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1.
J Exp Med ; 217(3)2020 Mar 02.
Artigo em Inglês | MEDLINE | ID: mdl-31961917

RESUMO

Cancer cells often proliferate under hypoxia and reprogram their metabolism. However, how to find targets to effectively block the hypoxia-associated metabolic pathways remains unclear. Here, we developed a tool to conveniently calculate electrons dissipated in metabolic transformations. Based on the law of conservation of electrons in chemical reactions, we further built up an electron balance model for central carbon metabolism, and it can accurately outline metabolic plasticity under hypoxia. Our model specifies that glutamine metabolism reprogrammed for biosynthesis of lipid and/or proline actually acts as the alternative electron bin to enable electron transfer in proliferating cells under hypoxia. Inhibition of both proline biosynthesis and lipogenesis can synergistically suppress cancer cell growth under hypoxia and in vivo tumor onset. Therefore, our model helps to reveal combinations of potential targets to inhibit tumor growth by blocking hypoxia-rewired metabolism and provides a useful tool for future studies on cancer metabolism.

2.
Proc Natl Acad Sci U S A ; 116(51): 25624-25633, 2019 Dec 17.
Artigo em Inglês | MEDLINE | ID: mdl-31796584

RESUMO

The translesion synthesis (TLS) pathway is a double-edged sword in terms of genome integrity. Deficiency in TLS leads to generation of DNA double strand break (DSB) during replication stress, while excessive activation of the TLS pathway increases the risk of point mutation. Here we demonstrate that HSCARG, a cellular redox sensor, directly interacts with the key protein PCNA in the TLS pathway. HSCARG enhances the interaction between PCNA and the deubiquitinase complex USP1/UAF1 and inhibits the monoubiquitination of PCNA, thereby impairing the recruitment of Y-family polymerases and increasing cell sensitivity to stimuli that trigger replication fork blockades. In response to oxidative stress, disaggregation of HSCARG dimers into monomers and the nuclear transport of HSCARG activate the regulatory function of HSCARG in the TLS pathway. Moreover, HSCARG, which is highly expressed in breast carcinoma, promotes the accumulation of DSBs and mutations. HSCARG knockout PyMT transgenic mice exhibit delayed mammary tumorigenesis compared with that in HSCARG wild-type or heterozygous PyMT mice. Taken together, these findings expand our understanding of TLS regulatory mechanisms and establish a link between the cellular redox status and the DNA damage response (DDR).

3.
Cancer Res ; 2019 Dec 24.
Artigo em Inglês | MEDLINE | ID: mdl-31874856

RESUMO

The DNA damage response (DDR) is essential for maintaining genome integrity. Mounting evidence reveals that protein modifications play vital roles in the DDR. Here, we show that USP38 is involved in the DDR by regulating the activity of HDAC1. In response to DNA damage, USP38 interacted with HDAC1 and specifically removed the K63-linked ubiquitin chain promoting the deacetylase activity of HDAC1. As a result, HDAC1 was able to deacetylate H3K56. USP38 deletion resulted in persistent focal accumulation of NHEJ factors at DNA damage sites and impaired NHEJ efficiency, causing genome instability and sensitizing cancer cells to genotoxic insults. Knockout of USP38 rendered mice hypersensitive to IR and shortened survival. In addition, USP38 was expressed at low levels in certain types of cancers including renal cell carcinoma, indicating dysregulation of USP38 expression contributes to genomic instability and may lead to tumorigenesis. In summary, this study identifies a critical role of USP38 in modulating genome integrity and cancer cell resistance to genotoxic insults by deubiquitinating HDAC1 and regulating its deacetylation activity.

4.
Nat Commun ; 10(1): 201, 2019 01 14.
Artigo em Inglês | MEDLINE | ID: mdl-30643150

RESUMO

Under hypoxia, most of glucose is converted to secretory lactate, which leads to the overuse of glutamine-carbon. However, under such a condition how glutamine nitrogen is disposed to avoid over-accumulating ammonia remains to be determined. Here we identify a metabolic flux of glutamine to secretory dihydroorotate, which is indispensable to glutamine-carbon metabolism under hypoxia. We found that glutamine nitrogen is necessary to nucleotide biosynthesis, but enriched in dihyroorotate and orotate rather than processing to its downstream uridine monophosphate under hypoxia. Dihyroorotate, not orotate, is then secreted out of cells. Furthermore, we found that the specific metabolic pathway occurs in vivo and is required for tumor growth. The identified metabolic pathway renders glutamine mainly to acetyl coenzyme A for lipogenesis, with the rest carbon and nitrogen being safely removed. Therefore, our results reveal how glutamine carbon and nitrogen are coordinatively metabolized under hypoxia, and provide a comprehensive understanding on glutamine metabolism.


