4,5-trisphosphate 3-kinase A (IP3K-A) is a molecule enriched in the brain and neurons that regulates intracellular calcium levels via signaling through the inositol trisphosphate receptor. In the present study, we found that IP3K-A expression is highly enriched in the central nucleus of the amygdala (CeA), which plays a pivotal role in the processing and expression of emotional phenotypes in mammals. We used microarray to identify differentially expressed genes in the amydala of wild type (WT) and IP3K-A KO mice. Male IP3K-A KO mice and their wild-type (WT) littermate controls from a C57BL/6N background were used at 10 to 15 weeks old. Mice were housed in temperature (22°C-23°C)- and humidity (50%)-controlled quarters under a 12-hr light/dark photoperiod (lights on at 08:00a.m.) with free access to food and water. Mice were sacrificed by cervical dislocation between 02:00p.m. and 04:00p.m. Amygdala tissue was rapidly dissected from 1-mm thick slices and frozen in liquid nitrogen.
2016-01-21 | E-GEOD-77001 | biostudies-arrayexpress
Project description:Cryptococcus neoformans is the most common cause of fungal meningitis, with high mortality and morbidity. The reason for the frequent occurrence of Cryptococcus infection in the central nervous system (CNS) is poorly understood. In this study, we find that inositol plays an important role in the transversal of Cryptococcus across the blood-brain barrier (BBB) both in an in vitro human BBB model and in vivo animal models. The inositol stimulation of BBB crossing is dependent upon fungal inositol transporters. The upregulation of genes involved in the inositol catabolism pathway is evident in a microarray analysis. The expression of CPS1, a gene encoding the hyaluronic acid synthase in Cryptococcus, is also upregulated by the inositol treatment. The production of hyaluronic acid increased in cells treated with inositol, which leads to the enhanced binding ability of Cryptococcus cells to the human brain microvascular endothelial cells (HBMECs) constituting the BBB. Overall, our studies provide a mechanism for inositol-dependent Cryptococcus transversal of the BBB, supporting our hypothesis that host inositol utilization by the fungus contributes to Cryptococcus CNS infection. To understand the effect of inositol on gene expression profiles during cell development, microarray experiments were performed to monitor the genes regulated by inositol. H99 overnight culture was washed with dH2O once, and cells were inoculated on SD medium with or without inositol. Cells were collected from SD plates 24 hr post-inoculation, washed with dH2O, and total RNA was purified. Total RNAs were extracted using Trizol Reagents (Invitrogen) and purified using the Qiagen RNeasy cleanup kit (Qiagen). Cy3 and Cy5-labeled cDNA were generated by incorporating amino-allyl-dUTP during reverse transcription of 5 µg of total RNA as described previously and competitively hybridized to a JEC21 whole-genome array generated previously at Washington University in Saint Louis. After hybridization, arrays were scanned with a GenePix 4000B scanner (Axon Instruments, http://www.axon.com) and analyzed by using GenePix Pro version 4.0 and BRB array tools (developed by Richard Simon and Amy Peng Lam at the National Cancer Institute; http://linus.nci.nih.gov/BRB-ArrayTools.html)
2012-09-29 | E-GEOD-41211 | biostudies-arrayexpress
Project description:Messenger (m)RNA export from the nucleus is essential for eukaryotic gene expression. Here, we identify a transcript-selective nuclear export mechanism affecting certain human transcripts, enriched for functions in genome duplication and repair, controlled by inositol polyphosphate multikinase (IPMK), an enzyme catalyzing inositol polyphosphate and phosphoinositide turnover. We studied transcripts encoding RAD51, a protein essential for DNA repair by homologous recombination (HR), to characterize the mechanism underlying IPMK-regulated mRNA export. IPMK depletion or catalytic inactivation selectively decreases the nuclear export of RAD51 mRNA, and RAD51 protein abundance, thereby impairing HR. Recognition of a sequence motif in the untranslated region of RAD51 transcripts by the mRNA export factor ALY requires IPMK. Phosphatidylinositol (3,4,5)-trisphosphate (PIP3), an IPMK product, restores ALY recognition in IPMK-depleted cell extracts, suggesting a mechanism underlying transcript selection. Our findings implicate IPMK in a transcript-selective mRNA export pathway controlled by phosphoinositide turnover that preserves genome integrity in humans. We used gene expression profiling to compare the abundance of cytoplasmic RNAs after IPMK depletion
2013-09-27 | E-GEOD-49658 | biostudies-arrayexpress