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1.
Cell ; 181(3): 748-748.e1, 2020 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-32359442

RESUMO

In addition to their well-defined recycling function, lysosomes act as metabolic signaling hubs that adjust cellular metabolism according to the availability of nutrients and growth factors by regulating metabolic kinases and transcription factors on their surface. Moreover, lysosomal hydrolases and ions released to cytosol or extracellular space have recently emerged as important regulators of various cellular processes from cell death to cell division. To view this SnapShot, open or download the PDF.


Assuntos
Lisossomos/metabolismo , Lisossomos/fisiologia , Autofagia/fisiologia , Citosol/metabolismo , Espaço Extracelular/metabolismo , Humanos , Hidrolases/metabolismo , Fosfotransferases/metabolismo , Transdução de Sinais , Fatores de Transcrição/metabolismo
2.
EMBO J ; 42(9): e111241, 2023 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-36970883

RESUMO

The accumulation of senescent cells is recognised as a driver of tissue and organismal ageing. One of the gold-standard hallmarks of a senescent cell is an increase in lysosomal content, as measured by senescence-associated ß-galactosidase (Senß-Gal) activity. The lysosome plays a central role in integrating mitogenic and stress cues to control cell metabolism, which is known to be dysregulated in senescence. Despite this, little is known about the cause and consequence of lysosomal biogenesis in senescence. We find here that lysosomes in senescent cells are dysfunctional; they have higher pH, increased evidence of membrane damage and reduced proteolytic capacity. The significant increase in lysosomal content is however sufficient to maintain degradative capacity of the cell to a level comparable to proliferating control cells. We demonstrate that increased nuclear TFEB/TFE3 supports lysosome biogenesis, is a hallmark of multiple forms of senescence and is required for senescent cell survival. TFEB/TFE3 are hypo-phosphorylated and show constitutive nuclear localisation in senescence. Evidence suggests that several pathways may contribute to TFEB/TFE3 dysregulation in senescence.


Assuntos
Núcleo Celular , Lisossomos , Autofagia , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Núcleo Celular/metabolismo , Proliferação de Células , Lisossomos/metabolismo
4.
EMBO J ; 41(24): e112677, 2022 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-36408828

RESUMO

Lysosome integrity is essential for cell viability, and lesions in lysosome membranes are repaired by the ESCRT machinery. Here, we describe an additional mechanism for lysosome repair that is activated independently of ESCRT recruitment. Lipidomic analyses showed increases in lysosomal phosphatidylserine and cholesterol after damage. Electron microscopy demonstrated that lysosomal membrane damage is rapidly followed by the formation of contacts with the endoplasmic reticulum (ER), which depends on the ER proteins VAPA/B. The cholesterol-binding protein ORP1L was recruited to damaged lysosomes, accompanied by cholesterol accumulation by a mechanism that required VAP-ORP1L interactions. The PtdIns 4-kinase PI4K2A rapidly produced PtdIns4P on lysosomes upon damage, and knockout of PI4K2A inhibited damage-induced accumulation of ORP1L and cholesterol and led to the failure of lysosomal membrane repair. The cholesterol-PtdIns4P transporter OSBP was also recruited upon damage, and its depletion caused lysosomal accumulation of PtdIns4P and resulted in cell death. We conclude that ER contacts are activated on damaged lysosomes in parallel to ESCRTs to provide lipids for membrane repair, and that PtdIns4P generation and removal are central in this response.


Assuntos
Receptores de Esteroides , Receptores de Esteroides/metabolismo , Retículo Endoplasmático/metabolismo , Lisossomos/metabolismo , Colesterol/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo
5.
EMBO J ; 40(19): e108863, 2021 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-34459017

RESUMO

Autophagy is a core molecular pathway for the preservation of cellular and organismal homeostasis. Pharmacological and genetic interventions impairing autophagy responses promote or aggravate disease in a plethora of experimental models. Consistently, mutations in autophagy-related processes cause severe human pathologies. Here, we review and discuss preclinical data linking autophagy dysfunction to the pathogenesis of major human disorders including cancer as well as cardiovascular, neurodegenerative, metabolic, pulmonary, renal, infectious, musculoskeletal, and ocular disorders.


