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1.
Cell ; 187(2): 294-311.e21, 2024 01 18.
Artículo en Inglés | MEDLINE | ID: mdl-38128537

RESUMEN

Lactylation is a lactate-induced post-translational modification best known for its roles in epigenetic regulation. Herein, we demonstrate that MRE11, a crucial homologous recombination (HR) protein, is lactylated at K673 by the CBP acetyltransferase in response to DNA damage and dependent on ATM phosphorylation of the latter. MRE11 lactylation promotes its binding to DNA, facilitating DNA end resection and HR. Inhibition of CBP or LDH downregulated MRE11 lactylation, impaired HR, and enhanced chemosensitivity of tumor cells in patient-derived xenograft and organoid models. A cell-penetrating peptide that specifically blocks MRE11 lactylation inhibited HR and sensitized cancer cells to cisplatin and PARPi. These findings unveil lactylation as a key regulator of HR, providing fresh insights into the ways in which cellular metabolism is linked to DSB repair. They also imply that the Warburg effect can confer chemoresistance through enhancing HR and suggest a potential therapeutic strategy of targeting MRE11 lactylation to mitigate the effects.


Asunto(s)
Proteínas de Unión al ADN , Proteína Homóloga de MRE11 , Reparación del ADN por Recombinación , Humanos , ADN , Roturas del ADN de Doble Cadena , Reparación del ADN , Proteínas de Unión al ADN/metabolismo , Epigénesis Genética , Recombinación Homóloga , Proteína Homóloga de MRE11/metabolismo , Ácido Láctico/metabolismo
2.
Mol Cell ; 83(4): 539-555.e7, 2023 02 16.
Artículo en Inglés | MEDLINE | ID: mdl-36702126

RESUMEN

Replication protein A (RPA) is a major regulator of eukaryotic DNA metabolism involved in multiple essential cellular processes. Maintaining appropriate RPA dynamics is crucial for cells to prevent RPA exhaustion, which can lead to replication fork breakage and replication catastrophe. However, how cells regulate RPA availability during unperturbed replication and in response to stress has not been well elucidated. Here, we show that HNRNPA2B1SUMO functions as an endogenous inhibitor of RPA during normal replication. HNRNPA2B1SUMO associates with RPA through recognizing the SUMO-interacting motif (SIM) of RPA to inhibit RPA accumulation at replication forks and impede local ATR activation. Declining HNRNPA2SUMO induced by DNA damage will release nuclear soluble RPA to localize to chromatin and enable ATR activation. Furthermore, we characterize that HNRNPA2B1 hinders homologous recombination (HR) repair via limiting RPA availability, thus conferring sensitivity to PARP inhibitors. These findings establish HNRNPA2B1 as a critical player in RPA-dependent surveillance networks.


Asunto(s)
Replicación del ADN , Proteína de Replicación A , Proteína de Replicación A/genética , Proteína de Replicación A/metabolismo , Replicación del ADN/genética , Sumoilación , Daño del ADN , Cromatina/genética , Proteínas de la Ataxia Telangiectasia Mutada/genética , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo
3.
Mol Cell ; 83(7): 1043-1060.e10, 2023 04 06.
Artículo en Inglés | MEDLINE | ID: mdl-36854302

RESUMEN

Repair of DNA double-strand breaks (DSBs) elicits three-dimensional (3D) chromatin topological changes. A recent finding reveals that 53BP1 assembles into a 3D chromatin topology pattern around DSBs. How this formation of a higher-order structure is configured and regulated remains enigmatic. Here, we report that SLFN5 is a critical factor for 53BP1 topological arrangement at DSBs. Using super-resolution imaging, we find that SLFN5 binds to 53BP1 chromatin domains to assemble a higher-order microdomain architecture by driving damaged chromatin dynamics at both DSBs and deprotected telomeres. Mechanistically, we propose that 53BP1 topology is shaped by two processes: (1) chromatin mobility driven by the SLFN5-LINC-microtubule axis and (2) the assembly of 53BP1 oligomers mediated by SLFN5. In mammals, SLFN5 deficiency disrupts the DSB repair topology and impairs non-homologous end joining, telomere fusions, class switch recombination, and sensitivity to poly (ADP-ribose) polymerase inhibitor. We establish a molecular mechanism that shapes higher-order chromatin topologies to safeguard genomic stability.


