bims-lysosi 2022-09-11 papers

bims-lysosi

Biomed News

on Lysosomes and signaling

Issue of 2022–09–11
thirty-two papers selected by
Stephanie Fernandes, Max Planck Institute for Biology of Ageing

A phosphoinositide signalling pathway mediates rapid lysosomal repair.
Lysosomal enzyme trafficking factor LYSET enables nutritional usage of extracellular proteins.
The human disease gene LYSET is essential for lysosomal enzyme transport and viral infection.
Activation of mitochondrial TRAP1 stimulates mitochondria-lysosome crosstalk and correction of lysosomal dysfunction.
ATP6V0C variants impair vacuolar V-ATPase causing a neurodevelopmental disorder often associated with epilepsy.
Inhibition of the Niemann-Pick C1 protein is a conserved feature of multiple strains of pathogenic mycobacteria.
Increased glucose metabolism in TAMs fuels O-GlcNAcylation of lysosomal Cathepsin B to promote cancer metastasis and chemoresistance.
Bidirectional lysosome transport: a balancing act between ARL8 effectors.
Lysosome Inhibition Reduces Basal and Nutrient-Induced Fat Accumulation in Caenorhabditis elegans.
Lysosomal Pathogenesis of Parkinson's Disease: Insights From LRRK2 and GBA1 Rodent Models.
The crosstalk between MYC and mTORC1 during osteoclastogenesis.
The lysosomal proteome of senescent cells contributes to the senescence secretome.
Hyperactivation of mTOR and AKT in a cardiac hypertrophy animal model of Friedreich ataxia.
Role of Mechanistic Target of Rapamycin in Autophagy and Alcohol-Associated Liver Disease.
Imiquimod-induced ROS production causes lysosomal membrane permeabilization and activates caspase-8-mediated apoptosis in skin cancer cells.
Olmsted syndrome causing point mutants of TRPV3 (G568C and G568D) show defects in intracellular Ca2+-mobilization and induce lysosomal defects.
mTORC2 Is the Major Second Layer Kinase Negatively Regulating FOXO3 Activity.
Alpha-Synuclein: The Spark That Flames Dopaminergic Neurons, In Vitro and In Vivo Evidence.
Pharmacologic inhibition of LAT1 predominantly suppresses transport of large neutral amino acids and downregulates global translation in cancer cells.
LC-MS/MS-based enzyme assay for lysosomal acid lipase using dried blood spots.
GBA1 and The Immune System: A Potential Role in Parkinson's Disease?
Deubiquitinase OTUD5 modulates mTORC1 signaling to promote bladder cancer progression.
An alliance between lipid transfer proteins and scramblases for membrane expansion.
Delivery and assessment of a CRISPR/nCas9-based genome editing system on in vitro models of mucopolysaccharidoses IVA assisted by magnetite-based nanoparticles.
Exploring the link between GBA1 mutations and Dementia with Lewy bodies, A Mini-Review.
Publisher Correction: Long non-coding RNA SNHG6 couples cholesterol sensing with mTORC1 activation in hepatocellular carcinoma.
The TFEB-TGIF1 axis regulates EMT in mouse epicardial cells.
Vacuolar H+ -ATPase subunit VAB3 regulates cell growth and ion homeostasis in Arabidopsis.
Tyrosine phosphatase activity is restricted by basic charge substituting mutation of substrates.
Targeting PI3K/AKT/mTOR Signaling Pathway in Pancreatic Cancer: From Molecular to Clinical Aspects.
Germline homozygous missense DEPDC5 variants cause severe refractory early-onset epilepsy, macrocephaly and bilateral polymicrogyria.
Long-term treatment with low dose rapamycin extends lifespan and delays tumours in cancer-prone germline PTEN mutant mice.

Nature. 2022 Sep 07.

A phosphoinositide signalling pathway mediates rapid lysosomal repair.

Jay Xiaojun Tan, Toren Finkel.

Lysosomal dysfunction has been increasingly linked to disease and normal ageing1,2. Lysosomal membrane permeabilization (LMP), a hallmark of lysosome-related diseases, can be triggered by diverse cellular stressors3. Given the damaging contents of lysosomes, LMP must be rapidly resolved, although the underlying mechanisms are poorly understood. Here, using an unbiased proteomic approach, we show that LMP stimulates a phosphoinositide-initiated membrane tethering and lipid transport (PITT) pathway for rapid lysosomal repair. Upon LMP, phosphatidylinositol-4 kinase type 2α (PI4K2A) accumulates rapidly on damaged lysosomes, generating high levels of the lipid messenger phosphatidylinositol-4-phosphate. Lysosomal phosphatidylinositol-4-phosphate in turn recruits multiple oxysterol-binding protein (OSBP)-related protein (ORP) family members, including ORP9, ORP10, ORP11 and OSBP, to orchestrate extensive new membrane contact sites between damaged lysosomes and the endoplasmic reticulum. The ORPs subsequently catalyse robust endoplasmic reticulum-to-lysosome transfer of phosphatidylserine and cholesterol to support rapid lysosomal repair. Finally, the lipid transfer protein ATG2 is also recruited to damaged lysosomes where its activity is potently stimulated by phosphatidylserine. Independent of macroautophagy, ATG2 mediates rapid membrane repair through direct lysosomal lipid transfer. Together, our findings identify that the PITT pathway maintains lysosomal membrane integrity, with important implications for numerous age-related diseases characterized by impaired lysosomal function.

DOI: https://doi.org/10.1038/s41586-022-05164-4
Science. 2022 Sep 08. eabn5637

Lysosomal enzyme trafficking factor LYSET enables nutritional usage of extracellular proteins.

Catarina Pechincha, Sven Groessl, Robert Kalis, Melanie de Almeida, Andrea Zanotti, Marten Wittmann, Martin Schneider, Rafael P de Campos, Sarah Rieser, Marlene Brandstetter, Alexander Schleiffer, Karin Müller-Decker, Dominic Helm, Sabrina Jabs, David Haselbach, Marius K Lemberg, Johannes Zuber, Wilhelm Palm.

Mammalian cells can generate amino acids through macropinocytosis and lysosomal breakdown of extracellular proteins, which is exploited by cancer cells to grow in nutrient-poor tumors. Here, through genetic screens in defined nutrient conditions we characterized LYSET, a transmembrane protein (TMEM251) selectively required when cells consume extracellular proteins. LYSET was found to associate in the Golgi with GlcNAc-1-phosphotransferase, which targets catabolic enzymes to lysosomes through mannose-6-phosphate modification. Without LYSET, GlcNAc-1-phosphotransferase was unstable owing to a hydrophilic transmembrane domain. Consequently, LYSET-deficient cells were depleted of lysosomal enzymes and impaired in turnover of macropinocytic and autophagic cargoes. Thus, LYSET represents a core component of the lysosomal enzyme trafficking pathway, underlies the pathomechanism for hereditary lysosomal storage disorders, and may represent a target to suppress metabolic adaptations in cancer.

DOI: https://doi.org/10.1126/science.abn5637
Science. 2022 Sep 08. eabn5648

The human disease gene LYSET is essential for lysosomal enzyme transport and viral infection.

