bims-lysosi 2021-01-10 papers

bims-lysosi

Biomed News

on Lysosomes and signaling

Issue of 2021–01–10
37 papers selected by
Stephanie Fernandes, Max Planck Institute for Biology of Ageing

mTORC1 promotes TOP mRNA translation through site-specific phosphorylation of LARP1.
A septin GTPase scaffold of dynein-dynactin motors triggers retrograde lysosome transport.
Lysosomotropic agents including azithromycin, chloroquine and hydroxychloroquine activate the integrated stress response.
Modulating lysosomal pH: a molecular and nanoscale materials design perspective.
Membrane Protein Quality Control Mechanisms in the Endo-Lysosome System.
Endolysosomal TRPMLs in Cancer.
Targeting Cancer Lysosomes with Good Old Cationic Amphiphilic Drugs.
Enzyme Replacement Therapy for Mucopolysaccharidosis IIID using Recombinant Human α-N-Acetylglucosamine-6-Sulfatase in Neonatal Mice.
Exosomes induce endolysosomal permeabilization as a gateway by which exosomal tau seeds escape into the cytosol.
Autophagy sustains glutamate and aspartate synthesis in Saccharomyces cerevisiae during nitrogen starvation.
Transcription factor EB‑mediated autophagy promotes dermal fibroblast differentiation and collagen production by regulating endoplasmic reticulum stress and autophagy‑dependent secretion.
Placental trophoblast syncytialization potentiates macropinocytosis via mTOR signaling to adapt to reduced amino acid supply.
Revisiting Brain Tuberous Sclerosis Complex in Rat and Human: Shared Molecular and Cellular Pathology Leads to Distinct Neurophysiological and Behavioral Phenotypes.
Mechanism and restoration strategy of lysosomal abnormalities induced by urinary protein overload in proximal tubule epithelial cells.
mTORC1 and mTORC2 converge on the Arp2/3 complex to promote KrasG12D-induced acinar-to-ductal metaplasia and early pancreatic carcinogenesis.
Phosphorylation of PKCδ by FER tips the balance from EGFR degradation to recycling.
Exosome Release Is Modulated by the Mitochondrial-Lysosomal Crosstalk in Parkinson's Disease Stress Conditions.
Build-UPS and break-downs: metabolism impacts on proteostasis and aging.
Novel autophagic vacuolar myopathies: phenotype and genotype features.
The Harmonious Interplay of Amino Acid and Monocarboxylate Transporters Induces the Robustness of Cancer Cells.
Nano CaCO3 "Lysosomal Bombs" Enhance Chemotherapy Drug Efficacy via Rebalancing Tumor Intracellular pH.
Transcription factor EB regulates cardiovascular homeostasis.
MTORC1-Regulated Metabolism Controlled by TSC2 Limits Cardiac Reperfusion Injury.
Decrease in Myelin-Associated Lipids Precedes Neuronal Loss and Glial Activation in the CNS of the Sandhoff Mouse as Determined by Metabolomics.
AMPK/mTOR Signaling in Autophagy Regulation During Cisplatin-Induced Acute Kidney Injury.
Function of Deptor and its roles in hematological malignancies.
Harnessing the Co-vulnerabilities of Amino Acid-Restricted Cancers.
Amino Acid Transporters on the Guard of Cell Genome and Epigenome.
The amino acid transporter SLC7A5 is required for efficient growth of KRAS-mutant colorectal cancer.
A cross-platform approach identifies genetic regulators of human metabolism and health.
mTORC2 deploys the mRNA binding protein IGF2BP1 to regulate c-MYC expression and promote cell survival.
Metabolites and the tumour microenvironment: from cellular mechanisms to systemic metabolism.
Extracellular vesicles released from the filarial parasite Brugia malayi downregulate the host mTOR pathway.
Selective autophagy of intracellular organelles: recent research advances.
Cyclin-dependent Kinase 1 and Aurora Kinase choreograph mitotic storage and redistribution of a growth factor receptor.
Dysfunctional Autophagy and Endolysosomal System in Neurodegenerative Diseases: Relevance and Therapeutic Options.
Mitochondrial long non-coding RNA GAS5 tunes TCA metabolism in response to nutrient stress.

Nucleic Acids Res. 2021 Jan 04. pii: gkaa1239. [Epub ahead of print]

mTORC1 promotes TOP mRNA translation through site-specific phosphorylation of LARP1.

Jian-Jun Jia, Roni M Lahr, Michael T Solgaard, Bruno J Moraes, Roberta Pointet, An-Dao Yang, Giovanna Celucci, Tyson E Graber, Huy-Dung Hoang, Marius R Niklaus, Izabella A Pena, Anne K Hollensen, Ewan M Smith, Malik Chaker-Margot, Leonie Anton, Christopher Dajadian, Mark Livingstone, Jaclyn Hearnden, Xu-Dong Wang, Yonghao Yu, Timm Maier, Christian K Damgaard, Andrea J Berman, Tommy Alain, Bruno D Fonseca.

LARP1 is a key repressor of TOP mRNA translation. It binds the m7Gppp cap moiety and the adjacent 5'TOP motif of TOP mRNAs, thus impeding the assembly of the eIF4F complex on these transcripts. mTORC1 controls TOP mRNA translation via LARP1, but the details of the mechanism are unclear. Herein we elucidate the mechanism by which mTORC1 controls LARP1's translation repression activity. We demonstrate that mTORC1 phosphorylates LARP1 in vitro and in vivo, activities that are efficiently inhibited by rapamycin and torin1. We uncover 26 rapamycin-sensitive phospho-serine and -threonine residues on LARP1 that are distributed in 7 clusters. Our data show that phosphorylation of a cluster of residues located proximally to the m7Gppp cap-binding DM15 region is particularly sensitive to rapamycin and regulates both the RNA-binding and the translation inhibitory activities of LARP1. Our results unravel a new model of translation control in which the La module (LaMod) and DM15 region of LARP1, both of which can directly interact with TOP mRNA, are differentially regulated: the LaMod remains constitutively bound to PABP (irrespective of the activation status of mTORC1), while the C-terminal DM15 'pendular hook' engages the TOP mRNA 5'-end to repress translation, but only in conditions of mTORC1 inhibition.

DOI: https://doi.org/10.1093/nar/gkaa1239
J Cell Biol. 2021 Feb 01. pii: e202005219. [Epub ahead of print]220(2):

A septin GTPase scaffold of dynein-dynactin motors triggers retrograde lysosome transport.

Ilona A Kesisova, Benjamin P Robinson, Elias T Spiliotis.

The metabolic and signaling functions of lysosomes depend on their intracellular positioning and trafficking, but the underlying mechanisms are little understood. Here, we have discovered a novel septin GTPase-based mechanism for retrograde lysosome transport. We found that septin 9 (SEPT9) associates with lysosomes, promoting the perinuclear localization of lysosomes in a Rab7-independent manner. SEPT9 targeting to mitochondria and peroxisomes is sufficient to recruit dynein and cause perinuclear clustering. We show that SEPT9 interacts with both dynein and dynactin through its GTPase domain and N-terminal extension, respectively. Strikingly, SEPT9 associates preferentially with the dynein intermediate chain (DIC) in its GDP-bound state, which favors dimerization and assembly into septin multimers. In response to oxidative cell stress induced by arsenite, SEPT9 localization to lysosomes is enhanced, promoting the perinuclear clustering of lysosomes. We posit that septins function as GDP-activated scaffolds for the cooperative assembly of dynein-dynactin, providing an alternative mechanism of retrograde lysosome transport at steady state and during cellular adaptation to stress.

DOI: https://doi.org/10.1083/jcb.202005219
Cell Death Dis. 2021 Jan 06. 12(1): 6

Lysosomotropic agents including azithromycin, chloroquine and hydroxychloroquine activate the integrated stress response.

Ai-Ling Tian, Qi Wu, Peng Liu, Liwei Zhao, Isabelle Martins, Oliver Kepp, Marion Leduc, Guido Kroemer.

The integrated stress response manifests with the phosphorylation of eukaryotic initiation factor 2α (eIF2α) on serine residue 51 and plays a major role in the adaptation of cells to endoplasmic reticulum stress in the initiation of autophagy and in the ignition of immune responses. Here, we report that lysosomotropic agents, including azithromycin, chloroquine, and hydroxychloroquine, can trigger eIF2α phosphorylation in vitro (in cultured human cells) and, as validated for hydroxychloroquine, in vivo (in mice). Cells bearing a non-phosphorylatable eIF2α mutant (S51A) failed to accumulate autophagic puncta in response to azithromycin, chloroquine, and hydroxychloroquine. Conversely, two inhibitors of eIF2α dephosphorylation, nelfinavir and salubrinal, enhanced the induction of such autophagic puncta. Altogether, these results point to the unexpected capacity of azithromycin, chloroquine, and hydroxychloroquine to elicit the integrated stress response.