Asunto(s)
Cromatina , Reparación del ADN , Animales , Cromatina/genética , Roturas del ADN de Doble Cadena , Reparación del ADN por Unión de Extremidades , Mamíferos/metabolismo , Proteínas de Unión a Telómeros/genética , Proteína 1 de Unión al Supresor Tumoral P53/genética , Proteína 1 de Unión al Supresor Tumoral P53/metabolismo , Proteínas de Ciclo Celular/metabolismo
4.
Mol Cell ; 79(5): 824-835.e5, 2020 09 03.
Artículo en Inglés | MEDLINE | ID: mdl-32649882

RESUMEN

DNA-protein crosslinks (DPCs) are highly toxic DNA lesions that threaten genomic integrity. Recent findings highlight that SPRTN, a specialized DNA-dependent metalloprotease, is a central player in proteolytic cleavage of DPCs. Previous studies suggest that SPRTN deubiquitination is important for its chromatin association and activation. However, the regulation and consequences of SPRTN deubiquitination remain unclear. Here we report that, in response to DPC induction, the deubiquitinase VCPIP1/VCIP135 is phosphorylated and activated by ATM/ATR. VCPIP1, in turn, deubiquitinates SPRTN and promotes its chromatin relocalization. Deubiquitination of SPRTN is required for its subsequent acetylation, which promotes SPRTN relocation to the site of chromatin damage. Furthermore, Vcpip1 knockout mice are prone to genomic instability and premature aging. We propose a model where two sequential post-translational modifications (PTMs) regulate SPRTN chromatin accessibility to repair DPCs and maintain genomic stability and a healthy lifespan.


Asunto(s)
Envejecimiento/genética , Reparación del ADN , Proteínas de Unión al ADN/metabolismo , Acetilación , Envejecimiento/metabolismo , Animales , Línea Celular , Daño del ADN , Proteínas de Unión al ADN/genética , Enzimas Desubicuitinizantes/metabolismo , Endopeptidasas/metabolismo , Femenino , Inestabilidad Genómica , Células HEK293 , Humanos , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Fosforilación , Dominios Proteicos , Procesamiento Proteico-Postraduccional , Ubiquitinación
5.
Mol Cell ; 74(6): 1215-1226.e4, 2019 06 20.
Artículo en Inglés | MEDLINE | ID: mdl-31053471

RESUMEN

Programmed death ligand 1 (PD-L1, also called B7-H1) is an immune checkpoint protein that inhibits immune function through its binding of the programmed cell death protein 1 (PD-1) receptor. Clinically approved antibodies block extracellular PD-1 and PD-L1 binding, yet the role of intracellular PD-L1 in cancer remains poorly understood. Here, we discovered that intracellular PD-L1 acts as an RNA binding protein that regulates the mRNA stability of NBS1, BRCA1, and other DNA damage-related genes. Through competition with the RNA exosome, intracellular PD-L1 protects targeted RNAs from degradation, thereby increasing cellular resistance to DNA damage. RNA immunoprecipitation and RNA-seq experiments demonstrated that PD-L1 regulates RNA stability genome-wide. Furthermore, we developed a PD-L1 antibody, H1A, which abrogates the interaction of PD-L1 with CMTM6, thereby promoting PD-L1 degradation. Intracellular PD-L1 may be a potential therapeutic target to enhance the efficacy of radiotherapy and chemotherapy in cancer through the inhibition of DNA damage response and repair.


Asunto(s)
Antígeno B7-H1/genética , Reparación del ADN , ADN de Neoplasias/genética , Complejo Multienzimático de Ribonucleasas del Exosoma/genética , Regulación Neoplásica de la Expresión Génica , Receptor de Muerte Celular Programada 1/genética , Animales , Antineoplásicos/farmacología , Antígeno B7-H1/antagonistas & inhibidores , Antígeno B7-H1/metabolismo , Proteína BRCA1/genética , Proteína BRCA1/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Cisplatino/farmacología , Daño del ADN , ADN de Neoplasias/metabolismo , Complejo Multienzimático de Ribonucleasas del Exosoma/metabolismo , Rayos gamma/uso terapéutico , Células HCT116 , Células HeLa , Humanos , Proteínas con Dominio MARVEL , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Proteínas de la Mielina , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Receptor de Muerte Celular Programada 1/metabolismo , Proteolisis/efectos de los fármacos , Proteolisis/efectos de la radiación , Estabilidad del ARN/efectos de los fármacos , Estabilidad del ARN/efectos de la radiación , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Ensayos Antitumor por Modelo de Xenoinjerto
6.
Am J Pathol ; 194(5): 656-672, 2024 05.
Artículo en Inglés | MEDLINE | ID: mdl-38325552