Christopher M Richards, Sabrina Jabs, Wenjie Qiao, Lauren D Varanese, Michaela Schweizer, Peter R Mosen, Nicholas M Riley, Malte Klüssendorf, James R Zengel, Ryan A Flynn, Arjun Rustagi, John C Widen, Christine E Peters, Yaw Shin Ooi, Xuping Xie, Pei-Yong Shi, Ralf Bartenschlager, Andreas S Puschnik, Matthew Bogyo, Carolyn R Bertozzi, Catherine A Blish, Dominic Winter, Claude M Nagamine, Thomas Braulke, Jan E Carette.

Lysosomes are key degradative compartments of the cell. Transport to lysosomes relies on GlcNAc-1-phosphotransferase-mediated tagging of soluble enzymes with mannose 6-phosphate (M6P). GlcNAc-1-phosphotransferase deficiency leads to the severe lysosomal storage disorder mucolipidosis II (MLII). Several viruses require lysosomal cathepsins to cleave structural proteins and thus depend on functional GlcNAc-1-phosphotransferase. Here, we used genome-scale CRISPR screens to identify Lysosomal Enzyme Trafficking factor (LYSET) as essential for infection by cathepsin-dependent viruses including SARS-CoV-2. LYSET deficiency resulted in global loss of M6P tagging and mislocalization of GlcNAc-1-phosphotransferase from the Golgi complex to lysosomes. Lyset knockout mice exhibited MLII-like phenotypes and human pathogenic LYSET alleles failed to restore lysosomal sorting defects. Thus, LYSET is required for correct functioning of the M6P trafficking machinery, and mutations in LYSET can explain the phenotype of the associated disorder.

DOI: https://doi.org/10.1126/science.abn5648
iScience. 2022 Sep 16. 25(9): 104941

Activation of mitochondrial TRAP1 stimulates mitochondria-lysosome crosstalk and correction of lysosomal dysfunction.

Fannie W Chen, Joanna P Davies, Raul Calvo, Jagruti Chaudhari, Georgia Dolios, Mercedes K Taylor, Samarjit Patnaik, Jean Dehdashti, Rebecca Mull, Patricia Dranchack, Amy Wang, Xin Xu, Emma Hughes, Noel Southall, Marc Ferrer, Rong Wang, Juan J Marugan, Yiannis A Ioannou.

  Numerous studies have established the involvement of lysosomal and mitochondrial dysfunction in the pathogenesis of neurodegenerative disorders such as Alzheimer's and Parkinson diseases. Building on our previous studies of the neurodegenerative lysosomal lipidosis Niemann-Pick C1 (NPC1), we have unexpectedly discovered that activation of the mitochondrial chaperone tumor necrosis factor receptor-associated protein 1 (TRAP1) leads to the correction of the lysosomal storage phenotype in patient cells from multiple lysosomal storage disorders including NPC1. Using small compound activators specific for TRAP1, we find that activation of this chaperone leads to a generalized restoration of lysosomal and mitochondrial health. Mechanistically, we show that this process includes inhibition of oxidative phosphorylation and reduction of oxidative stress, which results in activation of AMPK and ultimately stimulates lysosome recycling. Thus, TRAP1 participates in lysosomal-mitochondrial crosstalk to maintain cellular homeostasis and could represent a potential therapeutic target for multiple disorders.

Keywords:  Cell biology; Cellular neuroscience; Neuroscience

DOI:  https://doi.org/10.1016/j.isci.2022.104941
Brain. 2022 Sep 08. pii: awac330. [Epub ahead of print]

ATP6V0C variants impair vacuolar V-ATPase causing a neurodevelopmental disorder often associated with epilepsy.

Kari A Mattison, Gilles Tossing, Fred Mulroe, Callum Simmons, Kameryn M Butler, Alison Schreiber, Adnan Alsadah, Derek E Neilson, Karin Naess, Anna Wedell, Anna Wredenberg, Arthur Sorlin, Emma McCann, George J Burghel, Beatriz Menendez, George E Hoganson, Lorenzo D Botto, Francis M Filloux, Ángel Aledo-Serrano, Antonio Gil-Nagel, Katrina Tatton-Brown, Nienke E Verbeek, Michele van Hirtum-Das, Jeroen Breckpot, Trine Bjørg Hammer, Rikke S Møller, Andrea Whitney, Andrew G L Douglas, Mira Kharbanda, Nicola Brunetti-Pierri, Manuela Morleo, Vincenzo Nigro, Halie J May, James X Tao, Emanuela Argili, Elliot H Sherr, William B Dobyns, Genomics England Research Consortium, Richard A Baines, Jim Warwicker, J Alex Parker, Siddharth Banka, Philippe M Campeau, Andrew Escayg.

  The vacuolar H+-ATPase (V-ATPase) is an enzymatic complex that functions in an ATP-dependent manner to pump protons across membranes and acidify organelles, thereby creating the proton/pH gradient required for membrane trafficking by several different types of transporters. We describe heterozygous point variants in ATP6V0C, encoding the c-subunit in the membrane bound integral domain of the V-ATPase, in 27 patients with neurodevelopmental abnormalities with or without epilepsy. Corpus callosum hypoplasia and cardiac abnormalities were also present in some patients. In silico modeling suggested that the patient variants interfere with the interactions between the ATP6V0C and ATP6V0A subunits during ATP hydrolysis. Consistent with decreased V-ATPase activity, functional analyses conducted in Saccharomyces cerevisiae revealed reduced LysoSensor fluorescence and reduced growth in media containing varying concentrations of CaCl2. Knockdown of ATP6V0C in Drosophila resulted in increased duration of seizure-like behavior, and the expression of selected patient variants in Caenorhabditis elegans led to reduced growth, motor dysfunction, and reduced lifespan. In summary, this study establishes ATP6V0C as an important disease gene, describes the clinical features of the associated neurodevelopmental disorder, and provides insight into disease mechanisms.

Keywords:  ATP6V0C; V-ATPase; VMA3; epilepsy genetics; neurodevelopmental disorders

DOI:  https://doi.org/10.1093/brain/awac330
Nat Commun. 2022 Sep 09. 13(1): 5320

Inhibition of the Niemann-Pick C1 protein is a conserved feature of multiple strains of pathogenic mycobacteria.

Yuzhe Weng, Dawn Shepherd, Yi Liu, Nitya Krishnan, Brian D Robertson, Nick Platt, Gerald Larrouy-Maumus, Frances M Platt.

Mycobacterium tuberculosis (Mtb) survives and replicates within host macrophages (MΦ) and subverts multiple antimicrobial defense mechanisms. Previously, we reported that lipids shed by pathogenic mycobacteria inhibit NPC1, the lysosomal membrane protein deficient in the lysosomal storage disorder Niemann-Pick disease type C (NPC). Inhibition of NPC1 leads to a drop in lysosomal calcium levels, blocking phagosome-lysosome fusion leading to mycobacterial survival. We speculated that the production of specific cell wall lipid(s) that inhibit NPC1 could have been a critical step in the evolution of pathogenicity. We therefore investigated whether lipid extracts from clinical Mtb strains from multiple Mtb lineages, Mtb complex (MTBC) members and non-tubercular mycobacteria (NTM) inhibit the NPC pathway. We report that inhibition of the NPC pathway was present in all clinical isolates from Mtb lineages 1, 2, 3 and 4, Mycobacterium bovis and the NTM, Mycobacterium abscessus and Mycobacterium avium. However, lipid extract from Mycobacterium canettii, which is considered to resemble the common ancestor of the MTBC did not inhibit the NPC1 pathway. We conclude that the evolution of NPC1 inhibitory mycobacterial cell wall lipids evolved early and post divergence from Mycobacterium canettii-related mycobacteria and that this activity contributes significantly to the promotion of disease.