DOI: https://doi.org/10.1038/s41419-020-03324-w
J Life Sci (Westlake Village). 2020 Dec;2(4): 25-37

Modulating lysosomal pH: a molecular and nanoscale materials design perspective.

Jialiu Zeng, Orian S Shirihai, Mark W Grinstaff.

Lysosomes, membrane-bound organelles, play important roles in cellular processes including endocytosis, phagocytosis, and autophagy. Lysosomes maintain cellular homeostasis by generating a highly acidic environment of pH 4.5 - 5.0 and by housing hydrolytic enzymes that degrade engulfed biomolecules. Impairment of lysosomal function, especially in its acidification, is a driving force in the pathogenesis of diseases including neurodegeneration, cancer, metabolic disorders, and infectious diseases. Therefore, lysosomal pH is an attractive and targetable site for therapeutic intervention. Currently, there is a dearth of strategies or materials available to specifically modulate lysosomal acidification. This review focuses on the key aspects of how lysosomal pH is implicated in various diseases and discusses design strategies and molecular or nanoscale agents for lysosomal pH modulation, with the ultimate goal of developing novel therapeutic solutions.

DOI: https://doi.org/10.36069/jols/20201204
Trends Cell Biol. 2021 Jan 04. pii: S0962-8924(20)30246-4. [Epub ahead of print]

Membrane Protein Quality Control Mechanisms in the Endo-Lysosome System.

Richa Sardana, Scott D Emr.

  Protein quality control (PQC) machineries play a critical role in selective identification and removal of mistargeted, misfolded, and aberrant proteins. This task is extremely complicated due to the enormous diversity of the proteome. It also requires nuanced and careful differentiation between 'normal' and 'folding intermediates' from 'abnormal' and 'misfolded' protein states. Multiple genetic and proteomic approaches have started to delineate the molecular underpinnings of how these machineries recognize their target and how their activity is regulated. In this review, we summarize our understanding of the various E3 ubiquitin ligases and associated machinery that mediate PQC in the endo-lysosome system in yeast and humans, how they are regulated, and mechanisms of target selection, with the intent of guiding future research in this area.

Keywords:  E3 ligase adaptors; E3 ubiquitin ligase; HECT; Nedd4; RING; RING-CH; Rsp5; endo-lysosome system; lysosome; membrane protein quality control

DOI:  https://doi.org/10.1016/j.tcb.2020.11.011
Biomolecules. 2021 Jan 06. pii: E65. [Epub ahead of print]11(1):

Endolysosomal TRPMLs in Cancer.

Mengnan Xu, Xian-Ping Dong.

  Lysosomes, the degradative endpoints and sophisticated cellular signaling hubs, are emerging as intracellular Ca2+ stores that govern multiple cellular processes. Dys-homeostasis of lysosomal Ca2+ is intimately associated with a variety of human diseases including cancer. Recent studies have suggested that the Ca2+-permeable channels Transient Receptor Potential (TRP) Mucolipins (TRPMLs, TRPML1-3) integrate multiple processes of cell growth, division and metabolism. Dysregulation of TRPMLs activity has been implicated in cancer development. In this review, we provide a summary of the latest development of TRPMLs in cancer. The expression of TRPMLs in cancer, TRPMLs in cancer cell nutrient sensing, TRPMLs-mediated lysosomal exocytosis in cancer development, TRPMLs in TFEB-mediated gene transcription of cancer cells, TRPMLs in bacteria-related cancer development and TRPMLs-regulated antitumor immunity are discussed. We hope to guide readers toward a more in-depth discussion of the importance of lysosomal TRPMLs in cancer progression and other human diseases.

Keywords:  autophagy; calcium; cancer; endolysosome; ion channel

DOI:  https://doi.org/10.3390/biom11010065
Rev Physiol Biochem Pharmacol. 2021 Jan 05.

Targeting Cancer Lysosomes with Good Old Cationic Amphiphilic Drugs.

Anne-Marie Ellegaard, Peter Bach, Marja Jäättelä.

  Being originally discovered as cellular recycling bins, lysosomes are today recognized as versatile signaling organelles that control a wide range of cellular functions that are essential not only for the well-being of normal cells but also for malignant transformation and cancer progression. In addition to their core functions in waste disposal and recycling of macromolecules and energy, lysosomes serve as an indispensable support system for malignant phenotype by promoting cell growth, cytoprotective autophagy, drug resistance, pH homeostasis, invasion, metastasis, and genomic integrity. On the other hand, malignant transformation reduces the stability of lysosomal membranes rendering cancer cells sensitive to lysosome-dependent cell death. Notably, many clinically approved cationic amphiphilic drugs widely used for the treatment of other diseases accumulate in lysosomes, interfere with their cancer-promoting and cancer-supporting functions and destabilize their membranes thereby opening intriguing possibilities for cancer therapy. Here, we review the emerging evidence that supports the supplementation of current cancer therapies with lysosome-targeting cationic amphiphilic drugs.

Keywords:  Cancer; Cathepsins; Cationic amphiphilic drugs; Cell death; Lysosome; SMPD1; pH

DOI:  https://doi.org/10.1007/112_2020_56
Mol Pharm. 2021 Jan 04. 18(1): 214-227

Enzyme Replacement Therapy for Mucopolysaccharidosis IIID using Recombinant Human α-N-Acetylglucosamine-6-Sulfatase in Neonatal Mice.

Feng Wang, Derek R Moen, Chelsee Sauni, Shih-Hsin Kan, Shan Li, Steven Q Le, Brett Lomenick, Xiaoyi Zhang, Sean Ekins, Srikanth Singamsetty, Jill Wood, Patricia I Dickson, Tsui-Fen Chou.

  There is currently no cure or effective treatment available for mucopolysaccharidosis type IIID (MPS IIID, Sanfilippo syndrome type D), a lysosomal storage disorder (LSD) caused by the deficiency of α-N-acetylglucosamine-6-sulfatase (GNS). The clinical symptoms of MPS IIID, like other subtypes of Sanfilippo syndrome, are largely localized to the central nervous system (CNS), and any treatments aiming to ameliorate or reverse the catastrophic and fatal neurologic decline caused by this disease need to be delivered across the blood-brain barrier. Here, we report a proof-of-concept enzyme replacement therapy (ERT) for MPS IIID using recombinant human α-N-acetylglucosamine-6-sulfatase (rhGNS) via intracerebroventricular (ICV) delivery in a neonatal MPS IIID mouse model. We overexpressed and purified rhGNS from CHO cells with a specific activity of 3.9 × 104 units/mg protein and a maximal enzymatic activity at lysosomal pH (pH 5.6), which was stable for over one month at 4 °C in artificial cerebrospinal fluid (CSF). We demonstrated that rhGNS was taken up by MPS IIID patient fibroblasts via the mannose 6-phosphate (M6P) receptor and reduced intracellular glycosaminoglycans to normal levels. The delivery of 5 μg of rhGNS into the lateral cerebral ventricle of neonatal MPS IIID mice resulted in normalization of the enzymatic activity in brain tissues; rhGNS was found to be enriched in lysosomes in MPS IIID-treated mice relative to the control. Furthermore, a single dose of rhGNS was able to reduce the accumulated heparan sulfate and β-hexosaminidase. Our results demonstrate that rhGNS delivered into CSF is a potential therapeutic option for MPS IIID that is worthy of further development.

Keywords:  CHO cells; DHFR; GNS; MPS IIID; enzyme replacement therapy; recombinant protein production

DOI:  https://doi.org/10.1021/acs.molpharmaceut.0c00831
Acta Neuropathol. 2021 Jan 08.

Exosomes induce endolysosomal permeabilization as a gateway by which exosomal tau seeds escape into the cytosol.

Juan Carlos Polanco, Gabriel Rhys Hand, Adam Briner, Chuanzhou Li, Jürgen Götz.