RESUMEN

Idiopathic pulmonary fibrosis is a progressive interstitial lung disease for which there is no curative therapy available. Repetitive alveolar epithelial injury repair, myofibroblast accumulation, and excessive collagen deposition are key pathologic features of idiopathic pulmonary fibrosis, eventually leading to cellular hypoxia and respiratory failure. The precise mechanism driving this complex maladaptive process remains inadequately understood. WD repeat and suppressor of cytokine signaling box containing 1 (WSB1) is an E3 ubiquitin ligase, the expression of which is associated strongly with hypoxia, and forms a positive feedback loop with hypoxia-inducible factor 1α (HIF-1α) under anoxic condition. This study explored the expression, cellular distribution, and function of WSB1 in bleomycin (BLM)-induced mouse lung injury and fibrosis. WSB1 expression was highly induced by BLM injury and correlated with the progression of lung fibrosis. Significantly, conditional deletion of Wsb1 in adult mice ameliorated BLM-induced pulmonary fibrosis. Phenotypically, Wsb1-deficient mice showed reduced lipofibroblast to myofibroblast transition, but enhanced alveolar type 2 proliferation and differentiation into alveolar type 1 after BLM injury. Proteomic analysis of mouse lung tissues identified caveolin 2 as a potential downstream target of WSB1, contributing to BLM-induced epithelial injury repair and fibrosis. These findings unravel a vital role for WSB1 induction in lung injury repair, thus highlighting it as a potential therapeutic target for pulmonary fibrosis.


Asunto(s)
Fibrosis Pulmonar Idiopática , Lesión Pulmonar , Animales , Ratones , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismo , Miofibroblastos/metabolismo , Lesión Pulmonar/patología , Proteómica , Pulmón/patología , Fibrosis , Hipoxia/patología , Fibrosis Pulmonar Idiopática/patología , Bleomicina/toxicidad , Regeneración , Péptidos y Proteínas de Señalización Intracelular
7.
Cell ; 140(3): 384-96, 2010 Feb 05.
Artículo en Inglés | MEDLINE | ID: mdl-20096447

RESUMEN

Stability and localization of p53 is essential for its tumor suppressor function. Ubiquitination by the E3 ubiquitin ligase Mdm2 is the major regulatory mechanism of p53, which induces p53 nuclear export and degradation. However, it is unclear whether ubiquitinated cytoplasmic p53 can be recycled. Here, we report that USP10, a cytoplasmic ubiquitin-specific protease, deubiquitinates p53, reversing Mdm2-induced p53 nuclear export and degradation. After DNA damage, USP10 is stabilized, and a fraction of USP10 translocates to the nucleus to activate p53. The translocation and stabilization of USP10 is regulated by ATM -mediated phosphorylation of USP10 at Thr42 and Ser337. Finally, USP10 suppresses tumor cell growth in cells with wild-type p53, with USP10 expression downregulated in a high percentage of clear cell carcinomas, known to have few p53 mutations. These findings reveal USP10 to be a novel regulator of p53, providing an alternative mechanism of p53 inhibition in cancers with wild-type p53.


Asunto(s)
Regulación Neoplásica de la Expresión Génica , Proteína p53 Supresora de Tumor/metabolismo , Ubiquitina Tiolesterasa/metabolismo , Transporte Activo de Núcleo Celular , Proteínas de la Ataxia Telangiectasia Mutada , Carcinoma de Células Renales/metabolismo , Proteínas de Ciclo Celular/metabolismo , Línea Celular , Reparación del ADN , Proteínas de Unión al ADN/metabolismo , Células HCT116 , Humanos , Neoplasias Renales/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Estabilidad Proteica , Proteína p53 Supresora de Tumor/análisis , Proteínas Supresoras de Tumor/metabolismo , Ubiquitinación
8.
Drug Resist Updat ; 74: 101085, 2024 May.
Artículo en Inglés | MEDLINE | ID: mdl-38636338

RESUMEN

Enhanced DNA repair is an important mechanism of inherent and acquired resistance to DNA targeted therapies, including poly ADP ribose polymerase (PARP) inhibition. Spleen associated tyrosine kinase (Syk) is a non-receptor tyrosine kinase acknowledged for its regulatory roles in immune cell function, cell adhesion, and vascular development. This study presents evidence indicating that Syk expression in high-grade serous ovarian cancer and triple-negative breast cancers promotes DNA double-strand break resection, homologous recombination (HR), and subsequent therapeutic resistance. Our investigations reveal that Syk is activated by ATM following DNA damage and is recruited to DNA double-strand breaks by NBS1. Once localized to the break site, Syk phosphorylates CtIP, a pivotal mediator of resection and HR, at Thr-847 to promote repair activity, particularly in Syk-expressing cancer cells. Inhibition of Syk or its genetic deletion impedes CtIP Thr-847 phosphorylation and overcomes the resistant phenotype. Collectively, our findings suggest a model wherein Syk fosters therapeutic resistance by promoting DNA resection and HR through a hitherto uncharacterized ATM-Syk-CtIP pathway. Moreover, Syk emerges as a promising tumor-specific target to sensitize Syk-expressing tumors to PARP inhibitors, radiation and other DNA-targeted therapies.