DOI: https://doi.org/10.1038/s41467-022-32553-0
Cancer Cell. 2022 Aug 30. pii: S1535-6108(22)00376-2. [Epub ahead of print]

Increased glucose metabolism in TAMs fuels O-GlcNAcylation of lysosomal Cathepsin B to promote cancer metastasis and chemoresistance.

Qingzhu Shi, Qicong Shen, Yanfang Liu, Yang Shi, Wenwen Huang, Xi Wang, Zhiqing Li, Yangyang Chai, Hao Wang, Xiangjia Hu, Nan Li, Qian Zhang, Xuetao Cao.

  How glucose metabolism remodels pro-tumor functions of tumor-associated macrophages (TAMs) needs further investigation. Here we show that M2-like TAMs bear the highest individual capacity to take up intratumoral glucose. Their increased glucose uptake fuels hexosamine biosynthetic pathway-dependent O-GlcNAcylation to promote cancer metastasis and chemoresistance. Glucose metabolism promotes O-GlcNAcylation of the lysosome-encapsulated protease Cathepsin B at serine 210, mediated by lysosome-localized O-GlcNAc transferase (OGT), elevating mature Cathepsin B in macrophages and its secretion in the tumor microenvironment (TME). Loss of OGT in macrophages reduces O-GlcNAcylation and mature Cathepsin B in the TME and disrupts cancer metastasis and chemoresistance. Human TAMs with high OGT are positively correlated with Cathepsin B expression, and both levels predict chemotherapy response and prognosis of individuals with cancer. Our study reports the biological and potential clinical significance of glucose metabolism in tumor-promoting TAMs and reveals insights into the underlying mechanisms.

Keywords:  O-GlcNAc transferase; O-GlcNAcylation; cathepsin B; glucose metabolism; lysosome; metastasis; tumor-associated macrophages

DOI:  https://doi.org/10.1016/j.ccell.2022.08.012
Nat Commun. 2022 Sep 07. 13(1): 5261

Bidirectional lysosome transport: a balancing act between ARL8 effectors.

Agnieszka A Kendrick, Jenna R Christensen.

DOI: https://doi.org/10.1038/s41467-022-32965-y
Mol Cells. 2022 Sep 30. 45(9): 649-659

Lysosome Inhibition Reduces Basal and Nutrient-Induced Fat Accumulation in Caenorhabditis elegans.

Rui Lu, Juan Chen, Fangbin Wang, Lu Wang, Jian Liu, Yan Lin.

  A long-term energy nutritional imbalance fundamentally causes the development of obesity and associated fat accumulation. Lysosomes, as nutrient-sensing and lipophagy centers, critically control cellular lipid catabolism in response to nutrient deprivation. However, whether lysosome activity is directly involved in nutrient-induced fat accumulation remains unclear. In this study, worm fat accumulation was induced by 1 mM glucose or 0.02 mM palmitic acid supplementation. Along with the elevation of fat accumulation, lysosomal number and acidification were also increased, suggesting that lysosome activity might be correlated with nutrient-induced fat deposition in Caenorhabditis elegans. Furthermore, treatments with the lysosomal inhibitors chloroquine and leupeptin significantly reduced basal and nutrient-induced fat accumulation in C. elegans. The knockdown of hlh-30, which is a critical gene in lysosomal biogenesis, also resulted in worm fat loss. Finally, the mutation of aak-2, daf-15, and rsks-1 showed that mTORC1 (mechanistic target of rapamycin complex-1) signaling mediated the effects of lysosomes on basal and nutrient-induced fat accumulation in C. elegans. Overall, this study reveals the previously undescribed role of lysosomes in overnutrition sensing, suggesting a new strategy for controlling body fat accumulation.

Keywords:  Caenorhabditis elegans; fat accumulation; lysosome; nutrient

DOI:  https://doi.org/10.14348/molcells.2022.0073
Neurotherapeutics. 2022 Sep 09.

Lysosomal Pathogenesis of Parkinson's Disease: Insights From LRRK2 and GBA1 Rodent Models.

Mattia Volta.

  The discovery of mutations in LRRK2 and GBA1 that are linked to Parkinson's disease provided further evidence that autophagy and lysosome pathways are likely implicated in the pathogenic process. Their protein products are important regulators of lysosome function. LRRK2 has kinase-dependent effects on lysosome activity, autophagic efficacy and lysosomal Ca2+ signaling. Glucocerebrosidase (encoded by GBA1) is a hydrolytic enzyme contained in the lysosomes and contributes to the degradation of alpha-synuclein. PD-related mutations in LRRK2 and GBA1 slow the degradation of alpha-synuclein, thus directly implicating the dysfunction of the process in the neuropathology of Parkinson's disease. The development of genetic rodent models of LRRK2 and GBA1 provided hopes of obtaining reliable preclinical models in which to study pathogenic processes and perform drug validation studies. Here, I will review the extensive characterization of these models, their impact on understanding lysosome alterations in the course of Parkinson's disease and what novel insights have been obtained. In addition, I will discuss how these models fare with respect to the features of a "gold standard" animal models and what could be attempted in future studies to exploit LRRK2 and GBA1 rodent models in the fight against Parkinson's disease.

Keywords:  Animal models; Glucocerebrosidase; LRRK2; Lysosomes; Neuropathology; Parkinson’s disease

DOI:  https://doi.org/10.1007/s13311-022-01290-z
Front Cell Dev Biol. 2022 ;10 920683

The crosstalk between MYC and mTORC1 during osteoclastogenesis.

Seyeon Bae, Brian Oh, Jefferson Tsai, Peter Sang Uk Park, Matthew Blake Greenblatt, Eugenia G Giannopoulou, Kyung-Hyun Park-Min.

  Osteoclasts are bone-resorbing cells that undergo extensive changes in morphology throughout their differentiation. Altered osteoclast differentiation and activity lead to changes in pathological bone resorption. The mammalian target of rapamycin (mTOR) is a kinase, and aberrant mTOR complex 1 (mTORC1) signaling is associated with altered bone homeostasis. The activation of mTORC1 is biphasically regulated during osteoclastogenesis; however, the mechanism behind mTORC1-mediated regulation of osteoclastogenesis and bone resorption is incompletely understood. Here, we found that MYC coordinates the dynamic regulation of mTORC1 activation during osteoclastogenesis. MYC-deficiency blocked the early activation of mTORC1 and also reversed the decreased activity of mTORC1 at the late stage of osteoclastogenesis. The suppression of mTORC1 activity by rapamycin in mature osteoclasts enhances bone resorption activity despite the indispensable role of high mTORC1 activation in osteoclast formation in both mouse and human cells. Mechanistically, MYC induces Growth arrest and DNA damage-inducible protein (GADD34) expression and suppresses mTORC1 activity at the late phase of osteoclastogenesis. Taken together, our findings identify a MYC-GADD34 axis as an upstream regulator of dynamic mTORC1 activation in osteoclastogenesis and highlight the interplay between MYC and mTORC1 pathways in determining osteoclast activity.

Keywords:  GADD34 (PPP1R15A); MYC (c-myc); bone resorption; mTORC1 (mechanistic target of rapamycin complex 1); osteoclast (OC)

DOI:  https://doi.org/10.3389/fcell.2022.920683
Aging Cell. 2022 Sep 10. e13707

The lysosomal proteome of senescent cells contributes to the senescence secretome.