  The microtubule-associated protein tau has a critical role in Alzheimer's disease and other tauopathies. A proposed pathomechanism in the progression of tauopathies is the trans-synaptic spreading of tau seeds, with a role for exosomes which are secretory nanovesicles generated by late endosomes. Our previous work demonstrated that brain-derived exosomes isolated from tau transgenic rTg4510 mice encapsulate tau seeds with the ability to induce tau aggregation in recipient cells. We had also shown that exosomes can hijack the endosomal pathway to spread through interconnected neurons. Here, we reveal how tau seeds contained within internalized exosomes exploit mechanisms of lysosomal degradation to escape the endosome and induce tau aggregation in the cytosol of HEK293T-derived 'tau biosensor cells'. We found that the majority of the exosome-containing endosomes fused with lysosomes to form endolysosomes. Exosomes induced their permeabilization, irrespective of the presence of tau seeds, or whether the exosomal preparations originated from mouse brains or HEK293T cells. We also found that permeabilization is a conserved mechanism, operating in both non-neuronal tau biosensor cells and primary neurons. However, permeabilization of endolysosomes only occurred in a small fraction of cells, which supports the notion that permeabilization occurs by a thresholded mechanism. Interestingly, tau aggregation was only induced in cells that exhibited permeabilization, presenting this as an escape route of exosomal tau seeds into the cytosol. Overexpression of RAB7, which is required for the formation of endolysosomes, strongly increased tau aggregation. Conversely, inhibition of lysosomal function with alkalinizing agents, or by knocking-down RAB7, decreased tau aggregation. Together, we conclude that the enzymatic activities of lysosomes permeabilize exosomal and endosomal membranes, thereby facilitating access of exosomal tau seeds to cytosolic tau to induce its aggregation. Our data underscore the importance of endosomal membrane integrity in mechanisms of cellular invasion by misfolded proteins that are resistant to lysosomal degradation.

Keywords:  Alzheimer’s disease; Autophagy; Endosome; Exosome; Lysosome; Microtubule-associated protein tau; Protein aggregation; Spreading; seeding

DOI:  https://doi.org/10.1007/s00401-020-02254-3
Nat Commun. 2021 01 04. 12(1): 57

Autophagy sustains glutamate and aspartate synthesis in Saccharomyces cerevisiae during nitrogen starvation.

Kuanqing Liu, Benjamin M Sutter, Benjamin P Tu.

Autophagy catabolizes cellular constituents to promote survival during nutrient deprivation. Yet, a metabolic comprehension of this recycling operation, despite its crucial importance, remains incomplete. Here, we uncover a specific metabolic function of autophagy that exquisitely adjusts cellular metabolism according to nitrogen availability in the budding yeast Saccharomyces cerevisiae. Autophagy enables metabolic plasticity to promote glutamate and aspartate synthesis, which empowers nitrogen-starved cells to replenish their nitrogen currency and sustain macromolecule synthesis. Our findings provide critical insights into the metabolic basis by which autophagy recycles cellular components and may also have important implications in understanding the role of autophagy in diseases such as cancer.

DOI: https://doi.org/10.1038/s41467-020-20253-6
Int J Mol Med. 2020 Dec 09.

Transcription factor EB‑mediated autophagy promotes dermal fibroblast differentiation and collagen production by regulating endoplasmic reticulum stress and autophagy‑dependent secretion.

Ling Zhou, Zeming Liu, Sichao Chen, Jing Qiu, Qianqian Li, Shipei Wang, Wei Zhou, Danyang Chen, Guang Yang, Liang Guo.

Autophagy is reported to be involved in the formation of skin hypertrophic scar (HTS). However, the role of autophagy in the process of fibrosis remains unclear, therefore an improved understanding of the molecular mechanisms associated with autophagy may accelerate the development of effective therapeutic strategies against HTS. The present study evaluated the roles of autophagy mediated by transcription factor EB (TFEB), a pivotal regulator of lysosome biogenesis and autophagy, in transforming growth factor‑β1 (TGF‑β1)‑induced fibroblast differentiation and collagen production. Fibroblasts were treated with TGF‑β1, TGF‑β1 + tauroursodeoxycholic acid (TUDCA) or TGF‑β1 + TFEB‑small interfering RNA (siRNA). TGF‑β1 induced phenotypic transformation of fibroblasts, as well as collagen synthesis and secretion in fibroblasts in a dose‑dependent manner. Western blotting and immunofluorescence analyses demonstrated that TGF‑β1 upregulated the expression of autophagy‑related proteins through the endoplasmic reticulum (ER) stress pathway, whereas TUDCA reversed TGF‑β1‑induced changes. Reverse transcription‑quantitative PCR (RT‑qPCR), western blotting and RFP‑GFP‑LC3 double fluorescence analyses demonstrated that knockdown of TFEB by TFEB‑siRNA decreased autophagic flux, upregulated the expression of proteins involved in the apoptotic pathway, such as phosphorylated‑α subunit of eukaryotic initiation factor 2, C/EBP homologous protein and cysteinyl aspartate specific proteinase 3, and also downregulated the expression of α‑smooth muscle actin and collagen I (COL I) in fibroblasts. Immunofluorescence confocal analyses and enzyme‑linked immunosorbent assay indicated that TGF‑β1 increased the colocalization of COL I with lysosomal‑associated membrane protein 1 and Ras‑related protein Rab‑8A, a marker of secretory vesicles, in fibroblasts, as well as the secretion of pro‑COL Iα1 in culture supernatants. Meanwhile, these effects were abolished by TFEB knockdown. The present results suggested that autophagy reduced ER stress, decreased cell apoptosis and maintained fibroblast activation not only through degradation of misfolded or unfolded proteins, but also through promotion of COL I release from the autolysosome to the extracellular environment.

DOI: https://doi.org/10.3892/ijmm.2020.4814
Proc Natl Acad Sci U S A. 2021 Jan 19. pii: e2017092118. [Epub ahead of print]118(3):

Placental trophoblast syncytialization potentiates macropinocytosis via mTOR signaling to adapt to reduced amino acid supply.

Xuan Shao, Guangming Cao, Dunjin Chen, Juan Liu, Bolan Yu, Ming Liu, Yu-Xia Li, Bin Cao, Yoel Sadovsky, Yan-Ling Wang.

  During pregnancy, the appropriate allocation of nutrients between the mother and the fetus is dominated by maternal-fetal interactions, which is primarily governed by the placenta. The syncytiotrophoblast (STB) lining at the outer surface of the placental villi is directly bathed in maternal blood and controls feto-maternal exchange. The STB is the largest multinucleated cell type in the human body, and is formed through syncytialization of the mononucleated cytotrophoblast. However, the physiological advantage of forming such an extensively multinucleated cellular structure remains poorly understood. Here, we discover that the STB uniquely adapts to nutrient stress by inducing the macropinocytosis machinery through repression of mammalian target of rapamycin (mTOR) signaling. In primary human trophoblasts and in trophoblast cell lines, differentiation toward a syncytium triggers macropinocytosis, which is greatly enhanced during amino acid shortage, induced by inhibiting mTOR signaling. Moreover, inhibiting mTOR in pregnant mice markedly stimulates macropinocytosis in the syncytium. Blocking macropinocytosis worsens the phenotypes of fetal growth restriction caused by mTOR-inhibition. Consistently, placentas derived from fetal growth restriction patients display: 1) Repressed mTOR signaling, 2) increased syncytialization, and 3) enhanced macropinocytosis. Together, our findings suggest that the unique ability of STB to undergo macropinocytosis serves as an essential adaptation to the cellular nutrient status, and support fetal survival and growth under nutrient deprivation.

Keywords:  amino acid shortage; fetal growth; mTOR; macropinocytosis; placental syncytiotrophoblast

DOI:  https://doi.org/10.1073/pnas.2017092118
Neurotherapeutics. 2021 Jan 04.

Revisiting Brain Tuberous Sclerosis Complex in Rat and Human: Shared Molecular and Cellular Pathology Leads to Distinct Neurophysiological and Behavioral Phenotypes.

Viera Kútna, Valerie B O'Leary, Ehren Newman, Cyril Hoschl, Saak V Ovsepian.

  Tuberous sclerosis complex (TSC) is a dominant autosomal genetic disorder caused by loss-of-function mutations in TSC1 and TSC2, which lead to constitutive activation of the mammalian target of rapamycin C1 (mTORC1) with its decoupling from regulatory inputs. Because mTORC1 integrates an array of molecular signals controlling protein synthesis and energy metabolism, its unrestrained activation inflates cell growth and division, resulting in the development of benign tumors in the brain and other organs. In humans, brain malformations typically manifest through a range of neuropsychiatric symptoms, among which mental retardation, intellectual disabilities with signs of autism, and refractory seizures, which are the most prominent. TSC in the rat brain presents the first-rate approximation of cellular and molecular pathology of the human brain, showing many instructive characteristics. Nevertheless, the developmental profile and distribution of lesions in the rat brain, with neurophysiological and behavioral manifestation, deviate considerably from humans, raising numerous research and translational questions. In this study, we revisit brain TSC in human and Eker rats to relate their histopathological, electrophysiological, and neurobehavioral characteristics. We discuss shared and distinct aspects of the pathology and consider factors contributing to phenotypic discrepancies. Given the shared genetic cause and molecular pathology, phenotypic deviations suggest an incomplete understanding of the disease. Narrowing the knowledge gap in the future should not only improve the characterization of the TSC rat model but also explain considerable variability in the clinical manifestation of the disease in humans.

