CRISPR knockout screen implicates three genes in lysosome function
Ontology highlight
ABSTRACT:
ORGANISM(S): synthetic construct Homo sapiens
PROVIDER: GSE129282 | GEO | 2019/05/01
REPOSITORIES: GEO
Similar Datasets
Project description:Cholesterol and phosphoinositides (PI) are two critically important lipids that are found in cellular membranes and dysregulated in many disorders. Therefore, uncovering molecular pathways connecting these essential lipids may offer new therapeutic insights. We report that loss of function of lysosomal Niemann-Pick Type C1 (NPC1) cholesterol transporter, which leads to neurodegenerative NPC disease, initiates a signaling cascade that alters the cholesterol/phosphatidylinositol 4-phosphate (PtdIns4P) countertransport cycle between Golgi-endoplasmic reticulum (ER), as well as lysosome-ER membrane contact sites (MCS). Central to these disruptions is increased recruitment of phosphatidylinositol 4-kinases-PI4KIIα and PI4KIIIβ-which boosts PtdIns4P metabolism at Golgi and lysosomal membranes. Aberrantly increased PtdIns4P levels elevate constitutive anterograde secretion from the Golgi complex, and mTORC1 recruitment to lysosomes. NPC1 disease mutations phenocopy the transporter loss of function and can be rescued by inhibition or knockdown of either key phosphoinositide enzymes or their recruiting partners. In summary, we show that the lysosomal NPC1 cholesterol transporter tunes the molecular content of Golgi and lysosome MCS to regulate intracellular trafficking and growth signaling in health and disease.
2021-09-15 | MTBLS2612 | MetaboLights
Project description:Phosphatase activities on PI(3,4,5)P3 and
PI(3,4)P2
This model describes the action of various phosphatases on
PI(3,4,5)P3 and PI(3,4)P2, in response to a stimulation by EGF. It
contains boolean switches to simulate knock-down and knock-out of
phosphatases as well as inhibition of PI3 kinase.
This model is described in the article:
PTEN Regulates PI(3,4)P2
Signaling Downstream of Class I PI3K
Mouhannad Malek, Anna Kielkowska,
Tamara Chessa, Karen E. Anderson, David Barneda, P?nar Pir,
Hiroki Nakanishi, Satoshi Eguchi, Atsushi Koizumi, Junko Sasaki,
Véronique Juvin, Vladimir Y. Kiselev, Izabella Niewczas,
Alexander Gray, Alexandre Valayer, Dominik Spensberger, Marine
Imbert, Sergio Felisbino, Tomonori Habuchi, Soren Beinke, Sabina
Cosulich, Nicolas Le Novère, Takehiko Sasaki, Jonathan
Clark, Phillip T. Hawkins and Len R. Stephens
Molecular Cell
Abstract:
The PI3K signaling pathway regulates cell growth and
movement and is heavily mutated in cancer. Class I PI3Ks
synthesize the lipid messenger PI(3,4,5)P3. PI(3,4,5)P3 can be
dephosphorylated by 3- or 5-phosphatases, the latter producing
PI(3,4)P2. The PTEN tumor suppressor is thought to function
primarily as a PI(3,4,5)P3 3-phosphatase, limiting activation
of this pathway. Here we show that PTEN also functions as a
PI(3,4)P2 3-phosphatase, both in vitro and in vivo. PTEN is a
major PI(3,4)P2 phosphatase in Mcf10a cytosol, and loss of PTEN
and INPP4B, a known PI(3,4)P2 4-phosphatase, leads to
synergistic accumulation of PI(3,4)P2, which correlated with
increased invadopodia in epidermal growth factor
(EGF)-stimulated cells. PTEN deletion increased PI(3,4)P2
levels in a mouse model of prostate cancer, and it inversely
correlated with PI(3,4)P2 levels across several EGF-stimulated
prostate and breast cancer lines. These results point to a role
for PI(3,4)P2 in the phenotype caused by loss-of-function
mutations or deletions in PTEN.
This model is hosted on
BioModels Database
and identified by:
MODEL1704190000.
To cite BioModels Database, please use:
Chelliah V et al. BioModels: ten-year
anniversary. Nucl. Acids Res. 2015, 43(Database
issue):D542-8.
