Tetrandrine is a novel inhibitor of NAADP-induced calcium release through targeting Lysosomal Integral Membrane Protein 2 (LIMP-2)
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ABSTRACT:
PROVIDER: PRJNA876411 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR21412069_1.fastq.gz | Fastqsanger.gz | |||
| SRR21412069_2.fastq.gz | Fastqsanger.gz | |||
| SRR21412070_1.fastq.gz | Fastqsanger.gz | |||
| SRR21412070_2.fastq.gz | Fastqsanger.gz | |||
| SRR21412071_1.fastq.gz | Fastqsanger.gz |
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Project description:Tetrandrine (Tet), a bis-benzylisoquinoline alkaloid isolated from Stephania tetrandra, has been implicated in a broad range of pharmacological activities, and is best known for its highly potent activity against Ebola virus infection by inhibiting NAADP-mediated calcium release via the endosomal/lysosomal two-pore channels (TPCs). It has been proposed that Tet inhibits the activity of TPCs directly, despite a lack of direct evidence. Using a synthetic, alkynyl diazirine derivative of Tet (AD-Tet), a clickable photoaffinity probe as a tool for target identification, we present results from pulldown SILAC-MS, transcriptomics, biochemical and cell analysis to show that Tet/AD-Tet specifically targets lysosomal integral membrane protein-2 (LIMP-2), a lysosomal cholesterol transporter. Accordingly, we delineated the principles underlying the autophagic and cytotoxic properties of Tet, and discovered that hypercholesteremia is a previously unknown side-effect of Tet in vivo. More importantly, we discover that LIMP-2 is an essential regulator of NAADP-mediated calcium release, and implies that LIMP-2 is a novel target for the development of anti-Ebola virus therapeutics.
2023-03-04 | GSE212648 | GEO
Project description:The intercalated disc of cardiac myocytes is emerging as a crucial structure in the heart. Loss of intercalated disc proteins like N-cadherin causes lethal cardiac abnormalities, mutations in intercalated disc proteins cause human cardiomyopathy. A comprehensive screen for novel mechanisms in failing hearts demonstrated that expression of the lysosomal integral membrane protein-2 (LIMP-2) is increased in cardiac hypertrophy and heart failure in both rat and human myocardium. Complete loss of LIMP-2 in genetically engineered mice did not affect cardiac development; however these LIMP-2 null mice failed to mount a hypertrophic response to increased blood pressure but developed cardiomyopathy. Disturbed cadherin localization in these hearts suggested that LIMP-2 has important functions outside lysosomes. Indeed, we also find LIMP-2 in the intercalated disc, where it associates with cadherin. RNAi-mediated knockdown of LIMP-2 decreases the binding of phosphorylated b-catenin to cadherin, while overexpression of LIMP-2 has the opposite effect. Taken together, our data show that lysosomal integrated membrane protein-2 is crucial to mount the adaptive hypertrophic response to cardiac loading. We demonstrate a novel role for LIMP-2 as an important mediator of the intercalated disc. Experiment Overall Design: overall design: Experiment Overall Design: 3 groups of rats, 1 sample per rat: Experiment Overall Design: - compensated = Ren2 rat, hypertensive, no heart failure (N=6) Experiment Overall Design: - failure = Ren2 rat, hypertensive, no heart failure (N=4) Experiment Overall Design: - SD = control group, non-hypertensive (N=4)
2008-06-13 | E-GEOD-4286 | biostudies-arrayexpress
Project description:The intercalated disc of cardiac myocytes is emerging as a crucial structure in the heart. Loss of intercalated disc proteins like N-cadherin causes lethal cardiac abnormalities, mutations in intercalated disc proteins cause human cardiomyopathy. A comprehensive screen for novel mechanisms in failing hearts demonstrated that expression of the lysosomal integral membrane protein-2 (LIMP-2) is increased in cardiac hypertrophy and heart failure in both rat and human myocardium. Complete loss of LIMP-2 in genetically engineered mice did not affect cardiac development; however these LIMP-2 null mice failed to mount a hypertrophic response to increased blood pressure but developed cardiomyopathy. Disturbed cadherin localization in these hearts suggested that LIMP-2 has important functions outside lysosomes. Indeed, we also find LIMP-2 in the intercalated disc, where it associates with cadherin. RNAi-mediated knockdown of LIMP-2 decreases the binding of phosphorylated b-catenin to cadherin, while overexpression of LIMP-2 has the opposite effect. Taken together, our data show that lysosomal integrated membrane protein-2 is crucial to mount the adaptive hypertrophic response to cardiac loading. We demonstrate a novel role for LIMP-2 as an important mediator of the intercalated disc. Keywords: heart failure, comparison
2007-10-05 | GSE4286 | GEO
Project description:We report the application of RNA-seq for molecular profiling of cultured, cortical astrocytes. Our data set is based on about 40 million unique reads per sample in four independent mRNA preparations. Cortical astrocytes are a prototypical cell model for investigating calcium signaling. For analysis of calcium fluxes, we performed direct calcium imaging in the endoplasmic reticulum and in the cytosol. We describe in our study the physiological profile of homeostatic and agonist-induced calcium fluxes. Furthermore, we show how ER calcium release shapes the cytosolic calcium signal. This transcriptome dataset was used to profile the calcium toolkit of astrocytes. The data suggest that a small number of calcium signaling-related proteins mediate calcium homeostasis in astrocytes.