Keywords:  TSC1; TSC2; autism spectrum disorders; hamartoma; mTOR signaling; neoplasia; refractory epilepsy

DOI:  https://doi.org/10.1007/s13311-020-01000-7
Dev Dyn. 2021 Jan 06.

Mechanism and restoration strategy of lysosomal abnormalities induced by urinary protein overload in proximal tubule epithelial cells.

Xiaoyu Li, Ting Zou, Shujun Wang, Hongluan Wu, Man Wu, Zejian Liu, Huafeng Liu.

   BACKGROUND: Persistent elevated concentrations of urinary protein can destroy proximal tubule epithelial cells (PTECs) by inducing lysosomal abnormalities, thereby aggravating PTEC damage and renal fibrosis. However, the specific mechanisms of these serial biochemical events and methods for treating or preventing PTEC damage upon proteinuria need further investigation.
RESULTS: In this study, electron microscopy and dual-labeled immunofluorescence analysis for identifying lysosome type revealed inadequate primary lysosome biogenesis and secondary lysosome accumulation in the PTECs of patients with minimal change nephrotic syndrome or membranous nephropathy who suffered from proteinuria. In vitro studies on HK-2 cells indicated that this abnormality was associated with decreased expression of transcription factor EB (TFEB). In contrast, TFEB overexpressing HK-2 cells under urinary protein overload exhibited significantly reduced accumulation of secondary lysosomes and increased proportion and quantity of primary lysosomes as indicated by dual-labeled immunofluorescence. Further, these cells could upregulate lysosomal degradation functions, as determined using Cathepsin L activity assays and flow cytometry for dye quenched-albumin.
CONCLUSIONS: These results indicate that abnormal TFEB expression is a key mechanism of lysosomal dyshomeostasis caused by protein overload in PTECs. TFEB is thus a potential therapeutic target for the treatment of urinary protein-related kidney disease.

Keywords:  lysosomal dyshomeostasis; proteinuria; proximal tubule epithelial cells; transcription factor EB

DOI:  https://doi.org/10.1002/dvdy.297
Gastroenterology. 2020 Dec 31. pii: S0016-5085(20)35626-2. [Epub ahead of print]

mTORC1 and mTORC2 converge on the Arp2/3 complex to promote KrasG12D-induced acinar-to-ductal metaplasia and early pancreatic carcinogenesis.

Yamin Zhao, Benjamin Schoeps, Dianbo Yao, Zhiheng Zhang, Kathleen Schuck, Vivien Tissen, Carsten Jäger, Anna Melissa Schlitter, Rob van der Kammen, Christina Ludwig, Jan G D'Haese, Susanne Raulefs, Nadja Maeritz, Shanshan Shen, Xiaoping Zou, Achim Krüger, Jörg Kleeff, Christoph W Michalski, Helmut Friess, Metello Innocenti, Bo Kong.

   BACKGROUND AND AIM: Oncogenic KrasG12D induces neoplastic transformation of pancreatic acinar cells through acinar-to-ductal metaplasia (ADM), an actin-based morphogenetic process, and drives pancreatic ductal adenocarcinoma (PDAC). mTOR (mechanistic target of rapamycin kinase) complex 1 (mTORC1) and 2 (mTORC2) contain Rptor and Rictor, respectively, and are activated downstream of KrasG12D thereby contributing to PDAC. Yet, whether and how mTORC1 and mTORC2 impact on ADM and the identity of the actin nucleator(s) mediating such actin rearrangements remain unknown.
METHODS: A mouse model of inflammation-accelerated KrasG12D-driven early pancreatic carcinogenesis was used. Rptor, Rictor and Arpc4 (Actin-related protein 2/3 complex subunit 4) were conditionally ablated in acinar cells to deactivate the function of mTORC1, mTORC2 and the Actin-related protein (Arp) 2/3 complex, respectively.
RESULTS: We found that mTORC1 and mTORC2 are markedly activated in human and mouse ADM lesions, and cooperate to promote KrasG12D-driven ADM in mice and in vitro. They utilize the Arp2/3 complex as a common downstream effector to induce the remodeling the actin cytoskeleton leading to ADM. In particular, mTORC1 regulates the translation of Rac1 (Rac family small GTPase 1) and the Arp2/3-complex subunit Arp3, whereas mTORC2 activates the Arp2/3 complex by promoting Akt/Rac1 signaling. Consistently, genetic ablation of the Arp2/3 complex prevents KrasG12D-driven ADM in vivo. In acinar cells, the Arp2/3 complex and its actin-nucleation activity mediated the formation of a basolateral actin cortex, which is indispensable for ADM and pre-neoplastic transformation.
CONCLUSION: Here, we show that mTORC1 and mTORC2 attain a dual, yet non-redundant regulatory role in ADM and early pancreatic carcinogenesis by promoting Arp2/3 complex function. Thus, the role of Arp2/3 complex as a common effector of mTORC1 and mTORC2 fills the gap between oncogenic signals and actin dynamics underlying PDAC initiation.

Keywords:  ADM; Arp2/3 complex; PDAC; Rictor; Rptor; mTOR

DOI:  https://doi.org/10.1053/j.gastro.2020.12.061
J Cell Biol. 2021 Feb 01. pii: e201902073. [Epub ahead of print]220(2):

Phosphorylation of PKCδ by FER tips the balance from EGFR degradation to recycling.

Ana Lonic, Freya Gehling, Leila Belle, Xiaochun Li, Nicole L Schieber, Elizabeth V Nguyen, Gregory J Goodall, Robert G Parton, Roger J Daly, Yeesim Khew-Goodall.

Receptor degradation terminates signaling by activated receptor tyrosine kinases. Degradation of EGFR occurs in lysosomes and requires the switching of RAB5 for RAB7 on late endosomes to enable their fusion with the lysosome, but what controls this critical switching is poorly understood. We show that the tyrosine kinase FER alters PKCδ function by phosphorylating it on Y374, and that phospho-Y374-PKCδ prevents RAB5 release from nascent late endosomes, thereby inhibiting EGFR degradation and promoting the recycling of endosomal EGFR to the cell surface. The rapid association of phospho-Y374-PKCδ with EGFR-containing endosomes is diminished by PTPN14, which dephosphorylates phospho-Y374-PKCδ. In triple-negative breast cancer cells, the FER-dependent phosphorylation of PKCδ enhances EGFR signaling and promotes anchorage-independent cell growth. Importantly, increased Y374-PKCδ phosphorylation correlating with arrested late endosome maturation was identified in ∼25% of triple-negative breast cancer patients, suggesting that dysregulation of this pathway may contribute to their pathology.

DOI: https://doi.org/10.1083/jcb.201902073
Mol Neurobiol. 2021 Jan 06.

Exosome Release Is Modulated by the Mitochondrial-Lysosomal Crosstalk in Parkinson's Disease Stress Conditions.

Fatema Currim, Jyoti Singh, Anjali Shinde, Dhruv Gohel, Milton Roy, Kritarth Singh, Shatakshi Shukla, Minal Mane, Hitesh Vasiyani, Rajesh Singh.

  Parkinson's disease (PD) is a neurodegenerative disorder characterized by the progressive loss of dopaminergic neurons in the substantia nigra (SN) pars compacta region of the brain. The main pathological hallmark involves cytoplasmic inclusions of α-synuclein and mitochondrial dysfunction, which is observed in other part of the central nervous system other than SN suggesting the spread of pathogenesis to bystander neurons. The inter-neuronal communication through exosomes may play an important role in the spread of the disease; however, the mechanisms are not well elucidated. Mitochondria and its role in inter-organellar crosstalk with multivesicular body (MVB) and lysosome and its role in modulation of exosome release in PD is not well understood. In the current study, we investigated the mitochondria-lysosome crosstalk modulating the exosome release in neuronal and glial cells. We observed that PD stress showed enhanced release of exosomes in dopaminergic neurons and glial cells. The PD stress condition in these cells showed fragmented network and mitochondrial dysfunction which further leads to functional deficit of lysosomes and hence inhibition of autophagy flux. Neuronal and glial cells treated with rapamycin showed enhanced autophagy and inhibited the exosomal release. The results here suggest that maintenance of mitochondrial function is important for the lysosomal function and hence exosomal release which is important for the pathogenesis of PD.