Miguel Rovira, Rebecca Sereda, David Pladevall-Morera, Valentina Ramponi, Ines Marin, Mate Maus, Julio Madrigal-Matute, Antonio Díaz, Fernando García, Javier Muñoz, Ana María Cuervo, Manuel Serrano.

  Senescent cells accumulate in tissues over time, favoring the onset and progression of multiple age-related diseases. Senescent cells present a remarkable increase in lysosomal mass and elevated autophagic activity. Here, we report that two main autophagic pathways macroautophagy (MA) and chaperone-mediated autophagy (CMA) are constitutively upregulated in senescent cells. Proteomic analyses of the subpopulations of lysosomes preferentially engaged in each of these types of autophagy revealed profound quantitative and qualitative changes in senescent cells, affecting both lysosomal resident proteins and cargo proteins delivered to lysosomes for degradation. These studies have led us to identify resident lysosomal proteins that are highly augmented in senescent cells and can be used as novel markers of senescence, such as arylsulfatase ARSA. The abundant secretome of senescent cells, known as SASP, is considered their main pathological mediator; however, little is known about the mechanisms of SASP secretion. Some secretory cells, including melanocytes, use the small GTPase RAB27A to perform lysosomal secretion. We found that this process is exacerbated in the case of senescent melanoma cells, as revealed by the exposure of lysosomal membrane integral proteins LAMP1 and LAMP2 in their plasma membrane. Interestingly, a subset of SASP components, including cytokines CCL2, CCL3, CXCL12, cathepsin CTSD, or the protease inhibitor SERPINE1, are secreted in a RAB27A-dependent manner in senescent melanoma cells. Finally, proteins previously identified as plasma biomarkers of aging are highly enriched in the lysosomes of senescent cells, including CTSD. We conclude that the lysosomal proteome of senescent cells is profoundly reconfigured, and that some senescent cells can be highly active in lysosomal exocytosis.

Keywords:  SASP; aging; autophagy; cellular senescence; exocytosis; lysosome

DOI:  https://doi.org/10.1111/acel.13707
Heliyon. 2022 Aug;8(8): e10371

Hyperactivation of mTOR and AKT in a cardiac hypertrophy animal model of Friedreich ataxia.

Wing-Hang Tong, Hayden Ollivierre, Audrey Noguchi, Manik C Ghosh, Danielle A Springer, Tracey A Rouault.

  Cardiomyopathy is a primary cause of death in Friedreich ataxia (FRDA) patients with defective iron-sulfur cluster (ISC) biogenesis due to loss of functional frataxin and in rare patients with functional loss of other ISC biogenesis factors. The mechanistic target of rapamycin (mTOR) and AKT signaling cascades that coordinate eukaryotic cell growth and metabolism with environmental inputs, including nutrients and growth factors, are crucial regulators of cardiovascular growth and homeostasis. We observed increased phosphorylation of AKT and dysregulation of multiple downstream effectors of mTORC1, including S6K1, S6, ULK1 and 4EBP1, in a cardiac/skeletal muscle specific FRDA conditional knockout (cKO) mouse model and in human cell lines depleted of ISC biogenesis factors. Knockdown of several mitochondrial metabolic proteins that are downstream targets of ISC biogenesis, including lipoyl synthase and subunit B of succinate dehydrogenase, also resulted in activation of mTOR and AKT signaling, suggesting that mTOR and AKT hyperactivations are part of the metabolic stress response to ISC deficiencies. Administration of rapamycin, a specific inhibitor of mTOR signaling, enhanced the survival of the Fxn cKO mice, providing proof of concept for the potential of mTOR inhibition to ameliorate cardiac disease in patients with defective ISC biogenesis. However, AKT phosphorylation remained high in rapamycin-treated Fxn cKO hearts, suggesting that parallel mTOR and AKT inhibition might be necessary to further improve the lifespan and healthspan of ISC deficient individuals.

Keywords:  AKT; Cardiac hypertrophy; FXN; Frataxin; Friedreich ataxia; ISCU; Iron-sulfur cluster biogenesis; Metabolic stress; mTOR

DOI:  https://doi.org/10.1016/j.heliyon.2022.e10371
Am J Physiol Cell Physiol. 2022 Sep 05.

Role of Mechanistic Target of Rapamycin in Autophagy and Alcohol-Associated Liver Disease.

Xiaojuan Chao, Sha Neisha Williams, Wen-Xing Ding.

  Mechanistic target of rapamycin (mTOR) is a serine-threonine kinase and a cellular sensor for nutrient and energy status, which is critical in regulating cell metabolism and growth by governing the anabolic (protein and lipid synthesis) and catabolic process (autophagy). Alcohol-associated liver disease (ALD) is a major chronic liver disease worldwide that carries a huge financial burden. The spectrum of the pathogenesis of ALD includes steatosis, fibrosis, inflammation, ductular reaction and eventual hepatocellular carcinoma, which is closely associated with metabolic changes that are regulated by mTOR. In this review, we summarized recent progress of alcohol consumption on the changes of mTORC1 and mTORC2 activity, the potential mechanisms and possible impact of the mTORC1 changes on autophagy in ALD. We also discussed the potential beneficial effects and limitations of targeting mTORC1 against ALD.

Keywords:  ALD; Autophagy; MTOR; TFEB; ethanol

DOI:  https://doi.org/10.1152/ajpcell.00281.2022
J Dermatol Sci. 2022 Aug 28. pii: S0923-1811(22)00197-9. [Epub ahead of print]

Imiquimod-induced ROS production causes lysosomal membrane permeabilization and activates caspase-8-mediated apoptosis in skin cancer cells.

Shu-Hao Chang, Pei-Ying Lin, Tsai-Kun Wu, Chien-Sheng Hsu, Shi-Wei Huang, Zheng-Yi Li, Kuang-Ting Liu, Jun-Kai Kao, Yi-Ju Chen, Tak-Wah Wong, Chun-Ying Wu, Jeng-Jer Shieh.

   BACKGROUND: Lysosomal cell death is induced by lysosomal membrane permeabilization (LMP) and the subsequent release of lysosomal proteolytic enzymes, including cathepsins (CTSs), which results in mitochondrial dysfunction and apoptosis. Imiquimod (IMQ), a synthetic TLR7 ligand, has both antiviral and antitumor activity against various skin malignancies in clinical treatment. Previously, we demonstrated IMQ not only caused lysosomal dysfunction but also triggered lysosome biogenesis to achieve lysosomal adaptation in cancer cells.
OBJECTIVE: To determine whether lysosomes are involved in IMQ-induced apoptosis.
METHODS: The human skin cancer cell lines BCC, A375 and mouse melanoma cell line B16F10 were used in all experiments. Cell death was determined by the Cell Counting Kit-8 (CCK-8) assay and DNA content assay. Protein expression was determined by immunoblotting. Caspase-8 activity was assessed using a fluorescence caspase-8 kit and determined by flow cytometry and confocal microscopy.
RESULTS: IMQ not only induced lysosome damage but also abrogated lysosome function in skin cancer cells. IMQ-induced caspase-8 activation contributed to the processes of lysosomal cell death. Moreover, the use of ROS scavengers significantly abolished caspase-8 activation and inhibited IMQ-induced LMP. Additionally, pharmacological inhibition of CTSD not only abrogated caspase-8 activation but also rescued IMQ-induced cell death. Finally, lysosome-alkalizing agents enhanced the cytotoxicity of IMQ in vitro and in vivo.
CONCLUSIONS: IMQ-induced ROS accumulation promotes LMP, releases CTSs into the cytosol, stimulates caspase-8 activation and finally causes lysosomal cell death. Lysosomal cell death and the CTSD/caspase-8 axis may play a crucial role in IMQ-induced cell death.