Assuntos
Glutamina/metabolismo , Redes e Vias Metabólicas , Metaboloma , Neoplasias/metabolismo , Ácido Orótico/análogos & derivados , Acetilcoenzima A/metabolismo , Amônia/metabolismo , Amônia/toxicidade , Animais , Carbono/química , Carbono/metabolismo , Hipóxia Celular , Linhagem Celular Tumoral , Sobrevivência Celular , Feminino , Glucose/metabolismo , Glutamina/química , Células HEK293 , Humanos , Ácido Láctico/metabolismo , Lipogênese , Metabolômica , Camundongos , Camundongos Nus , Neoplasias/sangue , Neoplasias/mortalidade , Neoplasias/patologia , Nitrogênio/química , Nitrogênio/metabolismo , Nucleotídeos/biossíntese , Ácido Orótico/metabolismo , Microambiente Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto
5.
J Biol Chem ; 294(8): 2827-2838, 2019 02 22.
Artigo em Inglês | MEDLINE | ID: mdl-30598506

RESUMO

Ribosomal proteins are the building blocks of ribosome biogenesis. Beyond their known participation in ribosome assembly, the ribosome-independent functions of ribosomal proteins are largely unknown. Here, using immunoprecipitation, subcellular fractionation, His-ubiquitin pulldown, and immunofluorescence microscopy assays, along with siRNA-based knockdown approaches, we demonstrate that ribosomal protein L6 (RPL6) directly interacts with histone H2A and is involved in the DNA damage response (DDR). We found that in response to DNA damage, RPL6 is recruited to DNA damage sites in a poly(ADP-ribose) polymerase (PARP)-dependent manner, promoting its interaction with H2A. We also observed that RPL6 depletion attenuates the interaction between mediator of DNA damage checkpoint 1 (MDC1) and H2A histone family member X, phosphorylated (γH2AX), impairs the accumulation of MDC1 at DNA damage sites, and reduces both the recruitment of ring finger protein 168 (RNF168) and H2A Lys-15 ubiquitination (H2AK15ub). These RPL6 depletion-induced events subsequently inhibited the recruitment of the following downstream repair proteins: tumor protein P53-binding protein 1 (TP53BP1) and BRCA1, DNA repair-associated (BRCA1). Moreover, the RPL6 knockdown resulted in defects in the DNA damage-induced G2-M checkpoint, DNA damage repair, and cell survival. In conclusion, our study identifies RPL6 as a critical regulatory factor involved in the DDR. These findings expand our knowledge of the extraribosomal functions of ribosomal proteins in cell physiology and deepen our understanding of the molecular mechanisms underlying DDR regulation.


Assuntos
Proteína BRCA1/metabolismo , Dano ao DNA , Reparo do DNA , Histonas/metabolismo , Poli(ADP-Ribose) Polimerase-1/metabolismo , Proteínas Ribossômicas/metabolismo , Proteína BRCA1/genética , Ciclo Celular , Sobrevivência Celular , Células HEK293 , Células HeLa , Histonas/genética , Humanos , Poli(ADP-Ribose) Polimerase-1/genética , Proteínas Ribossômicas/genética , Transdução de Sinais , Ubiquitina , Ubiquitinação
6.
Sci Total Environ ; 639: 617-623, 2018 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-29803035

RESUMO

Horizontal transfer of ARGs was generally considered to be mediated by three methods - transformation, conjugation and transduction through phages - during which the contribution of bacteriophages to gene transfer in the environment is unclear or even questioned. In this study, a multiple-antibiotic-resistant Escherichia coli strain and its phage (YZ1) were isolated from a municipal wastewater treatment system. The results of the morphological and genomic analyses of phage YZ1 showed that it is a member of the T7 viral genus in the subfamily Autographivirinae. Its genome is similar to that of the E. coli phage K1F in both organization and sequence and does not encode ARGs. However, 28 paired reads in the raw sequencing data aligned to ARGs, including those promoting ß-lactam, aminoglycoside, and fluoroquinolone resistance, among others. Quantitative PCR showed that ARGs were present in bacteriophage DNA (approximately 103 copies/mL) and were also detected in the bacterial host DNA. The results suggested that while infrequent, some ARG-carrying transducing phages were presumably generated by erroneous packaging during infection of antibiotic-resistant bacteria, which may create the possibility of horizontal transfer of ARGs.


Assuntos
Bacteriófagos/crescimento & desenvolvimento , Farmacorresistência Bacteriana/genética , Escherichia coli/virologia , Genes Bacterianos , Antibacterianos , Bactérias , Escherichia coli/isolamento & purificação , Transferência Genética Horizontal , Águas Residuárias/microbiologia , Águas Residuárias/virologia
7.
Cancer Res ; 78(4): 1069-1082, 2018 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-29233925