Keywords:  Exosome release; Mitochondria-lysosome crosstalk; Mitochondrial dysfunctions; Parkinson’s disease

DOI:  https://doi.org/10.1007/s12035-020-02243-3
Cell Death Differ. 2021 Jan 04.

Build-UPS and break-downs: metabolism impacts on proteostasis and aging.

Franziska Ottens, André Franz, Thorsten Hoppe.

Perturbation of metabolism elicits cellular stress which profoundly modulates the cellular proteome and thus protein homeostasis (proteostasis). Consequently, changes in the cellular proteome due to metabolic shift require adaptive mechanisms by molecular protein quality control. The mechanisms vitally controlling proteostasis embrace the entire life cycle of a protein involving translational control at the ribosome, chaperone-assisted native folding, and subcellular sorting as well as proteolysis by the proteasome or autophagy. While metabolic imbalance and proteostasis decline have been recognized as hallmarks of aging and age-associated diseases, both processes are largely considered independently. Here, we delineate how proteome stability is governed by insulin/IGF1 signaling (IIS), mechanistic target of Rapamycin (TOR), 5' adenosine monophosphate-activated protein kinase (AMPK), and NAD-dependent deacetylases (Sir2-like proteins known as sirtuins). This comprehensive overview is emphasizing the regulatory interconnection between central metabolic pathways and proteostasis, indicating the relevance of shared signaling nodes as targets for future therapeutic interventions.

DOI: https://doi.org/10.1038/s41418-020-00682-y
Neuropathol Appl Neurobiol. 2021 Jan 04.

Novel autophagic vacuolar myopathies: phenotype and genotype features.

Filomena Napolitano, Chiara Terracciano, Giorgia Bruno, Paolo De Blasiis, Luca Lombardi, Alessandro Gialluisi, Fernando Gianfrancesco, Donatella De Giovanni, Albina Tummolo, Giuseppe Di Iorio, Giuseppe Limongelli, Teresa Esposito, Mariarosa Anna Beatrice Melone, Simone Sampaolo.

   BACKGROUND: Autophagic Vacuolar Myopathies (AVMs) are an emerging group of heterogeneous myopathies sharing histopathological features on muscle pathology, in which autophagic vacuoles are the pathognomonic morphologic hallmarks. Glycogen storage disease type II (GSDII) caused by lysosomal acid α-glucosidase (GAA) deficiency, is the best-characterized AVM.
AIMS: This study aimed to investigate the mutational profiling of seven neuromuscular outpatients sharing clinical, myopathological and biochemical findings with AVMs.
METHODS: We applied a diagnostic protocol, recently published by our research group for suspected late onset GSDII (LO-GSDII), including counting PAS-positive lymphocytes on blood smears, dried blood spot (DBS)-GAA, muscle biopsy histological and immunofluorescence studies, GAA activity assay and expression studies on muscle homogenate, GAA sequencing, GAA multiplex ligation-dependent probe amplification (MLPA) and whole exome sequencing (WES).
RESULTS: The patients had a limb girdle-like muscular pattern with persistent hyperCKaemia; vacuolated PAS-positive lymphocytes, glycogen accumulation and impaired autophagy at muscle biopsy. Decreased GAA activity was also measured. While GAA sequencing identified no pathogenic mutations, WES approach allowed us to identify for each patient an unexpected mutational pattern in genes cooperating in lysosomal-autophagic machinery, some of which have never been linked to human diseases.
CONCLUSIONS: Our data suggest that reduced GAA activity may occur in any condition of impaired autophagy and that WES approach is advisable in all genetically undefined cases of autophagic myopathy. Therefore, deficiency of GAA activity and PAS-positive lymphocytes should be considered as AVM markers together with LC3/p62-positive autophagic vacuoles.

Keywords:  AVM condition; GAA activity; autophagic myopathies; complex genetic profiling; genetic heterogeneity; muscle biopsy; vacuolated PAS-positive lymphocytes; whole exome sequencing

DOI:  https://doi.org/10.1111/nan.12690
Metabolites. 2021 Jan 02. pii: E27. [Epub ahead of print]11(1):

The Harmonious Interplay of Amino Acid and Monocarboxylate Transporters Induces the Robustness of Cancer Cells.

Go J Yoshida.

  There is a growing body of evidence that metabolic reprogramming contributes to the acquisition and maintenance of robustness associated with malignancy. The fine regulation of expression levels of amino acid and monocarboxylate transporters enables cancer cells to exhibit the metabolic reprogramming that is responsible for therapeutic resistance. Amino acid transporters characterized by xCT (SLC7A11), ASCT2 (SLC1A5), and LAT1 (SLC7A5) function in the uptake and export of amino acids such as cystine and glutamine, thereby regulating glutathione synthesis, autophagy, and glutaminolysis. CD44 variant, a cancer stem-like cell marker, stabilizes the xCT antiporter at the cellular membrane, and tumor cells positive for xCT and/or ASCT2 are susceptible to sulfasalazine, a system Xc(-) inhibitor. Inhibiting the interaction between LAT1 and CD98 heavy chain prevents activation of the mammalian target of rapamycin (mTOR) complex 1 by glutamine and leucine. mTOR signaling regulated by LAT1 is a sensor of dynamic alterations in the nutrient tumor microenvironment. LAT1 is overexpressed in various malignancies and positively correlated with poor clinical outcome. Metabolic reprogramming of glutamine occurs often in cancer cells and manifests as ASCT2-mediated glutamine addiction. Monocarboxylate transporters (MCTs) mediate metabolic symbiosis, by which lactate in cancer cells under hypoxia is exported through MCT4 and imported by MCT1 in less hypoxic regions, where it is used as an oxidative metabolite. Differential expression patterns of transporters cause functional intratumoral heterogeneity leading to the therapeutic resistance. Therefore, metabolic reprogramming based on these transporters may be a promising therapeutic target. This review highlights the pathological function and therapeutic targets of transporters including xCT, ASCT2, LAT1, and MCT.

Keywords:  ASCT2 (SLC1A5); CD44 variant; LAT1 (SLC7A5); cancer stem-like cells; glutamine addiction; metabolic symbiosis; monocarboxylate transporter; redox stress; sulfasalazine; system Xc(-)

DOI:  https://doi.org/10.3390/metabo11010027
ACS Biomater Sci Eng. 2019 Jul 08. 5(7): 3398-3408

Nano CaCO3 "Lysosomal Bombs" Enhance Chemotherapy Drug Efficacy via Rebalancing Tumor Intracellular pH.

Chenxu Zhang, Shihui Li, Ao Yu, Yongjian Wang.

  Successful delivery of drugs to the target site is half the battle against tumors as intracellular alkalization pH (pHi) microenvironments severely restricted the efficacy of chemotherapy drugs delivered into tumor cells. Herein, a redox-selective pH-triggered "lysosomal bomb" (DSA/CC-DOX) is developed based on vaterite calcium carbonate and disulfide-cross-linked sodium alginate (DSA) with doxorubicin (DOX) encapsulated. Benefiting from the acid-triggered volume expansion of CaCO3, DSA/CC-DOX NPs can act like a "lysosomal bomb" that rapidly tears the lysosomal membrane with the release of acidic inclusions and the loaded DOX, and then the alkalized pHi in human liver tumor cells (HepG2) can be decreased from 7.61 to 7.09, thus promoting the intracellular accumulation of DOX nearly 3 times more than the free drug. In addition, facilitated by the responsive break of the disulfide bond to GSH, the release of DOX in HepG2 is nearly 8 times that of human normal liver cell (LO2). Notably, DSA/CC-DOX treatment increased the tumor inhibition rate of free drug by 16% and effectively reduced the cardiotoxicity of DOX in the mouse H22 liver cancer model. Overall, acidifying the tumor intracellular environment is a prospective way to improve the antitumor capacity of chemotherapy drug.

Keywords:  acidify pHi; drug accumulation; lysosomes disruption; tumor selective; vaterite calcium carbonate

DOI:  https://doi.org/10.1021/acsbiomaterials.9b00436
EBioMedicine. 2021 Jan 05. pii: S2352-3964(20)30583-1. [Epub ahead of print]63 103207

Transcription factor EB regulates cardiovascular homeostasis.