Assuntos
Autofagia , Suscetibilidade a Doenças , Animais , Autofagia/efeitos dos fármacos , Autofagia/genética , Autofagia/imunologia , Biomarcadores , Regulação da Expressão Gênica , Predisposição Genética para Doença , Homeostase , Interações Hospedeiro-Patógeno , Humanos , Especificidade de Órgãos , Transdução de Sinais
6.
Rev Physiol Biochem Pharmacol ; 185: 107-152, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-33398504

RESUMO

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.


Assuntos
Neoplasias , Humanos , Morte Celular , Neoplasias/metabolismo , Membranas Intracelulares/metabolismo , Membranas Intracelulares/patologia , Lisossomos/metabolismo , Lisossomos/patologia , Transdução de Sinais
7.
EMBO J ; 36(13): 1811-1836, 2017 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-28596378

RESUMO

Over the past two decades, the molecular machinery that underlies autophagic responses has been characterized with ever increasing precision in multiple model organisms. Moreover, it has become clear that autophagy and autophagy-related processes have profound implications for human pathophysiology. However, considerable confusion persists about the use of appropriate terms to indicate specific types of autophagy and some components of the autophagy machinery, which may have detrimental effects on the expansion of the field. Driven by the overt recognition of such a potential obstacle, a panel of leading experts in the field attempts here to define several autophagy-related terms based on specific biochemical features. The ultimate objective of this collaborative exchange is to formulate recommendations that facilitate the dissemination of knowledge within and outside the field of autophagy research.


Assuntos
Autofagia , Terminologia como Assunto , Animais , Caenorhabditis elegans/fisiologia , Drosophila melanogaster/fisiologia , Redes Reguladoras de Genes , Camundongos , Saccharomyces cerevisiae/fisiologia
8.
Cancer Cell Int ; 21(1): 530, 2021 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-34641874

RESUMO

The transcription factor cyclic-AMP response element-binding protein 1 (CREB1) responds to cAMP level and controls the expression of target genes, which regulates nutrition partitioning. The promoters of CREB1-targeted genes responsive to cAMP have been extensively investigated and characterized with the presence of both cAMP response element and TATA box. Compelling evidence demonstrates that CREB1 also plays an essential role in promoting tumor development. However, only very few genes required for cell survival, proliferation and migration are known to be constitutively regulated by CREB1 in tumors. Their promoters mostly do not harbor any cAMP response element. Thus, it is very likely that CREB1 regulates the expressions of distinct sets of target genes in normal tissues and tumors. The whole gene network constitutively regulated by CREB1 in tumors has remained unrevealed. Here, we employ a systematical and integrative approach to decipher this gene network in the context of both tissue cultured cancer cells and patient samples. We combine transcriptomic, Rank-Rank Hypergeometric Overlap, and Chipseq analysis, to define and characterize CREB1-regulated genes in a multidimensional fashion. A strong cancer relevance of those top-ranked targets, which meet the most stringent criteria, is eventually verified by overall survival analysis of cancer patients. These findings strongly suggest the importance of genes constitutively regulated by CREB1 for their implicative involvement in promoting tumorigenesis.

9.
EMBO Rep ; 20(10): e48014, 2019 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-31432621

RESUMO

The autophagic clearance of damaged lysosomes by lysophagy involves extensive modification of the organelle with ubiquitin, but the underlying ubiquitination machinery is still poorly characterized. Here, we use an siRNA screening approach and identify human UBE2QL1 as a major regulator of lysosomal ubiquitination, lysophagy, and cell survival after lysosomal damage. UBE2QL1 translocates to permeabilized lysosomes where it associates with damage sensors, ubiquitination targets, and lysophagy effectors. UBE2QL1 knockdown reduces ubiquitination and accumulation of the critical autophagy receptor p62 and abrogates recruitment of the AAA-ATPase VCP/p97, which is essential for efficient lysophagy. Crucially, it affects association of LC3B with damaged lysosomes indicating that autophagosome formation was impaired. Already in unchallenged cells, depletion of UBE2QL1 leads to increased lysosomal damage, mTOR dissociation from lysosomes, and TFEB activation pointing to a role in lysosomal homeostasis. In line with this, mutation of the homologue ubc-25 in Caenorhabditis elegans exacerbates lysosome permeability in worms lacking the lysosome stabilizing protein SCAV-3/LIMP2. Thus, UBE2QL1 coordinates critical steps in the acute endolysosomal damage response and is essential for maintenance of lysosomal integrity.


