scholarly journals Regulation of lysosome biogenesis by phosphoinositides and phagocytosis

2021 ◽  
Author(s):  
Christopher Choy
Keyword(s):  
Cell Function ◽  
De Novo ◽  
Mammalian Target Of Rapamycin ◽  
Protein Translation ◽  
Nuclear Accumulation ◽  
Fcγ Receptor ◽  
Lysosome Biogenesis ◽  
Transcription Factor Eb ◽  
Acidic Organelles ◽  
Pathogen Clearance

Lysosomes are acidic organelles responsible for molecular degradation, energy balance, and pathogen clearance. Consequently, lysosome dysfunction is linked to numerous diseases, including lysosome storage diseases. Notably, enhancing lysosome biogenesis ameliorates cell function and helps clear metabolites. The transcription factor EB (TFEB) is a master regulator of lysosome biogenesis, and thus a potential therapeutic target. Among known regulators of TFEB, the mammalian target of rapamycin complex 1 (mTORC1) is best understood. In nutrient-rich cells, mTORC1 is activated and represses TFEB by phosphorylation. Upon starvation, mTORC1 is inactivated and TFEB enters the nucleus, upregulating lysosomal gene expression to enhance cellular degradation for energy recovery. Numerous other TFEB-dependent pathways have been identified. We aim to understand how TFEB is regulated in two additional contexts: in lysosome enlargement during phosphatidylinositol 3,5-bisphosphate [PtdIns(3,5)P2] depletion and in phagocytosis. First, PtdIns(3,5)P2 is required for maintaining lysosome size by an incompletely understood mechanism. We hypothesized that TFEB-mediated lysosome biogenesis contributes de novo lysosomal material. Acute depletion of PtdIns(3,5)P2-synthesizing kinase PIKfyve induced TFEB nuclear accumulation. Despite increases in transcription, little to no protein translation was observed. Furthermore, tfeb-/-cells and cells blocked with cycloheximide were similar to wild-type cells, with regard to the number and size of lysosomes during PIKfyve inhibition cells, suggesting biosynthesis is not necessary for lysosome enlargement. However, TFEB still becomes active by an known mechanism. We show that TFEB nuclear localization during PIKfyve inhibition was not due to mTORC1 inactivation but may result from GSK3 inhibition. Secondly, phagocytosis allows immune cells to sequester potential pathogens by engulfing them into phagosomes. These phagosomes are then degraded by the lysosome. We postulated that phagocytosis would enhance TFEB-mediated lysosome biogenesis to promote pathogen killing. Fcγ receptor-mediated phagocytosis activated TFEB and increased biosynthesis of select lysosomal genes, augmenting existing lysosomes and enhancing proteolysis. To understand how TFEB was activated by the Fcγ receptor, we inhibited key signaling and trafficking mediators. Particle internalization, phagosome formation, and phagosome maturation appear to be necessary for TFEB activation. Overall, our work uncovers two additional mechanisms that may govern TFEBactivation.

scholarly journals Regulation of lysosome biogenesis by phosphoinositides and phagocytosis

2021 ◽  
Author(s):  
Christopher Choy
Keyword(s):  
Cell Function ◽  
De Novo ◽  
Mammalian Target Of Rapamycin ◽  
Protein Translation ◽  
Nuclear Accumulation ◽  
Fcγ Receptor ◽  
Lysosome Biogenesis ◽  
Transcription Factor Eb ◽  
Acidic Organelles ◽  
Pathogen Clearance