Haocheng Lu, Jinjian Sun, Milton H Hamblin, Y Eugene Chen, Yanbo Fan.

  Cardiovascular diseases (CVDs) are the leading cause of death and a major cause of disability globally. Transcription factor EB (TFEB), as a member of the microphthalmia transcription factor (MITF) family, has been demonstrated to be a master regulator of autophagy and lysosomal biogenesis. Emerging studies suggest that TFEB regulates homeostasis in the cardiovascular system and shows beneficial effects on CVDs, including atherosclerosis, aortic aneurysm, postischemic angiogenesis, and cardiotoxicity, constituting a promising molecular target for the prevention and treatment of these diseases. Post-translational modifications regulate TFEB nuclear translocation and its transcriptional activity. Therapeutic strategies have been pursued to enhance TFEB activity and facilitate TFEB beneficial effects on CVDs. The elucidation of TFEB function and the precise underlying mechanisms will accelerate drug development and potential applications of TFEB drugs in the treatment of human diseases.

Keywords:  Autophagy; Cardiovascular disease; Drug development; Lysosome; Post-translational modification

DOI:  https://doi.org/10.1016/j.ebiom.2020.103207
Circ Res. 2021 Jan 06.

MTORC1-Regulated Metabolism Controlled by TSC2 Limits Cardiac Reperfusion Injury.

Christian U Oeing, Seungho Jun, Sumita Mishra, Brittany Dunkerly-Eyring, Anna Chen, Maria I Grajeda, Usman Tahir, Robert E Gerszten, Nazareno Paolocci, Mark J Ranek, David A Kass.

  Rationale: The mechanistic target of rapamycin complex-1 (mTORC1) controls metabolism and protein homeostasis, and is activated following ischemic reperfusion (IR) injury and by ischemic preconditioning (IPC). However, studies vary as to whether this activation is beneficial or detrimental, and its influence on metabolism after IR is little studied. A limitation of prior investigations is their use of broad gain/loss of mTORC1 function, mostly applied prior to ischemic stress. This can be circumvented by regulating one serine (S1365) on tuberous sclerosis complex (TSC2) to achieve bi-directional mTORC1 modulation but only with TCS2-regulated co-stimulation. Objective: We tested the hypothesis that reduced TSC2 S1365 phosphorylation protects the myocardium against IR and IPC by amplifying mTORC1 activity to favor glycolytic metabolism. Methods and Results: Mice with either S1365A (TSC2SA; phospho-null) or S1365E (TSC2SE; phosphomimetic) knock-in mutations were studied ex vivo and in vivo. In response to IR, hearts from TSC2SA mice had amplified mTORC1 activation and improved heart function compared to WT and TSC2SE hearts. The magnitude of protection matched IPC. IPC requited less S1365 phosphorylation, as TSC2SE hearts gained no benefit and failed to activate mTORC1 with IPC. IR metabolism was altered in TSC2SA, with increased mitochondrial oxygen consumption rate and glycolytic capacity (stressed/maximal extracellular acidification) after myocyte hypoxia-reperfusion. In whole heart, lactate increased and long-chain acyl-carnitine levels declined during ischemia. The relative IR protection in TSC2SA was lost by lowering glucose in the perfusate by 36%. Adding fatty acid (palmitate) compensated for reduced glucose in WT and TSC2SE but not TSC2SA which had the worst post-IR function under these conditions. Conclusions: TSC2-S1365 phosphorylation status regulates myocardial substrate utilization, and its decline activates mTORC1 biasing metabolism away from fatty acid oxidation to glycolysis to confer protection against IR. This pathway is also engaged and reduced TSC2 S1365 phosphorylation required for effective IPC.

Keywords:  mechanistic target of rapamycin; tuberous sclerosis complex

DOI:  https://doi.org/10.1161/CIRCRESAHA.120.317710
Metabolites. 2020 Dec 30. pii: E18. [Epub ahead of print]11(1):

Decrease in Myelin-Associated Lipids Precedes Neuronal Loss and Glial Activation in the CNS of the Sandhoff Mouse as Determined by Metabolomics.

Emmanuelle Lecommandeur, Maria Begoña Cachón-González, Susannah Boddie, Ben D McNally, Andrew W Nicholls, Timothy M Cox, Julian L Griffin.

  Sandhoff disease (SD) is a lysosomal disease caused by mutations in the gene coding for the β subunit of β-hexosaminidase, leading to deficiency in the enzymes β-hexosaminidase (HEX) A and B. SD is characterised by an accumulation of gangliosides and related glycolipids, mainly in the central nervous system, and progressive neurodegeneration. The underlying cellular mechanisms leading to neurodegeneration and the contribution of inflammation in SD remain undefined. The aim of the present study was to measure global changes in metabolism over time that might reveal novel molecular pathways of disease. We used liquid chromatography-mass spectrometry and 1H Nuclear Magnetic Resonance spectroscopy to profile intact lipids and aqueous metabolites, respectively. We examined spinal cord and cerebrum from healthy and Hexb-/- mice, a mouse model of SD, at ages one, two, three and four months. We report decreased concentrations in lipids typical of the myelin sheath, galactosylceramides and plasmalogen-phosphatidylethanolamines, suggesting that reduced synthesis of myelin lipids is an early event in the development of disease pathology. Reduction in neuronal density is progressive, as demonstrated by decreased concentrations of N-acetylaspartate and amino acid neurotransmitters. Finally, microglial activation, indicated by increased amounts of myo-inositol correlates closely with the late symptomatic phases of the disease.

Keywords:  bis(monoacylglycero)phosphates; galactosylceramides; lipidomics; lysosomal disorders; metabolomics; plasmalogens; β-hexosaminidase

DOI:  https://doi.org/10.3390/metabo11010018
Front Physiol. 2020 ;11 619730

AMPK/mTOR Signaling in Autophagy Regulation During Cisplatin-Induced Acute Kidney Injury.

Ying Wang, Zhiwen Liu, Shaoqun Shu, Juan Cai, Chengyuan Tang, Zheng Dong.

  Autophagy is a conserved, multistep pathway that degrades and recycles dysfunctional organelles and macromolecules to maintain cellular homeostasis. Mammalian target of rapamycin (mTOR) and adenosine-monophosphate activated-protein kinase (AMPK) are major negative and positive regulators of autophagy, respectively. In cisplatin-induced acute kidney injury (AKI) or nephrotoxicity, autophagy is rapidly induced in renal tubular epithelial cells and acts as a cytoprotective mechanism for cell survival. Both mTOR and AMPK have been implicated in the regulation of autophagy in cisplatin-induced AKI. Targeting mTOR and/or AMPK may offer effective strategies for kidney protection during cisplatin-mediated chemotherapy.

Keywords:  AMPK; acute kidney injury; autophagy; cisplatin; mTOR; nephrotoxicity

DOI:  https://doi.org/10.3389/fphys.2020.619730
Aging (Albany NY). 2021 Jan 07. 12

Function of Deptor and its roles in hematological malignancies.

Mario Morales-Martinez, Alan Lichtenstein, Mario I Vega.

  Deptor is a protein that interacts with mTOR and that belongs to the mTORC1 and mTORC2 complexes. Deptor is capable of inhibiting the kinase activity of mTOR. It is well known that the mTOR pathway is involved in various signaling pathways that are involved with various biological processes such as cell growth, apoptosis, autophagy, and the ER stress response. Therefore, Deptor, being a natural inhibitor of mTOR, has become very important in its study. Because of this, it is important to research its role regarding the development and progression of human malignancies, especially in hematologic malignancies. Due to its variation in expression in cancer, it has been suggested that Deptor can act as an oncogene or tumor suppressor depending on the cellular or tissue context. This review discusses recent advances in its transcriptional and post-transcriptional regulation of Deptor. As well as the advances regarding the activities of Deptor in hematological malignancies, its possible role as a biomarker, and its possible clinical relevance in these malignancies.

Keywords:  Deptor; Multiple Myeloma; Non-Hodgkin Lymphoma; hematological malignances; leukemia

DOI:  https://doi.org/10.18632/aging.202462
Cell Metab. 2021 Jan 05. pii: S1550-4131(20)30664-1. [Epub ahead of print]33(1): 9-20

Harnessing the Co-vulnerabilities of Amino Acid-Restricted Cancers.

Gaurav Pathria, Ze'ev A Ronai.