Assuntos
Autofagia , Endossomos/metabolismo , Lisossomos/metabolismo , Enzimas de Conjugação de Ubiquitina/metabolismo , Adenosina Trifosfatases , Animais , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Sobrevivência Celular , Endossomos/ultraestrutura , Galectinas/metabolismo , Células HeLa , Humanos , Lisina/metabolismo , Lisossomos/ultraestrutura , Proteínas Associadas aos Microtúbulos/metabolismo , Proteínas Nucleares , Permeabilidade , RNA Interferente Pequeno/metabolismo , Proteína Sequestossoma-1/metabolismo , Ubiquitina/metabolismo , Ubiquitinação , Ubiquitinas/metabolismo
10.
Metabolomics ; 16(9): 91, 2020 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-32851548

RESUMO

INTRODUCTION: Repurposing of cationic amphiphilic drugs (CADs) emerges as an attractive therapeutic solution against various cancers, including leukemia. CADs target lysosomal lipid metabolism and preferentially kill cancer cells via induction of lysosomal membrane permeabilization, but the exact effects of CADs on the lysosomal lipid metabolism remain poorly illuminated. OBJECTIVES: We aimed to systematically monitor CAD-induced alterations in the quantitative lipid profiles of leukemia cell lines in order to chart effects of CADs on the metabolism of various lipid classes present in these cells. METHODS: We conducted this study on eight cultured cell lines representing two leukemia types, acute lymphoblastic leukemia and acute myeloid leukemia. Mass spectrometry-based quantitative shotgun lipidomics was employed to quantify the levels of around 400 lipid species of 26 lipid classes in the leukemia cell lines treated or untreated with a CAD, siramesine. RESULTS: The two leukemia types displayed high, but variable sensitivities to CADs and distinct profiles of cellular lipids. Treatment with siramesine rapidly altered the levels of diverse lipid classes in both leukemia types. These included sphingolipid classes previously reported to play key roles in CAD-induced cell death, but also lipids of other categories. We demonstrated that the treatment with siramesine additionally elevated the levels of numerous cytolytic lysoglycerophospholipids in positive correlation with the sensitivity of individual leukemia cell lines to siramesine. CONCLUSIONS: Our study shows that CAD treatment alters balance in the metabolism of glycerophospholipids, and proposes elevation in the levels of lysoglycerophospholipids as part of the mechanism leading to CAD-induced cell death of leukemia cells.


Assuntos
Morte Celular/efeitos dos fármacos , Leucemia/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Lipídeos , Preparações Farmacêuticas , Linhagem Celular Tumoral , Sobrevivência Celular , Humanos , Lipidômica , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Esfingolipídeos/metabolismo
11.
Mol Cell ; 45(6): 764-76, 2012 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-22464443

RESUMO

Aberrant ErbB2 receptor tyrosine kinase activation in breast cancer is strongly linked to an invasive disease. The molecular basis of ErbB2-driven invasion is largely unknown. We show that cysteine cathepsins B and L are elevated in ErbB2 positive primary human breast cancer and function as effectors of ErbB2-induced invasion in vitro. We identify Cdc42-binding protein kinase beta, extracellular regulated kinase 2, p21-activated protein kinase 4, and protein kinase C alpha as essential mediators of ErbB2-induced cysteine cathepsin expression and breast cancer cell invasiveness. The identified signaling network activates the transcription of cathepsin B gene (CTSB) via myeloid zinc finger-1 transcription factor that binds to an ErbB2-responsive enhancer element in the first intron of CTSB. This work provides a model system for ErbB2-induced breast cancer cell invasiveness, reveals a signaling network that is crucial for invasion in vitro, and defines a specific role and targets for the identified serine-threonine kinases.