Lysosomes are acidic organelles responsible for molecular degradation, energy balance, and pathogen clearance. Consequently, lysosome dysfunction is linked to numerous diseases, including lysosome storage diseases. Notably, enhancing lysosome biogenesis ameliorates cell function and helps clear metabolites. The transcription factor EB (TFEB) is a master regulator of lysosome biogenesis, and thus a potential therapeutic target. Among known regulators of TFEB, the mammalian target of rapamycin complex 1 (mTORC1) is best understood. In nutrient-rich cells, mTORC1 is activated and represses TFEB by phosphorylation. Upon starvation, mTORC1 is inactivated and TFEB enters the nucleus, upregulating lysosomal gene expression to enhance cellular degradation for energy recovery. Numerous other TFEB-dependent pathways have been identified. We aim to understand how TFEB is regulated in two additional contexts: in lysosome enlargement during phosphatidylinositol 3,5-bisphosphate [PtdIns(3,5)P2] depletion and in phagocytosis. First, PtdIns(3,5)P2 is required for maintaining lysosome size by an incompletely understood mechanism. We hypothesized that TFEB-mediated lysosome biogenesis contributes de novo lysosomal material. Acute depletion of PtdIns(3,5)P2-synthesizing kinase PIKfyve induced TFEB nuclear accumulation. Despite increases in transcription, little to no protein translation was observed. Furthermore, tfeb-/-cells and cells blocked with cycloheximide were similar to wild-type cells, with regard to the number and size of lysosomes during PIKfyve inhibition cells, suggesting biosynthesis is not necessary for lysosome enlargement. However, TFEB still becomes active by an known mechanism. We show that TFEB nuclear localization during PIKfyve inhibition was not due to mTORC1 inactivation but may result from GSK3 inhibition. Secondly, phagocytosis allows immune cells to sequester potential pathogens by engulfing them into phagosomes. These phagosomes are then degraded by the lysosome. We postulated that phagocytosis would enhance TFEB-mediated lysosome biogenesis to promote pathogen killing. Fcγ receptor-mediated phagocytosis activated TFEB and increased biosynthesis of select lysosomal genes, augmenting existing lysosomes and enhancing proteolysis. To understand how TFEB was activated by the Fcγ receptor, we inhibited key signaling and trafficking mediators. Particle internalization, phagosome formation, and phagosome maturation appear to be necessary for TFEB activation. Overall, our work uncovers two additional mechanisms that may govern TFEBactivation.

scholarly journals Oxidative Stress in the Male Germline: A Review of Novel Strategies to Reduce 4-Hydroxynonenal Production

Antioxidants ◽  
2018 ◽  
Vol 7 (10) ◽  
pp. 132 ◽  
Author(s):  
Jessica Walters ◽  
Geoffry De Iuliis ◽  
Brett Nixon ◽  
Elizabeth Bromfield
Keyword(s):  
Oxidative Stress ◽  
Germ Cells ◽  
Cell Function ◽  
De Novo ◽  
Protein Translation ◽  
Egg Recognition ◽  
Male Germline ◽  
New Strategy ◽  
Artery Disease ◽  
The Stability

Germline oxidative stress is intimately linked to several reproductive pathologies including a failure of sperm-egg recognition. The lipid aldehyde 4-hydroxynonenal (4HNE) is particularly damaging to the process of sperm-egg recognition as it compromises the function and the stability of several germline proteins. Considering mature spermatozoa do not have the capacity for de novo protein translation, 4HNE modification of proteins in the mature gametes has uniquely severe consequences for protein homeostasis, cell function and cell survival. In somatic cells, 4HNE overproduction has been attributed to the action of lipoxygenase enzymes that facilitate the oxygenation and degradation of ω-6 polyunsaturated fatty acids (PUFAs). Accordingly, the arachidonate 15-lipoxygenase (ALOX15) enzyme has been intrinsically linked with 4HNE production, and resultant pathophysiology in various complex conditions such as coronary artery disease and multiple sclerosis. While ALOX15 has not been well characterized in germ cells, we postulate that ALOX15 inhibition may pose a new strategy to prevent 4HNE-induced protein modifications in the male germline. In this light, this review focuses on (i) 4HNE-induced protein damage in the male germline and its implications for fertility; and (ii) new methods for the prevention of lipid peroxidation in germ cells.

scholarly journals TFEB Overexpression, Not mTOR Inhibition, Ameliorates RagCS75Y Cardiomyopathy