  Sustained proliferative potential of cancer cells creates heightened energetic and biosynthetic demands. The resulting overt dependence of cancer cells on unperturbed nutrient supply has prompted a widespread interest in amino acid restriction strategies as potential cancer therapeutics. However, owing to rapid signaling and metabolic reprogramming in cancer cells, the prospects for success of amino acid restriction approaches remain unclear. We thus recognize that the identification of co-vulnerabilities of amino acid-restricted cancers may inform actionable targets for effective combined interventions. In this perspective, we outline the current state of key cellular mechanisms underlying adaptation to amino acid restriction and discuss the role of signal transduction pathways governing cancer cell resistance to amino acid restriction, with potential ramifications for the design of future therapeutic efforts.

Keywords:  ATF4; MAPK; NRF2; adaptation; amino acids; c-MYC; cancer; mTORC1; metabolism; resistance

DOI:  https://doi.org/10.1016/j.cmet.2020.12.009
Cancers (Basel). 2021 Jan 02. pii: E125. [Epub ahead of print]13(1):

Amino Acid Transporters on the Guard of Cell Genome and Epigenome.

Uğur Kahya, Ayşe Sedef Köseer, Anna Dubrovska.

  Tumorigenesis is driven by metabolic reprogramming. Oncogenic mutations and epigenetic alterations that cause metabolic rewiring may also upregulate the reactive oxygen species (ROS). Precise regulation of the intracellular ROS levels is critical for tumor cell growth and survival. High ROS production leads to the damage of vital macromolecules, such as DNA, proteins, and lipids, causing genomic instability and further tumor evolution. One of the hallmarks of cancer metabolism is deregulated amino acid uptake. In fast-growing tumors, amino acids are not only the source of energy and building intermediates but also critical regulators of redox homeostasis. Amino acid uptake regulates the intracellular glutathione (GSH) levels, endoplasmic reticulum stress, unfolded protein response signaling, mTOR-mediated antioxidant defense, and epigenetic adaptations of tumor cells to oxidative stress. This review summarizes the role of amino acid transporters as the defender of tumor antioxidant system and genome integrity and discusses them as promising therapeutic targets and tumor imaging tools.

Keywords:  Enhancer of zeste homolog 2 (EZH2); SLC1A5/ASCT2; SLC7A11/xCT; SLC7A5/LAT1; amino acid transporters; epigenetic regulation; glutathione (GSH); oxidative stress; reactive oxygen species (ROS); α-ketoglutarate (αKG)

DOI:  https://doi.org/10.3390/cancers13010125
Nat Genet. 2021 Jan;53(1): 16-26

The amino acid transporter SLC7A5 is required for efficient growth of KRAS-mutant colorectal cancer.

CRUK Rosetta Grand Challenge Consortium

Oncogenic KRAS mutations and inactivation of the APC tumor suppressor co-occur in colorectal cancer (CRC). Despite efforts to target mutant KRAS directly, most therapeutic approaches focus on downstream pathways, albeit with limited efficacy. Moreover, mutant KRAS alters the basal metabolism of cancer cells, increasing glutamine utilization to support proliferation. We show that concomitant mutation of Apc and Kras in the mouse intestinal epithelium profoundly rewires metabolism, increasing glutamine consumption. Furthermore, SLC7A5, a glutamine antiporter, is critical for colorectal tumorigenesis in models of both early- and late-stage metastatic disease. Mechanistically, SLC7A5 maintains intracellular amino acid levels following KRAS activation through transcriptional and metabolic reprogramming. This supports the increased demand for bulk protein synthesis that underpins the enhanced proliferation of KRAS-mutant cells. Moreover, targeting protein synthesis, via inhibition of the mTORC1 regulator, together with Slc7a5 deletion abrogates the growth of established Kras-mutant tumors. Together, these data suggest SLC7A5 as an attractive target for therapy-resistant KRAS-mutant CRC.

DOI: https://doi.org/10.1038/s41588-020-00753-3
Nat Genet. 2021 Jan;53(1): 54-64

A cross-platform approach identifies genetic regulators of human metabolism and health.

MacTel Consortium

In cross-platform analyses of 174 metabolites, we identify 499 associations (P < 4.9 × 10-10) characterized by pleiotropy, allelic heterogeneity, large and nonlinear effects and enrichment for nonsynonymous variation. We identify a signal at GLP2R (p.Asp470Asn) shared among higher citrulline levels, body mass index, fasting glucose-dependent insulinotropic peptide and type 2 diabetes, with β-arrestin signaling as the underlying mechanism. Genetically higher serine levels are shown to reduce the likelihood (by 95%) and predict development of macular telangiectasia type 2, a rare degenerative retinal disease. Integration of genomic and small molecule data across platforms enables the discovery of regulators of human metabolism and translation into clinical insights.

DOI: https://doi.org/10.1038/s41588-020-00751-5
Cell Signal. 2021 Jan 01. pii: S0898-6568(20)30390-9. [Epub ahead of print]80 109912

mTORC2 deploys the mRNA binding protein IGF2BP1 to regulate c-MYC expression and promote cell survival.

Andromachi Lambrianidou, Evangelia Sereti, Katerina Soupsana, Chrysoula Komini, Konstantinos Dimas, Theoni Trangas.

  mTORC2 promotes cell survival by phosphorylating AKT and enhancing its activity. Inactivation of mTORC2 reduces viability through down-regulation of E2F1 caused by up-regulation of c-MYC. An additional target of mTORC2 is IGF2BP1, an oncofetal RNA binding protein expressed de novo in a wide array of malignancies. IGF2BP1 enhances c-MYC expression by protecting the coding region instability sequence (CRD) of its mRNA from endonucleolytic cleavage. Here we show that repression of mTORC2 signalling and prevention of Ser181 phosphorylation of IGF2BP1 enhanced translation and destabilization of the endogenous c-myc mRNA as well as the mRNA of reporter transcripts carrying the CRD sequence in frame. The consequent increase in c-MYC protein was accompanied by the emergence of an apoptotic c-MYC overexpressing population. On the other hand, preventing phosphorylation of IGF2BP1 on Tyr396 by Src kinase caused the accumulation of translationally silent transcripts through sequestration by IGF2BP1 into cytoplasmic granules. The apoptotic effect of mTORC2 signalling deprivation was augmented when preceded by inhibition of IGF2BP1 phosphorylation by the Src kinase in concert with further increase of c-MYC levels because of enhanced translation of the previously stored mRNA only in the presence of IGF2BP1. Furthermore, the combined administration of mTORC2 and Src inhibitors exhibited synergism in delaying xenograft growth in female NOD.CB17-Prkdcscid/J mice. The above in vitro and in vivo findings may be applied for the induction of targeted apoptosis of cells expressing de novo the oncofetal protein IGF2BP1, a feature of aggressive malignancies resulting in a more focused anticancer therapeutic approach.

Keywords:  Apoptosis; C-MYC; IGF2BP1; Src kinase; mTORC2 kinase

DOI:  https://doi.org/10.1016/j.cellsig.2020.109912
Nat Metab. 2021 Jan 04.

Metabolites and the tumour microenvironment: from cellular mechanisms to systemic metabolism.

Ilaria Elia, Marcia C Haigis.

Metabolic transformation is a hallmark of cancer and a critical target for cancer therapy. Cancer metabolism and behaviour are regulated by cell-intrinsic factors as well as metabolite availability in the tumour microenvironment (TME). This metabolic niche within the TME is shaped by four tiers of regulation: (1) intrinsic tumour cell metabolism, (2) interactions between cancer cells and non-cancerous cells, (3) tumour location and heterogeneity and (4) whole-body metabolic homeostasis. Here, we define these modes of metabolic regulation and review how distinct cell types contribute to the metabolite composition of the TME. Finally, we connect these insights to understand how each of these tiers offers unique therapeutic potential to modulate the metabolic profile and function of all cells inhabiting the TME.

DOI: https://doi.org/10.1038/s42255-020-00317-z
PLoS Negl Trop Dis. 2021 Jan;15(1): e0008884

Extracellular vesicles released from the filarial parasite Brugia malayi downregulate the host mTOR pathway.

Alessandra Ricciardi, Sasisekhar Bennuru, Sameha Tariq, Sukhbir Kaur, Weiwei Wu, Abdel G Elkahloun, Anush Arakelyan, Jahangheer Shaik, David W Dorward, Thomas B Nutman, Roshanak Tolouei Semnani.