Keywords:  Imiquimod; Lysosomal cell death; ROS

DOI:  https://doi.org/10.1016/j.jdermsci.2022.08.006
Biochem Biophys Res Commun. 2022 Aug 28. pii: S0006-291X(22)01139-1. [Epub ahead of print]628 32-39

Olmsted syndrome causing point mutants of TRPV3 (G568C and G568D) show defects in intracellular Ca2+-mobilization and induce lysosomal defects.

Anushka Jain, Ram Prasad Sahu, Chandan Goswami.

  TRPV3, a non-selective cation channel known to be activated by physiological temperature, is expressed in skin and is involved in different skin functions. Point mutations in TRPV3 cause severe pathological condition, known as Olmsted Syndrome (OS). Now we demonstrate that two OS-inducing point mutations (G568C and G568D) located at the lipid-water-interface region joining TM4 with the loop4 of TRPV3 cause reduced cell size and major defects in lysosomal numbers, and distribution. We detected these two mutants in the lysosome. However, G568C and G568D mutants differ from themselves and also from Wild-type in terms of Ca2+-influx in response to activation by agonist (FPP). These two mutants fail to mobilise Ca2+ from intracellular stores, especially when cytosolic Ca2+ is chelated and/or in absence of extracellular Ca2+. We demonstrate that OS-mutants cause defective pH-maintenance at the lysosomes. We propose that G568C and G568D mutants most-likely act as Ca2+-leaky channels from lysosomes with different abilities.

Keywords:  Ca(2+)-channel; Channelopathy; Lipid-water-interface; Lysosomal pH; Skin disorder; Thermosensitive ion channel

DOI:  https://doi.org/10.1016/j.bbrc.2022.08.026
Molecules. 2022 Aug 24. pii: 5414. [Epub ahead of print]27(17):

mTORC2 Is the Major Second Layer Kinase Negatively Regulating FOXO3 Activity.

Lucia Jimenez, Carlos Amenabar, Victor Mayoral-Varo, Thomas A Mackenzie, Maria C Ramos, Andreia Silva, Giampaolo Calissi, Inês Grenho, Carmen Blanco-Aparicio, Joaquin Pastor, Diego Megías, Bibiana I Ferreira, Wolfgang Link.

  Forkhead box O (FOXO) proteins are transcription factors involved in cancer and aging and their pharmacological manipulation could be beneficial for the treatment of cancer and healthy aging. FOXO proteins are mainly regulated by post-translational modifications including phosphorylation, acetylation and ubiquitination. As these modifications are reversible, activation and inactivation of FOXO factors is attainable through pharmacological treatment. One major regulatory input of FOXO signaling is mediated by protein kinases. Here, we use specific inhibitors against different kinases including PI3K, mTOR, MEK and ALK, and other receptor tyrosine kinases (RTKs) to determine their effect on FOXO3 activity. While we show that inhibition of PI3K efficiently drives FOXO3 into the cell nucleus, the dual PI3K/mTOR inhibitors dactolisib and PI-103 induce nuclear FOXO translocation more potently than the PI3Kδ inhibitor idelalisib. Furthermore, specific inhibition of mTOR kinase activity affecting both mTORC1 and mTORC2 potently induced nuclear translocation of FOXO3, while rapamycin, which specifically inhibits the mTORC1, failed to affect FOXO3. Interestingly, inhibition of the MAPK pathway had no effect on the localization of FOXO3 and upstream RTK inhibition only weakly induced nuclear FOXO3. We also measured the effect of the test compounds on the phosphorylation status of AKT, FOXO3 and ERK, on FOXO-dependent transcriptional activity and on the subcellular localization of other FOXO isoforms. We conclude that mTORC2 is the most important second layer kinase negatively regulating FOXO activity.

Keywords:  FOXO; aging; cancer; chemical biology; high content screening; kinases; mTOR

DOI:  https://doi.org/10.3390/molecules27175414
Int J Mol Sci. 2022 Aug 30. pii: 9864. [Epub ahead of print]23(17):

Alpha-Synuclein: The Spark That Flames Dopaminergic Neurons, In Vitro and In Vivo Evidence.

Alexandre Henriques, Laura Rouvière, Elodie Giorla, Clémence Farrugia, Bilal El Waly, Philippe Poindron, Noëlle Callizot.

  Mitochondria, α-syn fibrils and the endo-lysosomal system are key players in the pathophysiology of Parkinson's disease. The toxicity of α-syn is amplified by cell-to-cell transmission and aggregation of endogenous species in newly invaded neurons. Toxicity of α-syn PFF was investigated using primary cultures of dopaminergic neurons or on aged mice after infusion in the SNpc and combined with mild inhibition of GBA. In primary dopaminergic neurons, application of α-syn PFF induced a progressive cytotoxicity associated with mitochondrial dysfunction, oxidative stress, and accumulation of lysosomes suggesting that exogenous α-syn reached the lysosome (from the endosome). Counteracting the α-syn endocytosis with a clathrin inhibitor, dopaminergic neuron degeneration was prevented. In vivo, α-syn PFF induced progressive neurodegeneration of dopaminergic neurons associated with motor deficits. Histology revealed progressive aggregation of α-syn and microglial activation and accounted for the seeding role of α-syn, injection of which acted as a spark suggesting a triggering of cell-to-cell toxicity. We showed for the first time that a localized SNpc α-syn administration combined with a slight lysosomal deficiency and aging triggered a progressive lesion. The cellular and animal models described could help in the understanding of the human disease and might contribute to the development of new therapies.

Keywords:  GBA; Parkinson’s disease; mitochondrial dysfunction; spreading; α-synuclein

DOI:  https://doi.org/10.3390/ijms23179864
J Cell Mol Med. 2022 Sep 07.

Pharmacologic inhibition of LAT1 predominantly suppresses transport of large neutral amino acids and downregulates global translation in cancer cells.

Kou Nishikubo, Ryuichi Ohgaki, Hiroki Okanishi, Suguru Okuda, Minhui Xu, Hitoshi Endou, Yoshikatsu Kanai.

  L-type amino acid transporter 1 (LAT1; SLC7A5), which preferentially transports large neutral amino acids, is highly upregulated in various cancers. LAT1 supplies cancer cells with amino acids as substrates for enhanced biosynthetic and bioenergetic reactions and stimulates signalling networks involved in the regulation of survival, growth and proliferation. LAT1 inhibitors show anti-cancer effects and a representative compound, JPH203, is under clinical evaluation. However, pharmacological impacts of LAT1 inhibition on the cellular amino acid transport and the translational activity in cancer cells that are conceptually pivotal for its anti-proliferative effect have not been elucidated yet. Here, we demonstrated that JPH203 drastically inhibits the transport of all the large neutral amino acids in pancreatic ductal adenocarcinoma cells. The inhibitory effects of JPH203 were observed even in competition with high concentrations of amino acids in a cell culture medium. The analyses of the nutrient-sensing mTORC1 and GAAC pathways and the protein synthesis activity revealed that JPH203 downregulates the global translation. This study demonstrates a predominant contribution of LAT1 to the transport of large neutral amino acids in cancer cells and the suppression of protein synthesis by JPH203 supposed to underly its broad anti-proliferative effects across various types of cancer cells.