RESUMO

A competent DNA damage response (DDR) helps prevent cancer, but once cancer has arisen, DDR can blunt the efficacy of chemotherapy and radiotherapy that cause lethal DNA breakage in cancer cells. Thus, blocking DDR may improve the efficacy of these modalities. Here, we report a new DDR mechanism that interfaces with inflammatory signaling and might be blocked to improve anticancer outcomes. Specifically, we report that the ubiquitin-editing enzyme A20/TNFAIP3 binds and inhibits the E3 ubiquitin ligase RNF168, which is responsible for regulating histone H2A turnover critical for proper DNA repair. A20 induced after DNA damage disrupted RNF168-H2A interaction in a manner independent of its enzymatic activity. Furthermore, it inhibited accumulation of RNF168 and downstream repair protein 53BP1 during DNA repair. A20 was also required for disassembly of RNF168 and 53BP1 from damage sites after repair. Conversely, A20 deletion increased the efficiency of error-prone nonhomologous DNA end-joining and decreased error-free DNA homologous recombination, destablizing the genome and increasing sensitivity to DNA damage. In clinical specimens of invasive breast carcinoma, A20 was widely overexpressed, consistent with its candidacy as a therapeutic target. Taken together, our findings suggest that A20 is critical for proper functioning of the DDR in cancer cells and it establishes a new link between this NFκB-regulated ubiquitin-editing enzyme and the DDR pathway.Significance: This study identifies the ubiquitin-editing enzyme A20 as a key factor in mediating cancer cell resistance to DNA-damaging therapy, with implications for blocking its function to leverage the efficacy of chemotherapy and radiotherapy. Cancer Res; 78(4); 1069-82. ©2017 AACR.


Assuntos
Dano ao DNA , Neoplasias/tratamento farmacológico , Neoplasias/genética , Proteína 3 Induzida por Fator de Necrose Tumoral alfa/metabolismo , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Etoposídeo/farmacologia , Células HEK293 , Células HeLa , Humanos , Células MCF-7 , NF-kappa B/genética , NF-kappa B/metabolismo , Neoplasias/metabolismo , Regiões Promotoras Genéticas , Transdução de Sinais , Transfecção , Proteína 3 Induzida por Fator de Necrose Tumoral alfa/genética , Enzimas de Conjugação de Ubiquitina/genética , Enzimas de Conjugação de Ubiquitina/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação , Regulação para Cima
9.
Nat Commun ; 8: 15308, 2017 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-28516914

RESUMO

Targeting the specific metabolic phenotypes of colorectal cancer stem cells (CRCSCs) is an innovative therapeutic strategy for colorectal cancer (CRC) patients with poor prognosis and relapse. However, the context-dependent metabolic traits of CRCSCs remain poorly elucidated. Here we report that adenylate kinase hCINAP is overexpressed in CRC tissues. Depletion of hCINAP inhibits invasion, self-renewal, tumorigenesis and chemoresistance of CRCSCs with a loss of mesenchymal signature. Mechanistically, hCINAP binds to the C-terminal domain of LDHA, the key regulator of glycolysis, and depends on its adenylate kinase activity to promote LDHA phosphorylation at tyrosine 10, resulting in the hyperactive Warburg effect and the lower cellular ROS level and conferring metabolic advantage to CRCSC invasion. Moreover, hCINAP expression is positively correlated with the level of Y10-phosphorylated LDHA in CRC patients. This study identifies hCINAP as a potent modulator of metabolic reprogramming in CRCSCs and a promising drug target for CRC invasion and metastasis.


Assuntos
Neoplasias Colorretais/genética , Regulação Neoplásica da Expressão Gênica , L-Lactato Desidrogenase/genética , Células-Tronco Neoplásicas/metabolismo , Proteínas Nucleares/genética , Processamento de Proteína Pós-Traducional , Animais , Antineoplásicos/farmacologia , Carcinogênese/genética , Carcinogênese/metabolismo , Carcinogênese/patologia , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Proteínas de Ligação a DNA , Doxiciclina/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Glicólise/efeitos dos fármacos , Glicólise/genética , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , L-Lactato Desidrogenase/metabolismo , Camundongos , Camundongos Nus , Invasividade Neoplásica , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologia , Proteínas Nucleares/antagonistas & inibidores , Proteínas Nucleares/metabolismo , Fenótipo , Fosforilação , Ligação Proteica , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Tirosina/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Chemistry ; 16(12): 3763-9, 2010 Mar 22.
Artigo em Inglês | MEDLINE | ID: mdl-20157906

RESUMO

The monoammoniate of lithium amidoborane, Li(NH(3))NH(2)BH(3), was synthesized by treatment of LiNH(2)BH(3) with ammonia at room temperature. This compound exists in the amorphous state at room temperature, but at -20 degrees C crystallizes in the orthorhombic space group Pbca with lattice parameters of a = 9.711(4), b = 8.7027(5), c = 7.1999(1) A, and V = 608.51 A(3). The thermal decomposition behavior of this compound under argon and under ammonia was investigated. Through a series of experiments we have demonstrated that Li(NH(3))NH(2)BH(3) is able to absorb/desorb ammonia reversibly at room temperature. In the temperature range of 40-70 degrees C, this compound showed favorable dehydrogenation characteristics. Specifically, under ammonia this material was able to release 3.0 equiv hydrogen (11.18 wt %) rapidly at 60 degrees C, which represents a significant advantage over LiNH(2)BH(3). It has been found that the formation of the coordination bond between ammonia and Li(+) in LiNH(2)BH(3) plays a crucial role in promoting the combination of hydridic B-H bonds and protic N-H bonds, leading to dehydrogenation at low temperature.

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