To the extent possible under law, all copyright and related or
neighbouring rights to this encoded model have been dedicated to
the public domain worldwide. Please refer to
CC0
Public Domain Dedication for more information.
| MODEL1704190000 | BioModels
Project description:Emerging evidences suggest that both function and position of organelles are pivotal for tumor cell dissemination. Among them, lysosomes stand out as they integrate metabolic sensing with gene regulation and secretion of proteases. Yet, how lysosomes function is linked to their position and thereby control metastatic progression remains elusive. Here, we analyzed lysosome subcellular distribution in micropatterned patient-derived melanoma cells and found that lysosome spreading scales with their aggressiveness. Peripheral lysosomes promote invadopodia-based matrix degradation and invasion of melanoma cells which is directly linked to their lysosomal and cell transcriptional programs. When controlling lysosomal positioning using chemo-genetical heterodimerization in melanoma cells, we demonstrated that perinuclear clustering impairs lysosomal secretion, matrix degradation and invasion. Impairing lysosomal spreading in a zebrafish metastasis model significantly reduces invasive outgrowth. Our study provides a mechanistic demonstration that lysosomal positioning controls cell invasion, illustrating the importance of organelle adaptation in carcinogenesis.
2023-07-31 | E-MTAB-13165 | biostudies-arrayexpress
Project description:Transcription factors can affect autophagy activity by promoting or inhibiting the expression of autophagy and lysosomal related genes. As a zinc finger family DNA-binding protein, ZKSCAN3 has been reported to function as a transcriptional repressor of autophagy, silencing of which can induced autophagy and promoted lysosome biogenesis in cancer cells. However, the studies in Zkscan 3 knockout mice showed that the deficiency of Zkscan3 did not induce autophagy and in-crease lysosome biogenesis. In order to further explore the role of ZKSCAN3 in the transcriptional regulation of autophagy in human cells, we generated ZKSCAN3 knockout HK-2 and Hela cells by CRISPR/Cas9 system and analyzed the differences in gene expression between ZKSCAN3 deleted cells and non-deleted cells through fluorescence quantitative PCR, Western blot and transcriptome sequencing, with special attention to the differences in gene expression of autophagy and lysosomal biogenesis. We found that ZKSCAN3 is not an essential regulator of autophagic or lysosomal gene expression, as the absence ZKSCAN3 had no significantly effect on the expression of autophagy or lysosomal genes.
2023-01-31 | GSE218727 | GEO
Project description:Aging is characterized by the loss of homeostasis and the general decline of physiological functions, accompanied by various degenerative diseases and higher rates of mortality. Anti-aging targeted small molecule screens have been carried outperformed many times, however, few have focused on endogenous metabolic intermediates—metabolites. Here, using C. elegans lifespan assays, we conducted a worm metabolite screen and identified an conserved metabolite through eEukaryotes-conserved metabolite, myo-inositol (MI), to extend lifespan, increase mobility and reduce fat content. Genetic analysis of enzymes in MI metabolic pathway suggestsed that MI alleviates aging through its derivative PI(4,5)P2. MI and PI(4,5)P2 are precursors of PI(3,4,5)P3, which is negatively related to longevity. However, the mechanism study shows itslongevity effects could be fully blocked by a null-allele mutation of the tumor suppressor gene, daf-18 (homologous to mouse Pten), areand was independent of its classical pathway downstream elements, akt or daf-16., but Instead, we found MI effects on aging were dependent on its downstream mitophagy regulator pink-1. MI’s anti-aging effect is was also conserved in mouse model, implicatingindicating a conserved mechanism in mammals that may extend to human.
2020-08-03 | GSE154417 | GEO
Project description:Ammonia is a ubiquitous, toxic by-product of cell metabolism. Its high membrane permeability and proton affinity causes ammonia to accumulate inside acidic lysosomes in its poorly membrane-permeant form: ammonium (NH4+). Ammonium buildup compromises lysosomal function, suggesting the existence of mechanisms that protect cells from ammonium toxicity. Here, we identified SLC12A9 as a lysosomal regulator of ammonium export that preserves lysosomal homeostasis. SLC12A9 knockout cells showed grossly enlarged lysosomes and elevated ammonium content. These phenotypes were reversed upon removal of the metabolic source of ammonium or dissipation of the lysosomal pH gradient. Lysosomal chloride increased in SLC12A9 knockout cells and chloride binding by SLC12A9 was required for ammonium transport. Our data indicate that SLC12A9 function is central for the handling of lysosomal ammonium and chloride, an unappreciated, fundamental mechanism of lysosomal physiology that may have special relevance in tissues with elevated ammonia, such as tumors.