Keywords:  LAT1; PDAC; SLC7A5; amino acid transporter; anti-cancer agent; global translation; inhibitor; large neutral amino acids

DOI:  https://doi.org/10.1111/jcmm.17553
Mol Genet Metab Rep. 2022 Dec;33 100913

LC-MS/MS-based enzyme assay for lysosomal acid lipase using dried blood spots.

Mari Ohira, Marianne Barr, Torayuki Okuyama, Ryuichi Mashima.

  Lysosomal acid lipase deficiency (LAL-D) (OMIM: 278000) is a lysosomal storage disorder with two distinct disease phenotypes such as Wolman disease and cholesteryl ester storage disorder (CESD), characterized by an accumulation of endocytosed cholesterol in the body. Due to the presence of multiple lipases in DBS, previous studies measured LAL enzyme activity in the presence of Lalistat-2, an established LAL-specific inhibitor (Hamilton J et al Chim Clin Acta (2012) 413:1207-1210). Alternatively, a novel substrate specific for LAL has been reported very recently (Masi S. et al Clin Chem (2018) 64:690-696). In this study, we examined the LAL enzyme activity of a Japanese population with the LAL-specific substrate using liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based enzyme assay whether an affected individual can be identified among this population. To achieve this, we first performed assay validation using LC-MS/MS. Under our experimental setting, typically we obtained LAL enzyme activity for QC High (100% enzyme activity) as 261.9 ± 3.2 μmol/h/L (n = 5) and for QC Low as (5% enzyme activity) as 14.7 ± 0.5 μmol/h/L (n = 5). The percentage of coefficient of variation for interday assay for QC High was 9.6% (n = 4) and for QC Low was 7.9% (n = 4), respectively. Based on these results, we further examined the LAL enzyme activity of control Japanese population and that of affected individuals with Wolman disease and CESD. The averaged enzyme activity for control newborns, Wolman, and CESD was 123.9 ± 53.9 μmol/h/L (n = 131), 6.6 ± 0.9 μmol/h/L (n = 3), and 4.8 ± 0.3 μmol/h/L (n = 3), respectively. These results suggest that an LAL-D-affected individual can be readily identified by enzyme activity using LC-MS/MS-based technique.

Keywords:  Cholesteryl ester storage disorder; Enzyme assay; LC-MS/MS; Lysosomal storage disorder; Wolman disease

DOI:  https://doi.org/10.1016/j.ymgmr.2022.100913
J Parkinsons Dis. 2022 Aug 29.

GBA1 and The Immune System: A Potential Role in Parkinson's Disease?

Zaid A M Al-Azzawi, Saman Arfaie, Ziv Gan-Or.

  It is clear that the immune system and inflammation have a role in Parkinson's disease (PD), including sporadic PD and some genetic forms such as LRRK2-associated PD. One of the most important genes associated with PD is GBA1, as mutations in this gene are found in 5-20% of PD patients in different populations worldwide. Biallelic mutations in GBA1 may cause Gaucher disease, a lysosomal storage disorder with involvement of the immune system, and other lines of evidence link GBA1 to the immune system and inflammation. In this review, we discuss these different pieces of evidence and whether the interplay between GBA1 and the immune system may have a role in PD.

Keywords:  GBA1; Gaucher disease; Parkinson’s disease; immune response; inflammation

DOI:  https://doi.org/10.3233/JPD-223423
Cell Death Dis. 2022 Sep 09. 13(9): 778

Deubiquitinase OTUD5 modulates mTORC1 signaling to promote bladder cancer progression.

Tao Hou, Weichao Dan, Tianjie Liu, Bo Liu, Yi Wei, Chenyang Yue, Taotao Que, Bohan Ma, Yuzeshi Lei, Zixi Wang, Jin Zeng, Yizeng Fan, Lei Li.

The mechanistic (formally "mammalian") target of rapamycin (mTOR) pathway serves as a crucial regulator of various biological processes such as cell growth and cancer progression. In bladder cancer, recent discoveries showing the cancer-promoting role of mTOR complex 1 have attracted wide attention. However, the regulation of mTOR signaling in bladder cancer is complicated and the underlying mechanism remains elusive. Here, we report that the deubiquitinating enzyme, ovarian tumor domain-containing protein 5 (OTUD5), can activate the mTOR signaling pathway, promote cancer progression, and show its oncogenic potential in bladder cancer. In our study, we found that OTUD5 deubiquitinated a RING-type E3 ligase, RNF186, and stabilized its function. In addition, the stabilization of RNF186 further led to the degradation of sestrin2, which is an inhibitor of the mTOR signaling pathway. Together, we provide novel insights into the pathogenesis of bladder cancer and first prove that OTUD5 can promote bladder cancer progression through the OTUD5-RNF186-sestrin2-mTOR axis, which may be exploited in the future for the diagnosis and treatment of this malignancy.

DOI: https://doi.org/10.1038/s41419-022-05128-6
Fac Rev. 2022 ;11 22

An alliance between lipid transfer proteins and scramblases for membrane expansion.

Joost C M Holthuis, Helene Jahn, Anant K Menon, Noboru Mizushima.

  Membrane growth requires lipid supply, which is usually accomplished by lipid synthesis or vesicular trafficking. In the case of autophagosomes, these principles do not apply. Ghanbarpour et al. postulate that autophagosome expansion relies on non-vesicular lipid delivery from the ER, whereby the activity of a lipid transfer protein (LTP) is directly coupled to scramblase activities in the donor and acceptor bilayers1. This new concept opens the possibility that lipid traffic is controlled by scramblases that provide not only specific docking sites for LTPs, thereby directing lipid flow, but also support their activity by overcoming barriers for lipid extraction and deposition.

Keywords:  Scramblase; autophagosome; lipid; lipid transfer protein; membrane

DOI:  https://doi.org/10.12703/r-01-0000015
Sci Rep. 2022 Sep 03. 12(1): 15045

Delivery and assessment of a CRISPR/nCas9-based genome editing system on in vitro models of mucopolysaccharidoses IVA assisted by magnetite-based nanoparticles.

Andrés Felipe Leal, Javier Cifuentes, Carlos Emilio Torres, Diego Suárez, Valentina Quezada, Saúl Camilo Gómez, Juan C Cruz, Luis H Reyes, Angela Johana Espejo-Mojica, Carlos Javier Alméciga-Díaz.

Mucopolysaccharidosis IV A (MPS IVA) is a lysosomal disorder caused by mutations in the GALNS gene. Consequently, the glycosaminoglycans (GAGs) keratan sulfate and chondroitin 6-sulfate accumulate in the lysosomal lumen. Although enzyme replacement therapy has shown essential advantages for the patients, several challenges remain to overcome, such as the limited impact on the bone lesion and recovery of oxidative profile. Recently, we validated a CRISPR/nCas9-based gene therapy with promising results in an in vitro MPS IVA model. In this study, we have expanded the use of this CRISPR/nCas9 system to several MPS IVA fibroblasts carrying different GALNS mutations. Considering the latent need to develop more safety vectors for gene therapy, we co-delivered the CRISPR/nCas9 system with a novel non-viral vector based on magnetoliposomes (MLPs). We found that the CRISPR/nCas9 treatment led to an increase in enzyme activity between 5 and 88% of wild-type levels, as well as a reduction in GAGs accumulation, lysosomal mass, and mitochondrial-dependent oxidative stress, in a mutation-dependent manner. Noteworthy, MLPs allowed to obtain similar results to those observed with the conventional transfection agent lipofectamine. Overall, these results confirmed the potential of CRISPR/nCas9 as a genome editing tool for treating MPS IVA. We also demonstrated the potential use of MLPs as a novel delivery system for CRISPR/nCas9-based therapies.