Assuntos
Neoplasias da Mama/patologia , Catepsina B/genética , Catepsina B/metabolismo , Fatores de Transcrição Kruppel-Like/metabolismo , Receptor ErbB-2/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Catepsina L/genética , Catepsina L/metabolismo , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Fatores de Transcrição Kruppel-Like/genética , Proteína Quinase 1 Ativada por Mitógeno/genética , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Miotonina Proteína Quinase , Invasividade Neoplásica , Regiões Promotoras Genéticas , Proteína Quinase C-alfa/genética , Proteína Quinase C-alfa/metabolismo , Proteínas Tirosina Quinases/genética , Proteínas Tirosina Quinases/metabolismo , Proteína Proto-Oncogênica c-ets-1/genética , Proteína Proto-Oncogênica c-ets-1/metabolismo , Receptor ErbB-2/genética , Elementos de Resposta , Transdução de Sinais , Quinases Ativadas por p21/genética , Quinases Ativadas por p21/metabolismo
12.
Int J Cancer ; 143(6): 1315-1326, 2018 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-29658114

RESUMO

Proton pump inhibitors (PPIs) are commonly used as a supplement to cancer therapy. Yet, their effect on cancer mortality is largely unknown. Using data from Danish nationwide registries and Cox models regressing of both propensity scores and drug use, we estimated hazard ratios (HRs) with 95% confidence intervals (CIs) for cancer-specific and noncancer death among PPI users (≥2 prescriptions within six months after diagnosis; n = 36,066) compared with nonusers (<2 prescriptions, n = 311,853) or users of histamine H2 -receptor antagonists (H2 RA; n = 5,152). Adjusted HRs for cancer-specific mortality among postdiagnostic PPI users as compared with nonusers or H2 RA users were 1.29 (95% CI, 1.27-1.32) and 1.15 (95% CI, 1.10-1.20), respectively. HRs for cancer mortality associated with PPI use were highest for ovarian (1.35; 95% CI, 1.20-1.52) and lowest for esophageal cancer (0.91; 95% CI, 0.81-1.04). The associations were stronger among new PPI users after cancer diagnosis, indicating potential confounding. To test the effect of PPIs on tumor growth in a model system free for confounding factors, we investigated the effect of pantoprazole on tumor growth in mice. Pantoprazole (5 mg/kg/day) enhanced tumor growth (p = 0.033) and reduced the antitumor activity of gemcitabine (p = 0.008) in fibrosarcoma-bearing Balb/c mice, but not in immunodeficient Balb/c nude mice. In breast carcinoma-bearing FVB/N mice, pantoprazole had no effect on tumor growth alone but it reduced the life-prolonging effect of doxorubicin significantly (p = 0.007). Taken together, these data raise concerns about the increasing use of PPIs and calls for further studies addressing their safety among cancer patients.


Assuntos
Neoplasias/mortalidade , Inibidores da Bomba de Prótons/efeitos adversos , Idoso , Animais , Estudos de Coortes , Feminino , Seguimentos , Humanos , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Prognóstico , Taxa de Sobrevida , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
13.
Nucleic Acids Res ; 44(5): 2214-26, 2016 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-26721387

