A Gene Network Regulating Lysosomal Biogenesis and Function

Telethon Institute of Genetics and Medicine, Via P. Castellino 111, 80131 Naples, Italy.
Science (Impact Factor: 33.61). 07/2009; 325(5939):473-7. DOI: 10.1126/science.1174447
Source: PubMed


Lysosomes are organelles central to degradation and recycling processes in animal cells. Whether lysosomal activity is coordinated
to respond to cellular needs remains unclear. We found that most lysosomal genes exhibit coordinated transcriptional behavior
and are regulated by the transcription factor EB (TFEB). Under aberrant lysosomal storage conditions, TFEB translocated from
the cytoplasm to the nucleus, resulting in the activation of its target genes. TFEB overexpression in cultured cells induced
lysosomal biogenesis and increased the degradation of complex molecules, such as glycosaminoglycans and the pathogenic protein
that causes Huntington’s disease. Thus, a genetic program controls lysosomal biogenesis and function, providing a potential
therapeutic target to enhance cellular clearing in lysosomal storage disorders and neurodegenerative diseases.


Available from: Michela Palmieri
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    • "This mechanism is suggested by the upregulation of the lysosomal glycoproteins LAMP1 and LAMP2 in cells treated with some cationic drugs, imatinib or chloroquine (Ertmer et al., 2007; Chen, Gombart & Chen, 2011). Ultimately, the process may be controlled by the lysosomal genesis transcription factor TFEB that is subjected to nuclear translocation upon activation (Sardiello et al., 2009) and such a protracted response to basic drugs has been recently reported in cultured cells (chloroquine: Roczniak-Fergusson et al., 2012; propranolol: Logan, Kong & Krise, 2014). (4) Intriguing tissue or cell type specificities were described for the formation of cationic drug reservoirs. "
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    ABSTRACT: The proton pump vacuolar (V)-ATPase is the driving force that mediates the concentration of cationic drugs (weak bases) in the late endosome-lysosome continuum; secondary cell reactions include the protracted transformation of enlarged vacuoles into autophagosomes. We used the inherently fluorescent tertiary amine quinacrine in murine models to further assess the accumulation and signaling associated with cation trapping. Primary fibroblasts concentrate quinacrine ∼5,000-fold from their culture medium (K M 9.8 µM; transport studies). The drug is present in perinuclear granules that are mostly positive for Rab7 and LAMP1 (microscopy). Both drug uptake and retention are extensively inhibited by treatments with the V-ATPase inhibitor bafilomycin A1. The H + ionophore monensin also prevented quinacrine concentration by fibroblasts. However, inhibition of plasma membrane transporters or of the autophagic process with spautin-1 did not alter quinacrine transport parameters. Ancillary experiments did not support that low micromolar concentrations of quinacrine are substrates for organic cation transporters-1 to-3 or P-glycoprotein. The secondary autophagy induced by quinacrine in cells may derive from the accumulation of incompetent autophagolysosomes, as judged from the accumulation of p62/SQSTM1 and LC3 II (immunoblots). Accordingly, protracted lysosomogenesis is evidenced by increased expression of LAMP1 and LAMP2 in quinacrine-treated fibroblasts (48 h, immunoblots), a response that follows the nuclear translocation of the lysosomal genesis transcription factor TFEB and upreg-ulation of LAMP1 and −2 mRNAs (24 h). Quinacrine administration to live mice evidenced variable distribution to various organs and heterogeneous accumulation within the lung (stereo-microscopy, extraction). Dose-dependent in vivo autophagic and lysosomal accumulation was observed in the lung (immunoblots). No evidence has been found for transport or extrusion mechanisms modulating the cellular uptake of micromolar quinacrine at the plasma membrane level. As shown in vitro and in vivo, V-ATPase-mediated cation sequestration is associated, above a certain threshold, to autophagic flux inhibition and feedback lysosomogenesis. How to cite this article Parks et al. (2015), Autophagic flux inhibition and lysosomogenesis ensuing cellular capture and retention of the cationic drug quinacrine in murine models. PeerJ 3:e1314
    PeerJ 10/2015; 3(suppl 2):e1314. DOI:10.7717/peerj.1314 · 2.11 Impact Factor