We have previously shown that the microfilarial (mf) stage of Brugia malayi can inhibit the mammalian target of rapamycin (mTOR; a conserved serine/threonine kinase critical for immune regulation and cellular growth) in human dendritic cells (DC) and we have proposed that this mTOR inhibition is associated with the DC dysfunction seen in filarial infections. Extracellular vesicles (EVs) contain many proteins and nucleic acids including microRNAs (miRNAs) that might affect a variety of intracellular pathways. Thus, EVs secreted from mf may elucidate the mechanism by which the parasite is able to modulate the host immune response during infection. EVs, purified from mf of Brugia malayi and confirmed by size through nanoparticle tracking analysis, were assessed by miRNA microarrays (accession number GSE157226) and shown to be enriched (>2-fold, p-value<0.05, FDR = 0.05) for miR100, miR71, miR34, and miR7. The microarray analysis compared mf-derived EVs and mf supernatant. After confirming their presence in EVs using qPCR for these miRNA targets, web-based target predictions (using MIRPathv3, TarBAse and MicroT-CD) predicted that miR100 targeted mTOR and its downstream regulatory protein 4E-BP1. Our previous data with live parasites demonstrated that mf downregulate the phosphorylation of mTOR and its downstream effectors. Additionally, our proteomic analysis of the mf-derived EVs revealed the presence of proteins commonly found in these vesicles (data are available via ProteomeXchange with identifier PXD021844). We confirmed internalization of mf-derived EVs by human DCs and monocytes using confocal microscopy and flow cytometry, and further demonstrated through flow cytometry, that mf-derived EVs downregulate the phosphorylation of mTOR in human monocytes (THP-1 cells) to the same degree that rapamycin (a known mTOR inhibitor) does. Our data collectively suggest that mf release EVs that interact with host cells, such as DC, to modulate host responses.

DOI: https://doi.org/10.1371/journal.pntd.0008884
Theranostics. 2021 ;11(1): 222-256

Selective autophagy of intracellular organelles: recent research advances.

Wen Li, Pengcheng He, Yuge Huang, Yi-Fang Li, Jiahong Lu, Min Li, Hiroshi Kurihara, Zhuo Luo, Tian Meng, Mashun Onishi, Changle Ma, Lei Jiang, Yongquan Hu, Qing Gong, Dongxing Zhu, Yiming Xu, Rong Liu, Lei Liu, Cong Yi, Yushan Zhu, Ningfang Ma, Koji Okamoto, Zhiping Xie, Jinbao Liu, Rong-Rong He, Du Feng.

  Macroautophagy (hereafter called autophagy) is a highly conserved physiological process that degrades over-abundant or damaged organelles, large protein aggregates and invading pathogens via the lysosomal system (the vacuole in plants and yeast). Autophagy is generally induced by stress, such as oxygen-, energy- or amino acid-deprivation, irradiation, drugs, etc. In addition to non-selective bulk degradation, autophagy also occurs in a selective manner, recycling specific organelles, such as mitochondria, peroxisomes, ribosomes, endoplasmic reticulum (ER), lysosomes, nuclei, proteasomes and lipid droplets (LDs). This capability makes selective autophagy a major process in maintaining cellular homeostasis. The dysfunction of selective autophagy is implicated in neurodegenerative diseases (NDDs), tumorigenesis, metabolic disorders, heart failure, etc. Considering the importance of selective autophagy in cell biology, we systemically review the recent advances in our understanding of this process and its regulatory mechanisms. We emphasize the 'cargo-ligand-receptor' model in selective autophagy for specific organelles or cellular components in yeast and mammals, with a focus on mitophagy and ER-phagy, which are finely described as types of selective autophagy. Additionally, we highlight unanswered questions in the field, helping readers focus on the research blind spots that need to be broken.

Keywords:  ER-phagy; autophagy receptor; lipophagy; lysophagy; mitophagy; nucleophagy; pexophagy; proteaphagy; ribophagy; selective autophagy

DOI:  https://doi.org/10.7150/thno.49860
PLoS Biol. 2021 Jan 04. 19(1): e3001029

Cyclin-dependent Kinase 1 and Aurora Kinase choreograph mitotic storage and redistribution of a growth factor receptor.

Christina D Cota, Matthew S Dreier, William Colgan, Anna Cha, Twan Sia, Brad Davidson.

Endosomal trafficking of receptors and associated proteins plays a critical role in signal processing. Until recently, it was thought that trafficking was shut down during cell division. Thus, remarkably, the regulation of trafficking during division remains poorly characterized. Here we delineate the role of mitotic kinases in receptor trafficking during asymmetric division. Targeted perturbations reveal that Cyclin-dependent Kinase 1 (CDK1) and Aurora Kinase promote storage of Fibroblast Growth Factor Receptors (FGFRs) by suppressing endosomal degradation and recycling pathways. As cells progress through metaphase, loss of CDK1 activity permits differential degradation and targeted recycling of stored receptors, leading to asymmetric induction. Mitotic receptor storage, as delineated in this study, may facilitate rapid reestablishment of signaling competence in nascent daughter cells. However, mutations that limit or enhance the release of stored signaling components could alter daughter cell fate or behavior thereby promoting oncogenesis.

DOI: https://doi.org/10.1371/journal.pbio.3001029
Front Cell Neurosci. 2020 ;14 602116

Dysfunctional Autophagy and Endolysosomal System in Neurodegenerative Diseases: Relevance and Therapeutic Options.

Silvia Giovedì, Margherita Maria Ravanelli, Barbara Parisi, Barbara Bettegazzi, Fabrizia Claudia Guarnieri.

  Autophagy and endolysosomal trafficking are crucial in neuronal development, function and survival. These processes ensure efficient removal of misfolded aggregation-prone proteins and damaged organelles, such as dysfunctional mitochondria, thus allowing the maintenance of proper cellular homeostasis. Beside this, emerging evidence has pointed to their involvement in the regulation of the synaptic proteome needed to guarantee an efficient neurotransmitter release and synaptic plasticity. Along this line, an intimate interplay between the molecular machinery regulating synaptic vesicle endocytosis and synaptic autophagy is emerging, suggesting that synaptic quality control mechanisms need to be tightly coupled to neurosecretion to secure release accuracy. Defects in autophagy and endolysosomal pathway have been associated with neuronal dysfunction and extensively reported in Alzheimer's, Parkinson's, Huntington's and amyotrophic lateral sclerosis among other neurodegenerative diseases, with common features and emerging genetic bases. In this review, we focus on the multiple roles of autophagy and endolysosomal system in neuronal homeostasis and highlight how their defects probably contribute to synaptic default and neurodegeneration in the above-mentioned diseases, discussing the most recent options explored for therapeutic interventions.

Keywords:  autophagy; endocytosis; lysosomes; neurodegenerative diseases; synapse

DOI:  https://doi.org/10.3389/fncel.2020.602116
Nat Metab. 2021 Jan 04.

Mitochondrial long non-coding RNA GAS5 tunes TCA metabolism in response to nutrient stress.

Lingjie Sang, Huai-Qiang Ju, Zuozhen Yang, Qiwei Ge, Zhen Zhang, Fangzhou Liu, Luojia Yang, Hangdi Gong, Chengyu Shi, Lei Qu, Hui Chen, Minjie Wu, Hao Chen, Ruihua Li, Qianqian Zhuang, Hailong Piao, Qingfeng Yan, Weishi Yu, Liangjing Wang, Jianzhong Shao, Jian Liu, Wenqi Wang, Tianhua Zhou, Aifu Lin.

Organelles use specialized molecules to regulate their essential cellular processes. However, systematically elucidating the subcellular distribution and function of molecules such as long non-coding RNAs (lncRNAs) in cellular homeostasis and diseases has not been fully achieved. Here, we reveal the diverse and abundant subcellular distribution of organelle-associated lncRNAs from mitochondria, lysosomes and endoplasmic reticulum. Among them, we identify the mitochondrially localized lncRNA growth-arrest-specific 5 (GAS5) as a tumour suppressor in maintaining cellular energy homeostasis. Mechanistically, energy-stress-induced GAS5 modulates mitochondrial tricarboxylic acid flux by disrupting metabolic enzyme tandem association of fumarate hydratase, malate dehydrogenase and citrate synthase, the canonical members of the tricarboxylic acid cycle. GAS5 negatively correlates with levels of its associated mitochondrial metabolic enzymes in tumours and benefits overall survival in individuals with breast cancer. Together, our detailed annotation of subcellular lncRNA distribution identifies a functional role for lncRNAs in regulating cellular metabolic homeostasis, highlighting organelle-associated lncRNAs as potential clinical targets to manipulate cellular metabolism and diseases.

DOI: https://doi.org/10.1038/s42255-020-00325-z