2021 ◽  
Vol 22 (11) ◽  
pp. 5494
Author(s):  
Maengjo Kim ◽  
Linghui Lu ◽  
Alexey V. Dvornikov ◽  
Xiao Ma ◽  
Yonghe Ding ◽  
...  
Keyword(s):  
Nuclear Translocation ◽  
De Novo ◽  
Mammalian Target Of Rapamycin ◽  
Missense Variant ◽  
Neonatal Rat ◽  
Physical Interaction ◽  
Mtor Inhibition ◽  
Transcription Factor Eb ◽  
Rag Gtpase ◽  
Rat Ventricle

A de novo missense variant in Rag GTPase protein C (RagCS75Y) was recently identified in a syndromic dilated cardiomyopathy (DCM) patient. However, its pathogenicity and the related therapeutic strategy remain unclear. We generated a zebrafish RragcS56Y (corresponding to human RagCS75Y) knock-in (KI) line via TALEN technology. The KI fish manifested cardiomyopathy-like phenotypes and poor survival. Overexpression of RagCS75Y via adenovirus infection also led to increased cell size and fetal gene reprogramming in neonatal rat ventricle cardiomyocytes (NRVCMs), indicating a conserved mechanism. Further characterization identified aberrant mammalian target of rapamycin complex 1 (mTORC1) and transcription factor EB (TFEB) signaling, as well as metabolic abnormalities including dysregulated autophagy. However, mTOR inhibition failed to ameliorate cardiac phenotypes in the RagCS75Y cardiomyopathy models, concomitant with a failure to promote TFEB nuclear translocation. This observation was at least partially explained by increased and mTOR-independent physical interaction between RagCS75Y and TFEB in the cytosol. Importantly, TFEB overexpression resulted in more nuclear TFEB and rescued cardiomyopathy phenotypes. These findings suggest that S75Y is a pathogenic gain-of-function mutation in RagC that leads to cardiomyopathy. A primary pathological step of RagCS75Y cardiomyopathy is defective mTOR–TFEB signaling, which can be corrected by TFEB overexpression, but not mTOR inhibition.

Regulation of the SREBP transcription factors by mTORC1

2011 ◽  
Vol 39 (2) ◽  
pp. 495-499 ◽  
Author(s):  
Caroline A. Lewis ◽  
Beatrice Griffiths ◽  
Claudio R. Santos ◽  
Mario Pende ◽  
Almut Schulze
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Synthase ◽  
Target Genes ◽  
De Novo ◽  
Regulatory Element ◽  
Cholesterol Biosynthesis ◽  
Mammalian Target Of Rapamycin ◽  
Lipid Biosynthesis ◽  
De Novo Lipogenesis ◽  
Genes Encoding

In recent years several reports have linked mTORC1 (mammalian target of rapamycin complex 1) to lipogenesis via the SREBPs (sterol-regulatory-element-binding proteins). SREBPs regulate the expression of genes encoding enzymes required for fatty acid and cholesterol biosynthesis. Lipid metabolism is perturbed in some diseases and SREBP target genes, such as FASN (fatty acid synthase), have been shown to be up-regulated in some cancers. We have previously shown that mTORC1 plays a role in SREBP activation and Akt/PKB (protein kinase B)-dependent de novo lipogenesis. Our findings suggest that mTORC1 plays a crucial role in the activation of SREBP and that the activation of lipid biosynthesis through the induction of SREBP could be part of a regulatory pathway that co-ordinates protein and lipid biosynthesis during cell growth. In the present paper, we discuss the increasing amount of data supporting the potential mechanisms of mTORC1-dependent activation of SREBP as well as the implications of this signalling pathway in cancer.