DOI: https://doi.org/10.1038/s41598-022-19407-x
Neurosci Biobehav Rev. 2022 Sep 06. pii: S0149-7634(22)00345-1. [Epub ahead of print] 104856

Exploring the link between GBA1 mutations and Dementia with Lewy bodies, A Mini-Review.

Sinead Gaubert, Claire Hourregue, François Mouton-Liger, Périne Millot, Mélanie Franco, Elodie Amar-Bouaziz, Dag Aarsland, Jacques Hugon, Claire Paquet.

   IMPORTANCE: Dementia with Lewy bodies (DLB) is a neurodegenerative disease linked to abnormal accumulation of phosphorylated α-synuclein. GBA1 is the gene encoding the lysosomal enzyme glucocerebrosidase (GCase), whose mutations are a risk factor of DLB.
OBJECTIVE: To report all available data exploring the association between GBA1 mutations and DLB.
EVIDENCE REVIEW: All publications focused on GCase and DLB in humans between 2003 and 2022 were identified on PubMed, Cochrane and ClinicalTrials.gov.
FINDINGS: 29 studies were included and confirmed the strong association between GBA1 mutations and DLB (Odds Ratio [OR]: 8.28). GBA1 mutation carriers presented a more malignant phenotype, with earlier symptom onset, more severe motor and cognitive dysfunctions, more visual hallucinations and rapid eye movement sleep disorder. GBA1 mutations were associated with "purer" neuropathological DLB. No therapeutic recommendations exist and clinical trials targeting GCase are just starting in DLB patients.
CONCLUSIONS AND RELEVANCE: This review reports a link between GBA1 mutations and the DLB phenotype with limited evidence due to the small number of studies.

Keywords:  Dementia with Lewy bodies; GBA1 gene; Gaucher disease; Glucocerebrosidase

DOI:  https://doi.org/10.1016/j.neubiorev.2022.104856
Nat Metab. 2022 Sep 06.

Publisher Correction: Long non-coding RNA SNHG6 couples cholesterol sensing with mTORC1 activation in hepatocellular carcinoma.

Fangzhou Liu, Tian Tian, Zhen Zhang, Shanshan Xie, Jiecheng Yang, Linyu Zhu, Wen Wang, Chengyu Shi, Lingjie Sang, Kaiqiang Guo, Zuozhen Yang, Lei Qu, Xiangrui Liu, Jian Liu, Qingfeng Yan, Huai-Qiang Ju, Wenqi Wang, Hai-Long Piao, Jianzhong Shao, Tianhua Zhou, Aifu Lin.

DOI: https://doi.org/10.1038/s42255-022-00650-5
Nat Commun. 2022 Sep 03. 13(1): 5191

The TFEB-TGIF1 axis regulates EMT in mouse epicardial cells.

Elena Astanina, Gabriella Doronzo, Davide Corà, Francesco Neri, Salvatore Oliviero, Tullio Genova, Federico Mussano, Emanuele Middonti, Edoardo Vallariello, Chiara Cencioni, Donatella Valdembri, Guido Serini, Federica Limana, Eleonora Foglio, Andrea Ballabio, Federico Bussolino.

Epithelial-mesenchymal transition (EMT) is a complex and pivotal process involved in organogenesis and is related to several pathological processes, including cancer and fibrosis. During heart development, EMT mediates the conversion of epicardial cells into vascular smooth muscle cells and cardiac interstitial fibroblasts. Here, we show that the oncogenic transcription factor EB (TFEB) is a key regulator of EMT in epicardial cells and that its genetic overexpression in mouse epicardium is lethal due to heart defects linked to impaired EMT. TFEB specifically orchestrates the EMT-promoting function of transforming growth factor (TGF) β, and this effect results from activated transcription of thymine-guanine-interacting factor (TGIF)1, a TGFβ/Smad pathway repressor. The Tgif1 promoter is activated by TFEB, and in vitro and in vivo findings demonstrate its increased expression when Tfeb is overexpressed. Furthermore, Tfeb overexpression in vitro prevents TGFβ-induced EMT, and this effect is abolished by Tgif1 silencing. Tfeb loss of function, similar to that of Tgif1, sensitizes cells to TGFβ, inducing an EMT response to low doses of TGFβ. Together, our findings reveal an unexpected function of TFEB in regulating EMT, which might provide insights into injured heart repair and control of cancer progression.

DOI: https://doi.org/10.1038/s41467-022-32855-3
Plant J. 2022 Sep 07.

Vacuolar H+ -ATPase subunit VAB3 regulates cell growth and ion homeostasis in Arabidopsis.

Wenbo Li, Laifu Luo, Lili Gu, Haimin Li, Qian Zhang, Yajin Ye, Laigeng Li.

  Vacuolar H+ -ATPase (V-ATPase) has diverse functions related to plant development and growth. It creates the turgor pressure that drives cell growth by generating the energy needed for the active transport of solutes across the tonoplast. Additionally, V-ATPase is a large protein complex made up of multiheteromeric subunits, some of which have unknown functions. In this study, a forward genetics-based strategy was employed to identify the vab3 mutant, which displayed resistance to isoxaben, a cellulose synthase inhibitor that could induce excessive transverse cell expansion. Map-based cloning and genetic complementary assays demonstrated that V-ATPase B subunit 3 (VAB3) was associated with the observed insensitivity of the mutant to isoxaben. Analysis of the vab3 mutant revealed defective ionic homeostasis and hypersensitivity to salt stress. Treatment with a V-ATPase inhibitor exacerbated ionic tolerance and cell elongation defects in vab3 mutant. Notably, exogenous low-dose Ca2+ or Na+ could partially restore the isoxben resistance of vab3 mutant, suggesting a relationship between VAB3-regulated cell growth and ion homeostasis. Taken together, the results of this study suggest that the V-ATPase subunit VAB3 is required for cell growth and ion homeostasis in Arabidopsis.

Keywords:  Arabidopsis thaliana; Cell growth; V-ATPase; VAB3; VHA-B; ion

DOI:  https://doi.org/10.1111/tpj.15971
Sci Rep. 2022 Sep 05. 12(1): 15095

Tyrosine phosphatase activity is restricted by basic charge substituting mutation of substrates.

Che-Fan Huang, Cara J Gottardi, Milan Mrksich.

Phosphorylation controls important cellular signals and its dysregulation leads to disease. While most phospho-regulation studies are focused on kinases, phosphatases are comparatively overlooked. Combining peptide arrays with SAMDI mass spectrometry, we show that tyrosine phosphatase activity is restricted by basic amino acids adjacent to phosphotyrosines. We validate this model using two β-catenin mutants associated with cancer (T653R/K) and a mouse model for intellectual disability (T653K). These mutants introduce a basic residue next to Y654, an established phosphorylation site where modification shifts β-catenin from cell-cell adhesions and towards its essential nuclear role as Wnt-signaling effector. We show that T653-basic mutant β-catenins are less efficiently dephosphorylated by phosphatases, leading to sustained Y654 phosphorylation and elevated Wnt signals, similar to those observed for Y654E phospho-mimic mutant mice. This model rationalizes how basic mutations proximal to phosphotyrosines can restrict counter-regulation by phosphatases, providing new mechanismistic and treatment insights for 6000+ potentially relevant cancer mutations.