2024-09-12 | GSE276655 | GEO
Project description:Understanding how cells mitigate lysosomal damage is critical for unraveling pathogenic mechanisms of lysosome related diseases. Here we use organelle-specific proteomics in iPSC derived neurons (i3Neuron) and an in vitro lysosome damaging assay to demonstrate that lysosome damage, caused by the aggregation of Ceroid Lipofuscinosis Neuronal 4 (CLN4) linked DNAJC5 mutants on lysosomal membranes, serves as a critical pathogenic linchpin in CLN4 associated neurodegeneration. Intriguingly, in non neuronal cells, a ubiquitin dependent microautophagy mechanism downregulates CLN4 aggregates to counteract CLN4 associated lysotoxicity. Genome wide CRISPR screens identify the ubiquitin ligase CHIP as a central microautophagy regulator that confers ubiquitin dependent lysosome protection. Importantly, CHIP lysosome protection function is transferrable, as ectopic CHIP improves lysosomal function in CLN4 i3Neurons, and effectively alleviates lipofuscin accumulation and neurodegeneration in a Drosophila CLN4 disease model. Our study establishes CHIP mediated microautophagy as a key organelle damage guardian that preserves lysosome integrity, offering new insights into therapeutic development for CLN4 and other lysosome related neurodegenerative diseases.
2024-12-11 | MSV000096646 | MassIVE
Project description:Lysosome plays important roles in cellular homeostasis and its dysregulation is associated with cancer. However, the regulation and underlying mechanism of lysosome in cancer survival remain incompletely understood. Here, we reveal a role for a histone acetylation-regulated long non-coding RNA termed as lysosome cell death regulator (LCDR) in lung cancer cell survival, knockdown of which promotes apoptosis. Mechanistically, LCDR binds to heterogenous nuclear ribonucleoprotein K (hnRNP K) to regulate the stability of lysosomal-associated protein transmembrane 5 (LAPTM5) transcript that maintain integrity of lysosomal membrane. Consequently, knockdown of LCDR, hnRNP K or LAPTM5 promotes lysosomal membrane permeabilization and lysosomal cell death-mediated apoptosis. LAPTM5 overexpression or cathepsin B inhibitor partially restores the effects of this axis on lysosomal cell death. Clinically, LCDR/hnRNP K/LAPTM5 are upregulated in lung cancer tissues and coexpression of this axis shows the increased diagnostic value for lung cancer. These findings shed light on LCDR/hnRNP K/LAPTM5 as potential therapeutic targets and targeting lysosome is a promising strategy in cancer treatment.
2021-05-05 | GSE173849 | GEO
Project description:We screened proteins in close proximity to PI(4,5)P2 in epithelial cells. We established HaCaT cells stably expressing APEX2 N-terminally fused to the PH domain of PLCd1, which specifically binds to PI(4,5)P2. In the presence of hydrogen peroxide and biotin-phenol, proteins proximal to APEX2 are biotinylated by APEX2, allowing for their enrichment using streptavidin beads. APEX2-fused non-PI(4,5) P2-binding mutant of the PH domain of PLCd1 (R40L) was used as the negative control.
2022-03-26 | PXD031266 | Pride
Project description:Lysosomal failure underlies pathogenesis of numerous congenital neurodegenerative disorders and is an early and progressive feature of Alzheimer’s disease (AD) pathogenesis. Here, we report that lysosomal dysfunction in Down Syndrome (Trisomy 21) requires the extra gene copy of amyloid precursor protein (APP) and is mediated by the beta cleaved carboxy terminal fragment of APP (APP-βCTF, C99). In primary fibroblasts from individuals with Down Syndrome (DS), lysosomal degradation of autophagic and endocytic substrates is selectively impaired causing them to accumulate in enlarged autolysosomes/lysosomes. Direct measurements of lysosomal pH uncovered a significant elevation (0.6 units) associated with slowed LC3 turnover and the inactivation of cathepsin D (CTSD) and other lysosomal hydrolases known to be unstable or less active when lysosomal pH is persistently elevated. RNA sequencing analysis excluded a transcriptional contribution to hydrolase declines. Normalizing lysosome pH by delivering acidic nanoparticles to lysosomes ameliorated lysosomal deficits, implicating pH elevation as their primary basis. Cortical neurons cultured from the Ts2 mouse model of DS exhibited lysosomal deficits similar to those in DS cells. Lowering APP expression with siRNA or BACE1 inhibition reversed cathepsin deficits in both fibroblasts and neurons. Deleting one BACE1 allele from adult Ts2 mice had similar rescue effects in vivo. The modest elevation of endogenous APP-βCTF needed to disrupt lysosomal function in DS is relevant to sporadic AD where APP-βCTF, but not APP, is also elevated. Our results extend evidence that impaired lysosomal acidification drives progressive lysosomal failure in multiple forms of AD.
2019-05-30 | GSE127880 | GEO