Antileukemic activity of rapamycin in acute myeloid leukemia

Blood ◽  
2005 ◽  
Vol 105 (6) ◽  
pp. 2527-2534 ◽  
Author(s):  
Christian Récher ◽  
Odile Beyne-Rauzy ◽  
Cécile Demur ◽  
Gaëtan Chicanne ◽  
Cédric Dos Santos ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
De Novo ◽  
Mammalian Target Of Rapamycin ◽  
Cell Types ◽  
Mtor Inhibition ◽  
Growth And Survival ◽  
Clinical Responses ◽  
De Novo Aml ◽  
Acute Myeloid

AbstractThe mammalian target of rapamycin (mTOR) is a key regulator of growth and survival in many cell types. Its constitutive activation has been involved in the pathogenesis of various cancers. In this study, we show that mTOR inhibition by rapamycin strongly inhibits the growth of the most immature acute myeloid leukemia (AML) cell lines through blockade in G0/G1 phase of the cell cycle. Accordingly, 2 downstream effectors of mTOR, 4E-BP1 and p70S6K, are phosphorylated in a rapamycin-sensitive manner in a series of 23 AML cases. Interestingly, the mTOR inhibitor markedly impairs the clonogenic properties of fresh AML cells while sparing normal hematopoietic progenitors. Moreover, rapamycin induces significant clinical responses in 4 of 9 patients with either refractory/relapsed de novo AML or secondary AML. Overall, our data strongly suggest that mTOR is aberrantly regulated in most AML cells and that rapamycin and analogs, by targeting the clonogenic compartment of the leukemic clone, may be used as new compounds in AML therapy.

scholarly journals Furin controls β cell function via mTORC1 signaling

2020 ◽  
Author(s):  
Bas Brouwers ◽  
Ilaria Coppola ◽  
Katlijn Vints ◽  
Bastian Dislich ◽  
Nathalie Jouvet ◽  
...  
Keyword(s):  
Secretory Pathway ◽  
Cell Function ◽  
Mammalian Target Of Rapamycin ◽  
Human Islets ◽  
Β Cells ◽  
Β Cell ◽  
Differential Proteomics ◽  
Atpase Subunit ◽  
Proteolytic Activation ◽  
Mtorc1 Signaling

AbstractFurin is a proprotein convertase (PC) responsible for proteolytic activation of a wide array of precursor proteins within the secretory pathway. It maps to the PRC1 locus, a type 2 diabetes susceptibility locus, yet its specific role in pancreatic β cells is largely unknown. The aim of this study was to determine the role of furin in glucose homeostasis. We show that furin is highly expressed in human islets, while PCs that potentially could provide redundancy are expressed at considerably lower levels. β cell-specific furin knockout (βfurKO) mice are glucose intolerant, due to smaller islets with lower insulin content and abnormal dense core secretory granule morphology. RNA expression analysis and differential proteomics on βfurKO islets revealed activation of Activating Transcription Factor 4 (ATF4), which was mediated by mammalian target of rapamycin C1 (mTORC1). βfurKO cells show impaired cleavage of the essential V-ATPase subunit Ac45, and by blocking this pump in β cells the mTORC1 pathway is activated. Furthermore, βfurKO cells show lack of insulin receptor cleavage and impaired response to insulin. Taken together, these results suggest a model of mTORC1-ATF4 hyperactivation in β cells lacking furin, which causes β cell dysfunction.

scholarly journals DNAM-1 regulates Foxp3 expression in regulatory T cells by interfering with TIGIT under inflammatory conditions

2021 ◽  
Vol 118 (21) ◽  
pp. e2021309118
Author(s):  
Kazuki Sato ◽  
Yumi Yamashita-Kanemaru ◽  
Fumie Abe ◽  
Rikito Murata ◽  
Yuho Nakamura-Shinya ◽  
...  
Keyword(s):  
Treg Cell ◽  
Intracellular Signaling ◽  
Cell Function ◽  
Inflammatory Diseases ◽  
Mammalian Target Of Rapamycin ◽  
Foxp3 Expression ◽  
Treg Cells ◽  
Cell Dysfunction ◽  
Inflammatory Conditions ◽  
Treg Cell Function