RESUMO

We have recently identified lens epithelium-derived growth factor (LEDGF/p75, also known as PSIP1) as a component of the homologous recombination DNA repair machinery. Through its Pro-Trp-Trp-Pro (PWWP) domain, LEDGF/p75 binds to histone marks associated with active transcription and promotes DNA end resection by recruiting DNA endonuclease retinoblastoma-binding protein 8 (RBBP8/CtIP) to broken DNA ends. Here we show that the structurally related PWWP domain-containing protein, hepatoma-derived growth factor-related protein 2 (HDGFRP2), serves a similar function in homologous recombination repair. Its depletion compromises the survival of human U2OS osteosarcoma and HeLa cervix carcinoma cells and impairs the DNA damage-induced phosphorylation of replication protein A2 (RPA2) and the recruitment of DNA endonuclease RBBP8/CtIP to DNA double strand breaks. In contrast to LEDGF/p75, HDGFRP2 binds preferentially to histone marks characteristic for transcriptionally silent chromatin. Accordingly, HDGFRP2 is found in complex with the heterochromatin-binding chromobox homologue 1 (CBX1) and Pogo transposable element with ZNF domain (POGZ). Supporting the functionality of this complex, POGZ-depleted cells show a similar defect in DNA damage-induced RPA2 phosphorylation as HDGFRP2-depleted cells. These data suggest that HDGFRP2, possibly in complex with POGZ, recruits homologous recombination repair machinery to damaged silent genes or to active genes silenced upon DNA damage.


Assuntos
Proteínas Cromossômicas não Histona/genética , Quebras de DNA de Cadeia Dupla , Histonas/genética , Peptídeos e Proteínas de Sinalização Intercelular/genética , Reparo de DNA por Recombinação , Transposases/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Sítios de Ligação , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular , Cromatina/química , Cromatina/metabolismo , Homólogo 5 da Proteína Cromobox , Proteínas Cromossômicas não Histona/metabolismo , Endodesoxirribonucleases , Células HeLa , Histonas/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Osteoblastos/metabolismo , Osteoblastos/patologia , Fosforilação , Ligação Proteica , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteína de Replicação A/genética , Proteína de Replicação A/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Transposases/antagonistas & inibidores , Transposases/metabolismo
14.
Nature ; 463(7280): 549-53, 2010 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-20111001

RESUMO

Heat shock protein 70 (Hsp70) is an evolutionarily highly conserved molecular chaperone that promotes the survival of stressed cells by inhibiting lysosomal membrane permeabilization, a hallmark of stress-induced cell death. Clues to its molecular mechanism of action may lay in the recently reported stress- and cancer-associated translocation of a small portion of Hsp70 to the lysosomal compartment. Here we show that Hsp70 stabilizes lysosomes by binding to an endolysosomal anionic phospholipid bis(monoacylglycero)phosphate (BMP), an essential co-factor for lysosomal sphingomyelin metabolism. In acidic environments Hsp70 binds with high affinity and specificity to BMP, thereby facilitating the BMP binding and activity of acid sphingomyelinase (ASM). The inhibition of the Hsp70-BMP interaction by BMP antibodies or a point mutation in Hsp70 (Trp90Phe), as well as the pharmacological and genetic inhibition of ASM, effectively revert the Hsp70-mediated stabilization of lysosomes. Notably, the reduced ASM activity in cells from patients with Niemann-Pick disease (NPD) A and B-severe lysosomal storage disorders caused by mutations in the sphingomyelin phosphodiesterase 1 gene (SMPD1) encoding for ASM-is also associated with a marked decrease in lysosomal stability, and this phenotype can be effectively corrected by treatment with recombinant Hsp70. Taken together, these data open exciting possibilities for the development of new treatments for lysosomal storage disorders and cancer with compounds that enter the lysosomal lumen by the endocytic delivery pathway.


Assuntos
Proteínas de Choque Térmico HSP70/metabolismo , Lisossomos/metabolismo , Lisossomos/patologia , Doenças de Niemann-Pick/metabolismo , Doenças de Niemann-Pick/patologia , Linhagem Celular Tumoral , Células Cultivadas , Humanos , Concentração de Íons de Hidrogênio , Membranas Intracelulares/metabolismo , Lisofosfolipídeos/metabolismo , Monoglicerídeos/metabolismo , Esfingomielina Fosfodiesterase/metabolismo
15.
Biochim Biophys Acta ; 1838(5): 1344-61, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24480410