DOI: https://doi.org/10.1038/s41598-022-19133-4
Int J Mol Sci. 2022 Sep 04. pii: 10132. [Epub ahead of print]23(17):

Targeting PI3K/AKT/mTOR Signaling Pathway in Pancreatic Cancer: From Molecular to Clinical Aspects.

Silviu Stanciu, Florentina Ionita-Radu, Constantin Stefani, Daniela Miricescu, Iulia-Ioana Stanescu-Spinu, Maria Greabu, Alexandra Ripszky Totan, Mariana Jinga.

  Although pancreatic cancer (PC) was considered in the past an orphan cancer type due to its low incidence, it may become in the future one of the leading causes of cancer death. Pancreatic ductal adenocarcinoma (PDAC) is the most frequent type of PC, being a highly aggressive malignancy and having a 5-year survival rate of less than 10%. Non-modifiable (family history, age, genetic susceptibility) and modifiable (smoking, alcohol, acute and chronic pancreatitis, diabetes mellitus, intestinal microbiota) risk factors are involved in PC pathogenesis. Chronic inflammation induced by various factors plays crucial roles in PC development from initiation to metastasis. In multiple malignant conditions such as PC, cytokines, chemokines, and growth factors activate the class I phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) (PI3K/AKT/mTOR) signaling pathway, which plays key roles in cell growth, survival, proliferation, metabolism, and motility. Currently, mTOR, AKT, and PI3K inhibitors are used in clinical studies. Moreover, PI3K/mTOR dual inhibitors are being tested in vitro and in vivo with promising results for PC patients. The main aim of this review is to present PC incidence, risk factors, tumor microenvironment development, and PI3K/AKT/mTOR dysregulation and inhibitors used in clinical, in vivo, and in vitro studies.

Keywords:  PI3K/AKT/mTOR; inhibitors; pancreatic cancer; risk factors; tumor microenvironment

DOI:  https://doi.org/10.3390/ijms231710132
Hum Mol Genet. 2022 Sep 06. pii: ddac225. [Epub ahead of print]

Germline homozygous missense DEPDC5 variants cause severe refractory early-onset epilepsy, macrocephaly and bilateral polymicrogyria.

Athina Ververi, Sara Zagaglia, Lara Menzies, Julia Baptista, Richard Caswell, Stephanie Baulac, Sian Ellard, Sally Lynch, Genomics England Research Consortium, Thomas S Jacques, Maninder Singh Chawla, Martin Heier, Mari Ann Kulseth, Inger-Lise Mero, Anne Katrine Våtevik, Ichraf Kraoua, Hanene Ben Rhouma, Thouraya Ben Younes, Zouhour Miladi, Ilhem Ben Youssef Turki, Wendy D Jones, Emma Clement, Christin Eltze, Kshitij Mankad, Ashirwad Merve, Jennifer Parker, Bethan Hoskins, Ronit Pressler, Sniya Sudhakar, Catherine DeVile, Tessa Homfray, Marios Kaliakatsos, Prab Prabhakar Ponnudas, Robert Robinson, Sara Margrete Bøen Keim, Imen Habibi, Alexandre Reymond, Sanjay M Sisodiya, Jane A Hurst.

PURPOSE: DEPDC5 (DEP Domain-Containing Protein 5) encodes an inhibitory component of the mTOR pathway and is commonly implicated in sporadic and familial focal epilepsies, both non-lesional and in association with focal cortical dysplasia. Germline pathogenic variants are typically heterozygous and inactivating. We describe a novel phenotype caused by germline biallelic missense variants in DEPDC5.
METHODS: Cases were identified clinically. Available records, including MRI and EEG, were reviewed. Genetic testing was performed by whole exome and whole genome sequencing and cascade screening. In addition, immunohistochemistry was performed on skin biopsy.
RESULTS: The phenotype was identified in nine children, eight of which are described in detail herein. Six of the children were of Irish Traveller, two of Tunisian and one of Lebanese origin. The Irish Traveller children shared the same DEPDC5 germline homozygous missense variant (p.Thr337Arg), whereas the Lebanese and Tunisian children shared a different germline homozygous variant (p.Arg806Cys). Consistent phenotypic features included extensive bilateral polymicrogyria, congenital macrocephaly and early-onset refractory epilepsy, in keeping with other mTOR-opathies. Eye and cardiac involvement, and severe neutropenia, were also observed in one or more patients. Five of the children died in infancy or childhood, the other four are currently aged between five months and six years. Skin biopsy immunohistochemistry was supportive of hyperactivation of the mTOR pathway.
DISCUSSION: The clinical, histopathological and genetic evidence supports a causal role for the homozygous DEPDC5 variants, expanding our understanding of the biology of this gene.

DOI: https://doi.org/10.1093/hmg/ddac225
J Pathol. 2022 Sep 08.

Long-term treatment with low dose rapamycin extends lifespan and delays tumours in cancer-prone germline PTEN mutant mice.

Priyanka Tibarewal, Victoria Rathbone, Georgia Constantinou, Wayne Pearce, Mahreen Adil, Zofia Varyova, Lisa Folkes, Alix Hampson, Gala Anastasia Electra Classen, Adriana Alves, Sara Carvalho, Cheryl L Scudamore, Bart Vanhaesebroeck.

  PTEN is one of the most commonly inactivated tumour suppressor genes in sporadic cancer. Germline heterozygous PTEN gene alterations also underly the PTEN Hamartoma Tumour Syndrome (PHTS), a rare human cancer-predisposition condition. A key feature of systemic PTEN deregulation is the inability to adequately dampen PI-3-kinase (PI3K)/mTORC1 signalling. PI3K/mTORC1 pathway inhibitors such as rapamycin are therefore expected to neutralize the impact of PTEN loss, rendering this a more druggable context compared to those of other tumour suppressor pathways such as loss of TP53. However, this has not been explored in cancer prevention in a model of germline cancer predisposition, such as PHTS. Clinical trials of short-term treatment with rapamycin have recently been initiated for PHTS, focusing on cognition and colon polyposis. Here, we administered a low dose of rapamycin from the age of 6-weeks onwards to mice with heterozygous germline PTEN loss, a mouse model that recapitulates most characteristics of human PHTS. Rapamycin was well-tolerated and led to a highly significant improvement of survival in both male and female mice. This was accompanied by a delay, but not full blockade of the development of a range of proliferative lesions, including gastro-intestinal and thyroid tumours and endometrial hyperplasia, with no impact on mammary and prostate tumours, and no effect on brain overgrowth. Our data indicate that rapamycin may have cancer prevention potential in human PHTS. This might also be the case for sporadic cancers in which genetic PI3K pathway activation is an early event in tumour development, such as endometrial cancer and some breast cancers. To the best of our knowledge, this is the first report of a long-term treatment of a germline cancer predisposition model with a PI3K/mTOR pathway inhibitor This article is protected by copyright. All rights reserved.

Keywords:  PHTS; PI 3-kinase; PTEN; cancer prevention; drug; hamartoma; kinase inhibitor; mTORC1; rapamycin; rare disease; syndrome

DOI:  https://doi.org/10.1002/path.6009