Asunto(s)
Roturas del ADN de Doble Cadena , Resistencia a Antineoplásicos , Recombinación Homóloga , Quinasa Syk , Quinasa Syk/metabolismo , Quinasa Syk/genética , Quinasa Syk/antagonistas & inhibidores , Humanos , Roturas del ADN de Doble Cadena/efectos de los fármacos , Femenino , Resistencia a Antineoplásicos/genética , Resistencia a Antineoplásicos/efectos de los fármacos , Fosforilación , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/uso terapéutico , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Reparación del ADN/efectos de los fármacos , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Proteínas de la Ataxia Telangiectasia Mutada/antagonistas & inhibidores , Proteínas de la Ataxia Telangiectasia Mutada/genética , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/patología , Animales , Línea Celular Tumoral , Daño del ADN/efectos de los fármacos
9.
Mol Cell ; 61(4): 614-624, 2016 Feb 18.
Artículo en Inglés | MEDLINE | ID: mdl-26876938

RESUMEN

The AMP-activated protein kinase (AMPK) is the master regulator of metabolic homeostasis by sensing cellular energy status. When intracellular ATP levels decrease during energy stress, AMPK is initially activated through AMP or ADP binding and phosphorylation of a threonine residue (Thr-172) within the activation loop of its kinase domain. Here we report a key molecular mechanism by which AMPK activation is amplified under energy stress. We found that ubiquitination on AMPKα blocks AMPKα phosphorylation by LKB1. The deubiquitinase USP10 specifically removes ubiquitination on AMPKα to facilitate AMPKα phosphorylation by LKB1. Under energy stress, USP10 activity in turn is enhanced through AMPK-mediated phosphorylation of Ser76 of USP10. Thus, USP10 and AMPK form a key feedforward loop ensuring amplification of AMPK activation in response to fluctuation of cellular energy status. Disruption of this feedforward loop leads to improper AMPK activation and multiple metabolic defects.


Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Ubiquitina Tiolesterasa/química , Ubiquitina Tiolesterasa/metabolismo , Animales , Metabolismo Energético , Activación Enzimática , Células HCT116 , Células HEK293 , Humanos , Ratones , Fosforilación , Proteínas Serina-Treonina Quinasas/metabolismo , Serina/metabolismo , Ubiquitinación
10.
Genes Dev ; 30(23): 2581-2595, 2016 12 01.
Artículo en Inglés | MEDLINE | ID: mdl-27941124

RESUMEN

Homologous recombination (HR) is one of the major DNA double-strand break (DSB) repair pathways in mammalian cells. Defects in HR trigger genomic instability and result in cancer predisposition. The defining step of HR is homologous strand exchange directed by the protein RAD51, which is recruited to DSBs by BRCA2. However, the regulation of the BRCA2-RAD51 axis remains unclear. Here we report that ubiquitination of RAD51 hinders RAD51-BRCA2 interaction, while deubiquitination of RAD51 facilitates RAD51-BRCA2 binding and RAD51 recruitment and thus is critical for proper HR. Mechanistically, in response to DNA damage, the deubiquitinase UCHL3 is phosphorylated and activated by ATM. UCHL3, in turn, deubiquitinates RAD51 and promotes the binding between RAD51 and BRCA2. Overexpression of UCHL3 renders breast cancer cells resistant to radiation and chemotherapy, while depletion of UCHL3 sensitizes cells to these treatments, suggesting a determinant role of UCHL3 in cancer therapy. Overall, we identify UCHL3 as a novel regulator of DNA repair and reveal a model in which a phosphorylation-deubiquitination cascade dynamically regulates the BRCA2-RAD51 pathway.


Asunto(s)
Proteína BRCA2/metabolismo , Neoplasias de la Mama/genética , Recombinación Homóloga/genética , Recombinasa Rad51/metabolismo , Ubiquitinación/genética , Proteína BRCA2/genética , Neoplasias de la Mama/enzimología , Neoplasias de la Mama/fisiopatología , Línea Celular Tumoral , Cisteína Endopeptidasas/genética , Cisteína Endopeptidasas/metabolismo , Reparación del ADN/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Técnicas de Inactivación de Genes , Células HEK293 , Recombinación Homóloga/efectos de los fármacos , Humanos , Células MCF-7 , Fosforilación/genética , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Unión Proteica/genética , Recombinasa Rad51/genética , Tolerancia a Radiación/efectos de los fármacos , Tolerancia a Radiación/genética , Transducción de Señal/genética , Análisis de Supervivencia , Ubiquitina Tiolesterasa , Ubiquitinación/efectos de los fármacos
11.
Genes Dev ; 30(8): 946-59, 2016 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-27083998