Regulatory T (Treg) cells that express forkhead box P3 (Foxp3) are pivotal for immune tolerance. Although inflammatory mediators cause Foxp3 instability and Treg cell dysfunction, their regulatory mechanisms remain incompletely understood. Here, we show that the transfer of Treg cells deficient in the activating immunoreceptor DNAM-1 ameliorated the development of graft-versus-host disease better than did wild-type Treg cells. We found that DNAM-1 competes with T cell immunoreceptor with Ig and ITIM domains (TIGIT) in binding to their common ligand CD155 and therefore regulates TIGIT signaling to down-regulate Treg cell function without DNAM-1–mediated intracellular signaling. DNAM-1 deficiency augments TIGIT signaling; this subsequently inhibits activation of the protein kinase B–mammalian target of rapamycin complex 1 pathway, resulting in the maintenance of Foxp3 expression and Treg cell function under inflammatory conditions. These findings demonstrate that DNAM-1 regulates Treg cell function via TIGIT signaling and thus, it is a potential molecular target for augmenting Treg function in inflammatory diseases.

scholarly journals Imaging the fate of histone Cse4 reveals de novo replacement in S phase and subsequent stable residence at centromeres

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Jan Wisniewski ◽  
Bassam Hajj ◽  
Jiji Chen ◽  
Gaku Mizuguchi ◽  
Hua Xiao ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Elevated Temperatures ◽  
De Novo ◽  
Histone H3 ◽  
S Phase ◽  
Nuclear Accumulation ◽  
Histone H3 Variant ◽  
Functionally Impaired ◽  
Cell Cycle Dynamics ◽  
Cell Lethality

The budding yeast centromere contains Cse4, a specialized histone H3 variant. Fluorescence pulse-chase analysis of an internally tagged Cse4 reveals that it is replaced with newly synthesized molecules in S phase, remaining stably associated with centromeres thereafter. In contrast, C-terminally-tagged Cse4 is functionally impaired, showing slow cell growth, cell lethality at elevated temperatures, and extra-centromeric nuclear accumulation. Recent studies using such strains gave conflicting findings regarding the centromeric abundance and cell cycle dynamics of Cse4. Our findings indicate that internally tagged Cse4 is a better reporter of the biology of this histone variant. Furthermore, the size of centromeric Cse4 clusters was precisely mapped with a new 3D-PALM method, revealing substantial compaction during anaphase. Cse4-specific chaperone Scm3 displays steady-state, stoichiometric co-localization with Cse4 at centromeres throughout the cell cycle, while undergoing exchange with a nuclear pool. These findings suggest that a stable Cse4 nucleosome is maintained by dynamic chaperone-in-residence Scm3.

scholarly journals Multi-regional characterisation of renal cell carcinoma and microenvironment at single cell resolution

2021 ◽  
Author(s):  
Ruoyan Li ◽  
John R. Ferdinand ◽  
Kevin W. Loudon ◽  
Georgina S. Bowyer ◽  
Lira Mamanova ◽  
...  
Keyword(s):  
Single Cell ◽  
Cell Function ◽  
De Novo ◽  
Epithelial Mesenchymal Transition ◽  
Spatial Location ◽  
De Novo Mutation ◽  
Cellular Interactions ◽  
Sequencing Data ◽  
Mesenchymal Transition ◽  
Multiple Regions

Tumour behaviour is dependent on the oncogenic properties of cancer cells and their multi-cellular interactions. These dependencies were examined through 270,000 single cell transcriptomes and 100 micro-dissected whole exomes obtained from 12 patients with kidney tumours. Tissue was sampled from multiple regions of tumour core, tumour-normal interface, normal surrounding tissues, and peripheral blood. We found the principal spatial location of CD8+ T cell clonotypes largely defined exhaustion state, with clonotypic heterogeneity not explained by somatic intra-tumoural heterogeneity. De novo mutation calling from single cell RNA sequencing data allows us to lineage-trace and infer clonality of cells. We discovered six meta-programmes that distinguish tumour cell function. An epithelial-mesenchymal transition meta-programme, enriched at the tumour-normal interface appears modulated through macrophage expressed IL1B, potentially forming a therapeutic target.

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