2021-12-14 | GSE166464 | GEO
Project description:Homeostatic calcium fluxes, ER calcium release, SOCE, and calcium oscillations in cultured astrocytes are interlinked by a small calcium toolkit
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Project description:Calcium signals are initiated in immune cells by the process of store-operated calcium entry (SOCE), where receptor activation triggers transient calcium release from the endoplasmic reticulum, followed by opening of plasma membrane calcium-release activated calcium (CRAC) channels. ORAI1, ORAI2 and ORAI3 are known to comprise the CRAC channel, however the contributions of individual isoforms to neutrophil function is not well understood. Here we show that loss of ORAI1 partially decreases calcium influx while loss of both ORAI1 and ORAI2 completely abolishes store-operated calcium entry. In other immune cell types, loss of ORAI2 enhances SOCE. In contrast, we find that ORAI2-deficient neutrophils display decreased calcium influx, which is correlated with measurable differences in regulation of neutrophil membrane potential via KCa3.1. Decreased SOCE in ORAI1-, ORAI2- and ORAI1/2-deficient neutrophils impairs multiple neutrophil functions including phagocytosis, degranulation, leukotriene and ROS production, rendering ORAI1/2-deficient mice highly susceptible to staphylococcal infection. This study demonstrates that ORAI1 and ORAI2 are the primary components of the neutrophil CRAC channel and identifies novel subpopulations of neutrophils where cell membrane potential functions as a rheostat to modulate the SOCE response. These findings have implications for new mechanisms that modulate neutrophil function during infection, acute and chronic inflammatory conditions, and cancer.
2020-09-01 | GSE157200 | GEO
Project description:To investigate the putative differential effect of chloride transport on lysosomal ion homeostasis,we generated an ODE mathematical model for this system. Our mathematical model builds upon a previously published model for lysosomal homeostasis (Grabe et al. 2001 J. Gen. Physiol, Ishida et al. 2013 J. Gen. Physiol), and further includes the (de)activation kinetics of the ClC-7 antiporter and Ca2+ uptake/release mechanisms. Our new model tracks the total number of ions within the lysosomal lumen over time. It considers different types of lysosomal ion channels and exchangers and two possible lysosomal Ca2+ transporters. The variation in the total number of each ion within the lysosome is described by an ordinary differential equation (ODE), and the rate of change is determined by the flux of the corresponding ion across the lysosomal membrane. The model considers different elements affecting lysosomal ion homeostasis, among which are: (i) the V-ATPase pump, (ii) a proton leak, (iii) the luminal proton buffering capacity, (iv) ClC-7 chloride/proton exchanger, (v) Ca2+ /proton exchanger (CAX), (vi) passive channels for K+ , Na+ , and Ca2+ , and (vii) Donnan particles, which are negatively charged particles or molecules trapped in the lysosomal lumen. It allows for the simulation of different scenarios mimicking the differential transport of chloride, its impact on Ca2+ uptake and release and ultimately on lysosomal homeostasis. We provide a Supplementary Information file (Astaburuaga et al. Cells 2019) with the full description of the mathematical model. The equations written in purple were newly developed, the other elements were retrieved from a previous mathematical model as indicated above.
2019-10-22 | MODEL1910220001 | BioModels
Project description:Lysosomal cathepsins regulate an exquisite range of biological functions, and their deregulation is associated with inflammatory, metabolic and degenerative disease in humans. Here, we identified a key cell-intrinsic role for cathepsin B as a negative feedback regulator of lysosomal biogenesis and autophagy. Mice and macrophages lacking cathepsin B activity had increased resistance to the cytosolic bacterial pathogen Francisella novicida. Genetic deletion or pharmacological inhibition of cathepsin B downregulated mTOR activity and prevented cleavage of the lysosomal calcium channel TRPML1. These events drove transcription of lysosomal and autophagy genes via the transcription factor TFEB, which increased lysosomal biogenesis and activation of autophagy-initiation kinase ULK1 for clearance of the bacteria. Our results identified a fundamental biological function of cathepsin B in providing a checkpoint for homeostatic maintenance of lysosome population and basic recycling functions in the cell. We used microarrays to explore the gene expression profiles differentially expressed in bone marrow-derived macrophages (BMDM) isolated from cathepsin B-/- and wild-type mice.
2016-08-04 | E-GEOD-79508 | biostudies-arrayexpress
Project description:Autophagic and endosomal dysfunctions are prominently observed at preclinical stages of Alzheimer's disease (AD) as well as in presenilin (PSEN) 1-deficient mice and neurons. In the latter, the defects relate to the γ-secretase-independent role of PSEN in lysosomal fusion and organelle turnover. While we demonstrated previously that the impaired capacity of lysosomal fusion is associated with a significant reduction in lysosomal calcium storage/release, the underlying mechanism remained unexplored. Here we demonstrate that PSEN-deficient cells are impaired in endosomal recycling of several cargo proteins reminiscent of clathrin-independent carriers and lipid rafts. This is accompanied by the accumulation of cholesterol in LAMP1-positive organelles. The small GTPase ARF6, an important regulator of lipid raft recycling, fully rescues the endo-lysosomal abnormalities in PSEN-/- cells, suggesting that defective recycling is upstream of lysosomal dysfunction. PSEN-/- cells and neurons present significantly reduced ARF6 expression levels. Importantly, similar decreased ARF6 levels are observed in aging murine neurons and brain and are even more pronounced in AD brains, suggesting a particular vulnerability of ARF6-mediated recycling in the early etiology of AD.
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