RESUMEN

Dynamic regulation of RNF168-mediated ubiquitylation of histone H2A Lys13,15 (H2AK13,15ub) at DNA double-strand breaks (DSBs) is crucial for preventing aberrant DNA repair and maintaining genome stability. However, it remains unclear which deubiquitylating enzyme (DUB) removes H2AK13,15ub. Here we show that USP51, a previously uncharacterized DUB, deubiquitylates H2AK13,15ub and regulates DNA damage response. USP51 depletion results in increased spontaneous DNA damage foci and elevated levels of H2AK15ub and impairs DNA damage response. USP51 overexpression suppresses the formation of ionizing radiation-induced 53BP1 and BRCA1 but not RNF168 foci, suggesting that USP51 functions downstream from RNF168 in DNA damage response. In vitro, USP51 binds to H2A-H2B directly and deubiquitylates H2AK13,15ub. In cells, USP51 is recruited to chromatin after DNA damage and regulates the dynamic assembly/disassembly of 53BP1 and BRCA1 foci. These results show that USP51 is the DUB for H2AK13,15ub and regulates DNA damage response.


Asunto(s)
Daño del ADN/fisiología , Reparación del ADN/fisiología , Histonas/metabolismo , Proteasas Ubiquitina-Específicas/metabolismo , Proteínas Adaptadoras Transductoras de Señales , Proteínas de Ciclo Celular , Línea Celular Tumoral , Supervivencia Celular/fisiología , ADN/metabolismo , ADN/efectos de la radiación , Humanos , Proteínas Nucleares/metabolismo , Unión Proteica , Radiación Ionizante , Transactivadores/metabolismo , Proteasas Ubiquitina-Específicas/genética , Ubiquitinación
12.
Semin Cancer Biol ; 85: 164-184, 2022 10.
Artículo en Inglés | MEDLINE | ID: mdl-33905873

RESUMEN

The acquisition of DNA damage is an early driving event in tumorigenesis. Premalignant lesions show activated DNA damage responses and inactivation of DNA damage checkpoints promotes malignant transformation. However, DNA damage is also a targetable vulnerability in cancer cells. This requires a detailed understanding of the cellular and molecular mechanisms governing DNA integrity. Here, we review current work on DNA damage in tumorigenesis. We discuss DNA double strand break repair, how repair pathways contribute to tumorigenesis, and how double strand breaks are linked to the tumor microenvironment. Next, we discuss the role of oncogenes in promoting DNA damage through replication stress. Finally, we discuss our current understanding on DNA damage in micronuclei and discuss therapies targeting these DNA damage pathways.


Asunto(s)
Daño del ADN , Reparación del ADN , Humanos , Reparación del ADN/genética , Roturas del ADN de Doble Cadena , Transformación Celular Neoplásica/metabolismo , ADN , Inestabilidad Genómica , Microambiente Tumoral
13.
J Biol Chem ; 298(2): 101563, 2022 02.
Artículo en Inglés | MEDLINE | ID: mdl-34998823

RESUMEN

The cytidine deaminase APOBEC3B (A3B) is an endogenous inducer of somatic mutations and causes chromosomal instability by converting cytosine to uracil in single-stranded DNA. Therefore, identification of factors and mechanisms that mediate A3B expression will be helpful for developing therapeutic approaches to decrease DNA mutagenesis. Arsenic (As) is one well-known mutagen and carcinogen, but the mechanisms by which it induces mutations have not been fully elucidated. Herein, we show that A3B is upregulated and required for As-induced DNA damage and mutagenesis. We found that As treatment causes a decrease of N6-methyladenosine (m6A) modification near the stop codon of A3B, consequently increasing the stability of A3B mRNA. We further reveal that the demethylase FTO is responsible for As-reduced m6A modification of A3B, leading to increased A3B expression and DNA mutation rates in a manner dependent on the m6A reader YTHDF2. Our in vivo data also confirm that A3B is a downstream target of FTO in As-exposed lung tissues. In addition, FTO protein is highly expressed and positively correlates with the protein levels of A3B in tumor samples from human non-small cell lung cancer patients. These findings indicate a previously unrecognized role of A3B in As-triggered somatic mutation and might open new avenues to reduce DNA mutagenesis by targeting the FTO/m6A axis.