RESUMO

While a significant fraction of heat shock protein 70 (Hsp70) is membrane associated in lysosomes, mitochondria, and the outer surface of cancer cells, the mechanisms of interaction have remained elusive, with no conclusive demonstration of a protein receptor. Hsp70 contains two Trps, W90 and W580, in its N-terminal nucleotide binding domain (NBD), and the C-terminal substrate binding domain (SBD), respectively. Our fluorescence spectroscopy study using Hsp70 and its W90F and W580F mutants, and Hsp70-∆SBD and Hsp70-∆NBD constructs, revealed that binding to liposomes depends on their lipid composition and involves both NBD and SBD. Association of Hsp70 with phosphatidylcholine (PC) liposomes is weak, with insertion of its Trps into the bilayer hydrocarbon region. In the presence of cardiolipin (CL), bis-monoacylglycero phosphate (BMP), or phosphatidylserine (PS) Hsp70 attaches to membranes peripherally, without penetration. Our data suggest that the organelle distribution of Hsp70 is determined by their specific lipid compositions, with Hsp70 associating with the above lipids in mitochondria, lysosomes, and the surface of cancer cells, respectively. NBD and SBD attach to lipids by extended phospholipid anchorage, with specific acidic phospholipids associating with Hsp70 in the extended conformation with acyl chains inserting into hydrophobic crevices within Hsp70, and other chains remaining in the bilayer. This anchorage is expected to cause a stringent orientation of Hsp70 on the surface. Our data further suggest that acidic phospholipids induce a transition of SBD into the molten globule state, which may be essential to allow SBD-substrate interaction also within the hydrophobic bilayer interior acyl chain region.


Assuntos
Membrana Celular/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Bicamadas Lipídicas/metabolismo , Proteínas de Membrana/metabolismo , Cardiolipinas/metabolismo , Humanos , Metabolismo dos Lipídeos , Lipossomos/metabolismo , Mitocôndrias/metabolismo , Modelos Moleculares , Fosfatidilcolinas/metabolismo , Fosfatidilserinas/metabolismo , Fosfolipídeos/metabolismo , Ligação Proteica , Estrutura Terciária de Proteína
16.
EMBO J ; 30(22): 4628-41, 2011 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-21915098

RESUMO

Autophagy is an evolutionarily conserved mechanism of cellular self-digestion in which proteins and organelles are degraded through delivery to lysosomes. Defects in this process are implicated in numerous human diseases including cancer. To further elucidate regulatory mechanisms of autophagy, we performed a functional screen in search of microRNAs (miRNAs), which regulate the autophagic flux in breast cancer cells. In this study, we identified the tumour suppressive miRNA, miR-101, as a potent inhibitor of basal, etoposide- and rapamycin-induced autophagy. Through transcriptome profiling, we identified three novel miR-101 targets, STMN1, RAB5A and ATG4D. siRNA-mediated depletion of these genes phenocopied the effect of miR-101 overexpression, demonstrating their importance in autophagy regulation. Importantly, overexpression of STMN1 could partially rescue cells from miR-101-mediated inhibition of autophagy, indicating a functional importance for this target. Finally, we show that miR-101-mediated inhibition of autophagy can sensitize breast cancer cells to 4-hydroxytamoxifen (4-OHT)-mediated cell death. Collectively, these data establish a novel link between two highly important and rapidly growing research fields and present a new role for miR-101 as a key regulator of autophagy.


Assuntos
Autofagia , MicroRNAs/genética , MicroRNAs/metabolismo , Estatmina/metabolismo , Proteínas Relacionadas à Autofagia , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Cisteína Endopeptidases/genética , Cisteína Endopeptidases/metabolismo , Etoposídeo/farmacologia , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Humanos , Análise de Sequência com Séries de Oligonucleotídeos , Interferência de RNA , RNA Interferente Pequeno , Sirolimo/farmacologia , Estatmina/biossíntese , Estatmina/genética , Tamoxifeno/análogos & derivados , Tamoxifeno/farmacologia , Proteínas rab5 de Ligação ao GTP/genética , Proteínas rab5 de Ligação ao GTP/metabolismo
17.
J Cell Sci ; 126(Pt 9): 1905-12, 2013 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-23720375

RESUMO

Lysosomes serve as the cellular recycling centre and are filled with numerous hydrolases that can degrade most cellular macromolecules. Lysosomal membrane permeabilization and the consequent leakage of the lysosomal content into the cytosol leads to so-called "lysosomal cell death". This form of cell death is mainly carried out by the lysosomal cathepsin proteases and can have necrotic, apoptotic or apoptosis-like features depending on the extent of the leakage and the cellular context. This article summarizes our current knowledge on lysosomal cell death with an emphasis on the upstream mechanisms that lead to lysosomal membrane permeabilization.