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    • "Our findings demonstrate that mitochondrial dysfunction results in autophagosome-lysosome fusion defects and reduced mitochondrial clearance by mitophagy, which are commonly observed in almost all sphingolipidoses (Lieberman et al., 2012). A predominant nuclear localization of TFEB has been detected in cells from mouse models of LSD (Sardiello et al., 2009). Mitochondrial respiration deficiency increases TFEB activation, suggesting that the TFEB pathway is activated as a cellular response to lysosomal stress after intralysosomal storage of nondegraded molecules. "
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    ABSTRACT: The endolysosomal system is critical for the maintenance of cellular homeostasis. However, how endolysosomal compartment is regulated by mitochondrial function is largely unknown. We have generated a mouse model with defective mitochondrial function in CD4(+) T lymphocytes by genetic deletion of the mitochondrial transcription factor A (Tfam). Mitochondrial respiration deficiency impairs lysosome function, promotes p62 and sphingomyelin accumulation, and disrupts endolysosomal trafficking pathways and autophagy, thus linking a primary mitochondrial dysfunction to a lysosomal storage disorder. The impaired lysosome function in Tfam-deficient cells subverts T cell differentiation toward proinflammatory subsets and exacerbates the in vivo inflammatory response. Restoration of NAD(+) levels improves lysosome function and corrects the inflammatory defects in Tfam-deficient T cells. Our results uncover a mechanism by which mitochondria regulate lysosome function to preserve T cell differentiation and effector functions, and identify strategies for intervention in mitochondrial-related diseases. Copyright © 2015 Elsevier Inc. All rights reserved.
    Cell metabolism 08/2015; 22(3). DOI:10.1016/j.cmet.2015.07.020 · 17.57 Impact Factor
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    • "The most abundant of these proteins are the lysosomal membrane associated proteins LAMP1 and LAMP2 that, among other functions, are important for the auto(phago)-lysosomal fusion during the autophagic process [4]. Lysosomal biogenesis is transcriptionally regulated by Transcription factor EB (TFEB) that regulates expression of multiple genes encoding lysosomal enzymes and membrane proteins [5] [6] [7]. "
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    ABSTRACT: We investigated the role of lysosomal biogenesis and hydrolase activity in the clinical behavior and postoperative outcome of lung cancer. Using immunohistochemistry we investigated the expression of the transcription factor EB (TFEB) which orchestrates lysosomal biogenesis, the lysosome membrane protein LAMP2a and of the lysosomal hydrolase cathepsin D in a series of 98 non-small cell lung carcinomas (NSCLC) treated with surgery alone. In vitro experiments with the A549 and H1299 lung cancer cell lines were also performed. Overexpression of TFEB, LAMP2a and Cathepsin D was noted in 47/98 (47.9%), 43/98 (43.9%) and 39/98 (39.8%) cases, respectively, and were significantly correlated with each other and with adenocarcinomas. High LAMP2a was related to high histology grade. Linear regression analysis confirmed significant association of TFEB with BNIP3 (p=0.0003, r=0.35) and LC3A with LAMP2a expression (p=0.0002, r=0.37). An inverse association of Cathepsin D expression with stone-like structures (SLS) was recorded (p=0.02, r=0.22). On univariate analysis all three lyososomal variables were associated with poor prognosis (p=0.05, 0.04 and 0.01, for TFEB, Cathepsin D and LAMP2a, respectively). Multivariate analysis showed that the SLS number (p=0.0001, HR5.37), Cathepsin D expression (p=0.01, HR=2.2) and stage (p=0.01, HR=1.5) were independent prognostic variables. Silencing of TFEB with siRNAs in the A549 and H1299 lung cancer cell lines did not affect proliferation but resulted in reduced migration ability. Lysosomal biogenesis is linked to autophagosomal protein expression in NSCLC and characterizes subgroups of high risk patients after complete surgical lung tumor resection. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.
    Lung cancer (Amsterdam, Netherlands) 07/2015; 90(1). DOI:10.1016/j.lungcan.2015.07.008 · 3.96 Impact Factor
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