Asunto(s)
Adenosina/análogos & derivados , Dioxigenasa FTO Dependiente de Alfa-Cetoglutarato , Arsénico , Carcinoma de Pulmón de Células no Pequeñas , Citidina Desaminasa , Neoplasias Pulmonares , Antígenos de Histocompatibilidad Menor , ARN Mensajero , Adenosina/genética , Adenosina/metabolismo , Dioxigenasa FTO Dependiente de Alfa-Cetoglutarato/genética , Dioxigenasa FTO Dependiente de Alfa-Cetoglutarato/metabolismo , Arsénico/toxicidad , Carcinoma de Pulmón de Células no Pequeñas/inducido químicamente , Carcinoma de Pulmón de Células no Pequeñas/genética , Citidina Desaminasa/genética , Citidina Desaminasa/metabolismo , Desmetilación/efectos de los fármacos , Humanos , Antígenos de Histocompatibilidad Menor/genética , Antígenos de Histocompatibilidad Menor/metabolismo , Mutagénesis , ARN Mensajero/genética , ARN Mensajero/metabolismo
14.
Acta Pharmacol Sin ; 44(10): 2004-2018, 2023 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-37225844

RESUMEN

Doxorubicin is a common chemotherapeutic agent in clinic, but myocardial toxicity limits its use. Fibroblast growth factor (FGF) 10, a multifunctional paracrine growth factor, plays diverse roles in embryonic and postnatal heart development as well as in cardiac regeneration and repair. In this study we investigated the role of FGF10 as a potential modulator of doxorubicin-induced cardiac cytotoxicity and the underlying molecular mechanisms. Fgf10+/- mice and an inducible dominant negative FGFR2b transgenic mouse model (Rosa26rtTA; tet(O)sFgfr2b) were used to determine the effect of Fgf10 hypomorph or blocking of endogenous FGFR2b ligands activity on doxorubicin-induced myocardial injury. Acute myocardial injury was induced by a single injection of doxorubicin (25 mg/kg, i.p.). Then cardiac function was evaluated using echocardiography, and DNA damage, oxidative stress and apoptosis in cardiac tissue were assessed. We showed that doxorubicin treatment markedly decreased the expression of FGFR2b ligands including FGF10 in cardiac tissue of wild type mice, whereas Fgf10+/- mice exhibited a greater degree of oxidative stress, DNA damage and apoptosis as compared with the Fgf10+/+ control. Pre-treatment with recombinant FGF10 protein significantly attenuated doxorubicin-induced oxidative stress, DNA damage and apoptosis both in doxorubicin-treated mice and in doxorubicin-treated HL-1 cells and NRCMs. We demonstrated that FGF10 protected against doxorubicin-induced myocardial toxicity via activation of FGFR2/Pleckstrin homology-like domain family A member 1 (PHLDA1)/Akt axis. Overall, our results unveil a potent protective effect of FGF10 against doxorubicin-induced myocardial injury and identify FGFR2b/PHLDA1/Akt axis as a potential therapeutic target for patients receiving doxorubicin treatment.


Asunto(s)
Factor 10 de Crecimiento de Fibroblastos , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos , Animales , Ratones , Doxorrubicina , Factor 10 de Crecimiento de Fibroblastos/metabolismo , Factores de Crecimiento de Fibroblastos/metabolismo , Ratones Transgénicos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/metabolismo , Transducción de Señal/fisiología , Factores de Transcripción
15.
Mol Cell ; 60(1): 21-34, 2015 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-26387737

RESUMEN

Mutations in the E3 ubiquitin ligase Parkin have been linked to familial Parkinson's disease. Parkin has also been implicated in mitosis through mechanisms that are unclear. Here we show that Parkin interacts with anaphase promoting complex/cyclosome (APC/C) coactivators Cdc20 and Cdh1 to mediate the degradation of several key mitotic regulators independent of APC/C. We demonstrate that ordered progression through mitosis is orchestrated by two distinct E3 ligases through the shared use of Cdc20 and Cdh1. Furthermore, Parkin is phosphorylated and activated by polo-like kinase 1 (Plk1) during mitosis. Parkin deficiency results in overexpression of its substrates, mitotic defects, genomic instability, and tumorigenesis. These results suggest that the Parkin-Cdc20/Cdh1 complex is an important regulator of mitosis.