Assuntos
Apoptose/fisiologia , Catepsinas/metabolismo , Membranas Intracelulares/metabolismo , Lisossomos/enzimologia , Animais , Humanos , Permeabilidade
18.
Mol Cell Proteomics ; 11(3): M111.014035, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22311637

RESUMO

Autophagy is one of the major intracellular catabolic pathways, but little is known about the composition of autophagosomes. To study the associated proteins, we isolated autophagosomes from human breast cancer cells using two different biochemical methods and three stimulus types: amino acid deprivation or rapamycin or concanamycin A treatment. The autophagosome-associated proteins were dependent on stimulus, but a core set of proteins was stimulus-independent. Remarkably, proteasomal proteins were abundant among the stimulus-independent common autophagosome-associated proteins, and the activation of autophagy significantly decreased the cellular proteasome level and activity supporting interplay between the two degradation pathways. A screen of yeast strains defective in the orthologs of the human genes encoding for a common set of autophagosome-associated proteins revealed several regulators of autophagy, including subunits of the retromer complex. The combined spatiotemporal proteomic and genetic data sets presented here provide a basis for further characterization of autophagosome biogenesis and cargo selection.


Assuntos
Autofagia , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Testes Genéticos , Fagossomos/metabolismo , Proteínas/metabolismo , Proteômica , Aminoácidos/metabolismo , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/metabolismo , Antivirais/farmacologia , Neoplasias da Mama/patologia , Eletroforese em Gel de Poliacrilamida , Feminino , Proteínas de Fluorescência Verde/imunologia , Proteínas de Fluorescência Verde/metabolismo , Humanos , Imunoprecipitação , Imunossupressores/farmacologia , Marcação por Isótopo , Lisossomos/metabolismo , Macrolídeos/farmacologia , Fagossomos/efeitos dos fármacos , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Sirolimo/farmacologia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Inanição , Células Tumorais Cultivadas
19.
Nat Rev Cancer ; 5(11): 886-97, 2005 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-16239905

RESUMO

Lysosomal hydrolases participate in the digestion of endocytosed and autophagocytosed material inside the lysosomal/autolysosomal compartment in acute cell death when released into the cytosol and in cancer progression following their release into the extracellular space. Lysosomal alterations are common in cancer cells. The increased expression and altered trafficking of lysosomal enzymes participates in tissue invasion, angiogenesis and sensitization to the lysosomal death pathway. But lysosomal heat-shock protein 70 locally prevents lysosomal-membrane permeabilization. Similarly, alterations in the autophagic compartment are linked to carcinogenesis and resistance to chemotherapy. Targeting these pathways might constitute a novel approach to cancer therapy.


Assuntos
Autofagia/fisiologia , Transformação Celular Neoplásica/metabolismo , Lisossomos/fisiologia , Animais , Humanos , Hidrolases/metabolismo
20.
Cell Metab ; 36(3): 461-462, 2024 03 05.
Artigo em Inglês | MEDLINE | ID: mdl-38447528

RESUMO

Cancer metabolism produces large fluxes of lactate and H+, which are extruded by membrane transporters. However, H+ production and extrusion must be coupled by diffusion, facilitated by mobile buffers. Yan et al. propose that carnosine, generated by CARNS2, provides this mobile buffering and enables lysosomal functions that block T cell surveillance.


Assuntos
Carnosina , Carnosina/farmacologia , Linfócitos T , Ácido Láctico , Proteínas de Membrana Transportadoras
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