Asunto(s)
Cadherinas/metabolismo , Proteínas Cdc20/metabolismo , Inestabilidad Genómica , Mitosis , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismo , Animales , Carcinogénesis/genética , Proteínas de Ciclo Celular/metabolismo , Células Cultivadas , Embrión de Mamíferos/citología , Fibroblastos/citología , Fibroblastos/metabolismo , Técnicas de Inactivación de Genes , Células HEK293 , Humanos , Ratones , Mutación , Fosforilación , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Quinasa Tipo Polo 1
16.
Nucleic Acids Res ; 49(19): 11224-11240, 2021 11 08.
Artículo en Inglés | MEDLINE | ID: mdl-34606619

RESUMEN

The human RecQ helicase BLM is involved in the DNA damage response, DNA metabolism, and genetic stability. Loss of function mutations in BLM cause the genetic instability/cancer predisposition syndrome Bloom syndrome. However, the molecular mechanism underlying the regulation of BLM in cancers remains largely elusive. Here, we demonstrate that the deubiquitinating enzyme USP37 interacts with BLM and that USP37 deubiquitinates and stabilizes BLM, thereby sustaining the DNA damage response (DDR). Mechanistically, DNA double-strand breaks (DSB) promotes ATM phosphorylation of USP37 and enhances the binding between USP37 and BLM. Moreover, knockdown of USP37 increases BLM polyubiquitination, accelerates its proteolysis, and impairs its function in DNA damage response. This leads to enhanced DNA damage and sensitizes breast cancer cells to DNA-damaging agents in both cell culture and in vivo mouse models. Collectively, our results establish a novel molecular mechanism for the USP37-BLM axis in regulating DSB repair with an important role in chemotherapy and radiotherapy response in human cancers.


Asunto(s)
Neoplasias de la Mama/genética , Reparación del ADN , Endopeptidasas/genética , Regulación Neoplásica de la Expresión Génica , RecQ Helicasas/genética , Animales , Proteínas de la Ataxia Telangiectasia Mutada/genética , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/mortalidad , Neoplasias de la Mama/patología , Línea Celular Tumoral , ADN/genética , ADN/metabolismo , Roturas del ADN de Doble Cadena , Replicación del ADN , Endopeptidasas/metabolismo , Femenino , Células HEK293 , Células HeLa , Humanos , Células MCF-7 , Ratones , Fosforilación , Unión Proteica , Estabilidad Proteica , Proteolisis , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , RecQ Helicasas/metabolismo , Análisis de Supervivencia , Ubiquitinación , Ensayos Antitumor por Modelo de Xenoinjerto
17.
Nucleic Acids Res ; 49(6): 3322-3337, 2021 04 06.
Artículo en Inglés | MEDLINE | ID: mdl-33704464

RESUMEN

RPA is a critical factor for DNA replication and replication stress response. Surprisingly, we found that chromatin RPA stability is tightly regulated. We report that the GDP/GTP exchange factor DOCK7 acts as a critical replication stress regulator to promote RPA stability on chromatin. DOCK7 is phosphorylated by ATR and then recruited by MDC1 to the chromatin and replication fork during replication stress. DOCK7-mediated Rac1/Cdc42 activation leads to the activation of PAK1, which subsequently phosphorylates RPA1 at S135 and T180 to stabilize chromatin-loaded RPA1 and ensure proper replication stress response. Moreover, DOCK7 is overexpressed in ovarian cancer and depleting DOCK7 sensitizes cancer cells to camptothecin. Taken together, our results highlight a novel role for DOCK7 in regulation of the replication stress response and highlight potential therapeutic targets to overcome chemoresistance in cancer.


Asunto(s)
Cromatina/metabolismo , Replicación del ADN , Proteínas Activadoras de GTPasa/fisiología , Factores de Intercambio de Guanina Nucleótido/fisiología , Proteína de Replicación A/metabolismo , Animales , Línea Celular Tumoral , Reparación del ADN , Femenino , Proteínas Activadoras de GTPasa/antagonistas & inhibidores , Proteínas Activadoras de GTPasa/metabolismo , Factores de Intercambio de Guanina Nucleótido/antagonistas & inhibidores , Factores de Intercambio de Guanina Nucleótido/metabolismo , Células HEK293 , Humanos , Ratones , Ratones Desnudos , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/metabolismo , Fosforilación , Proteolisis , Transducción de Señal , Estrés Fisiológico/genética , Proteína de Unión al GTP cdc42/metabolismo , Quinasas p21 Activadas/metabolismo , Proteína de Unión al GTP rac1/metabolismo
18.
Genes Dev ; 29(21): 2244-57, 2015 Nov 01.
Artículo en Inglés | MEDLINE | ID: mdl-26545811

RESUMEN

The von Hippel-Lindau tumor suppressor pVHL is an E3 ligase that targets hypoxia-inducible factors (HIFs). Mutation of VHL results in HIF up-regulation and contributes to processes related to tumor progression such as invasion, metastasis, and angiogenesis. However, very little is known with regard to post-transcriptional regulation of pVHL. Here we show that WD repeat and SOCS box-containing protein 1 (WSB1) is a negative regulator of pVHL through WSB1's E3 ligase activity. Mechanistically, WSB1 promotes pVHL ubiquitination and proteasomal degradation, thereby stabilizing HIF under both normoxic and hypoxic conditions. As a consequence, WSB1 up-regulates the expression of HIF-1α's target genes and promotes cancer invasion and metastasis through its effect on pVHL. Consistent with this, WSB1 protein level negatively correlates with pVHL level and metastasis-free survival in clinical samples. This work reveals a new mechanism of pVHL's regulation by which cancer acquires invasiveness and metastatic tendency.


Asunto(s)
Regulación Neoplásica de la Expresión Génica , Metástasis de la Neoplasia , Proteínas/metabolismo , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo , Hipoxia de la Célula , Línea Celular Tumoral , Movimiento Celular/genética , Células HEK293 , Células HT29 , Células HeLa , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Péptidos y Proteínas de Señalización Intracelular , Mutación , Invasividad Neoplásica/genética , Neoplasias/genética , Neoplasias/fisiopatología , Complejo de la Endopetidasa Proteasomal/metabolismo , Estabilidad Proteica , Estructura Terciaria de Proteína , Proteínas/genética , Ubiquitinación , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genética
19.
Mol Cell ; 56(5): 681-95, 2014 Dec 04.
Artículo en Inglés | MEDLINE | ID: mdl-25454945

RESUMEN

DNA replication is executed only when cells have sufficient metabolic resources and undamaged DNA. Nutrient limitation and DNA damage cause a metabolic checkpoint and DNA damage checkpoint, respectively. Although SIRT1 activity is regulated by metabolic stress and DNA damage, its function in these stress-mediated checkpoints remains elusive. Here we report that the SIRT1-TopBP1 axis functions as a switch for both checkpoints. With glucose deprivation, SIRT1 is activated and deacetylates TopBP1, resulting in TopBP1-Treslin disassociation and DNA replication inhibition. Conversely, SIRT1 activity is inhibited under genotoxic stress, resulting in increased TopBP1 acetylation that is important for the TopBP1-Rad9 interaction and activation of the ATR-Chk1 pathway. Mechanistically, we showed that acetylation of TopBP1 changes the conformation of TopBP1, thereby facilitating its interaction with distinct partners in DNA replication and checkpoint activation. Taken together, our studies identify the SIRT1-TopBP1 axis as a key signaling mode in the regulation of the metabolic checkpoint and the DNA damage checkpoint.


Asunto(s)
Proteínas Portadoras/metabolismo , Daño del ADN , Sirtuina 1/metabolismo , Estrés Fisiológico , Acetilación , Animales , Puntos de Control del Ciclo Celular , Proteínas de Ciclo Celular/metabolismo , Línea Celular , Reparación del ADN , Replicación del ADN , Células HEK293 , Humanos , Ratones , Conformación Proteica , Transducción de Señal
20.
Nucleic Acids Res ; 48(22): 12711-12726, 2020 12 16.
Artículo en Inglés | MEDLINE | ID: mdl-33237263

RESUMEN

PrimPol has been recently identified as a DNA damage tolerant polymerase that plays an important role in replication stress response. However, the regulatory mechanisms of PrimPol are not well defined. In this study, we identify that the deubiquitinase USP36 interferes with degradation of PrimPol to regulate the replication stress response. Mechanistically, USP36 is deubiquitinated following DNA replication stress, which in turn facilitates its upregulation and interaction with PrimPol. USP36 deubiquitinates K29-linked polyubiquitination of PrimPol and increases its protein stability. Depletion of USP36 results in replication stress-related defects and elevates cell sensitivity to DNA-damage agents, such as cisplatin and olaparib. Moreover, USP36 expression positively correlates with the level of PrimPol protein and poor prognosis in patient samples. These findings indicate that the regulation of PrimPol K29-linked ubiquitination by USP36 plays a critical role in DNA replication stress and chemotherapy response.


Asunto(s)
ADN Primasa/genética , Replicación del ADN/efectos de los fármacos , ADN Polimerasa Dirigida por ADN/genética , Enzimas Multifuncionales/genética , Neoplasias Ováricas/genética , Ubiquitina Tiolesterasa/genética , Línea Celular Tumoral , Cisplatino/farmacología , Daño del ADN/efectos de los fármacos , Enzimas Desubicuitinizantes/genética , Resistencia a Antineoplásicos/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/patología , Ftalazinas/farmacología , Piperazinas/farmacología , Poliubiquitina/genética , Pronóstico , Estabilidad Proteica/efectos de los fármacos , Proteolisis/efectos de los fármacos
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