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1.
Sci Transl Med ; 14(626): eabf0992, 2022 01 05.
Artigo em Inglês | MEDLINE | ID: mdl-34985967

RESUMO

High CD8+ T cell infiltration in colorectal cancer (CRC) should suggest a favorable prognosis and a satisfactory response to immunotherapy; however, the vast majority of patients with CRC do not benefit from immunotherapy due to poor T cell infiltration. Therefore, a better understanding of the mechanisms for T cell exclusion from CRC tumors is needed. Tribbles homolog 3 (TRIB3) has been implicated as an oncoprotein, but its role in regulating antitumor immune responses has not been defined. Here, we demonstrated that TRIB3 inhibits CD8+ T cell infiltration in various CRC mouse models. We showed that TRIB3 was acetylated by acetyltransferase P300, which inhibited ubiquitination and subsequent proteasomal degradation of TRIB3. Ectopically expressed TRIB3 inhibited signal transducer and activator of transcription 1 (STAT1) activation and STAT1-mediated CXCL10 transcription by enhancing the epidermal growth factor receptor signaling pathway, causing a reduction in tumor-infiltrating T cells. Genetic ablation of Trib3 or pharmacological acceleration of TRIB3 degradation with a P300 inhibitor increased T cell recruitment and sensitized CRCs to immune checkpoint blockade therapy. These findings identified TRIB3 as a negative modulator of CD8+ T cell infiltration in CRCs, highlighting a potential therapeutic target for treating immunologically "cold" CRCs.


Assuntos
Proteínas de Ciclo Celular , Neoplasias Colorretais , Evasão da Resposta Imune , Proteínas Repressoras , Animais , Linfócitos T CD8-Positivos , Proteínas de Ciclo Celular/metabolismo , Quimiocina CXCL10/metabolismo , Neoplasias Colorretais/patologia , Humanos , Imunoterapia , Camundongos , Proteínas Repressoras/metabolismo , Fator de Transcrição STAT1/metabolismo , Transdução de Sinais
3.
Immunity ; 54(9): 2042-2056.e8, 2021 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-34407391

RESUMO

Recruitment of immune cells to the site of inflammation by the chemokine CCL1 is important in the pathology of inflammatory diseases. Here, we examined the role of CCL1 in pulmonary fibrosis (PF). Bronchoalveolar lavage fluid from PF mouse models contained high amounts of CCL1, as did lung biopsies from PF patients. Immunofluorescence analyses revealed that alveolar macrophages and CD4+ T cells were major producers of CCL1 and targeted deletion of Ccl1 in these cells blunted pathology. Deletion of the CCL1 receptor Ccr8 in fibroblasts limited migration, but not activation, in response to CCL1. Mass spectrometry analyses of CCL1 complexes identified AMFR as a CCL1 receptor, and deletion of Amfr impaired fibroblast activation. Mechanistically, CCL1 binding triggered ubiquitination of the ERK inhibitor Spry1 by AMFR, thus activating Ras-mediated profibrotic protein synthesis. Antibody blockade of CCL1 ameliorated PF pathology, supporting the therapeutic potential of targeting this pathway for treating fibroproliferative lung diseases.


Assuntos
Quimiocina CCL1/metabolismo , Fibroblastos/metabolismo , Proteínas de Membrana/metabolismo , Miofibroblastos/metabolismo , Fosfoproteínas/metabolismo , Fibrose Pulmonar/metabolismo , Receptores do Fator Autócrino de Motilidade/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Diferenciação Celular/fisiologia , Fibroblastos/patologia , Humanos , Camundongos , Miofibroblastos/patologia , Fibrose Pulmonar/patologia , Transdução de Sinais/fisiologia
4.
Sci Transl Med ; 13(586)2021 03 24.
Artigo em Inglês | MEDLINE | ID: mdl-33762435

RESUMO

Most basal-like breast cancers (BLBCs) are triple-negative breast cancers (TNBCs), which have the worst prognosis and distant metastasis-free survival among breast cancer subtypes. Now, no targeted therapies are available for patients with BLBC due to the lack of reliable and effective molecular targets. Here, we performed the BLBC tissue microarray-based immunohistochemical analysis and showed that Faciogenital Dysplasia 5 (FGD5) abundance is associated with poor prognosis in BLBCs. FGD5 deletion decreased the proliferation, invasion, and tumorsphere formation capacity of BLBC cells. Furthermore, genetic inhibition of Fgd5 in mouse mammary epithelial cells attenuated BLBC initiation and progression by reducing the self-renewal ability of tumor-initiating cells. In addition, FGD5 abundance was positively correlated with the abundance of epidermal growth factor receptor (EGFR) in BLBCs. FGD5 ablation decreased EGFR abundance by reducing EGFR stability in TNBC cells in 2D and 3D culture conditions. Mechanistically, FGD5 binds to EGFR and interferes with basal EGFR ubiquitination and degradation induced by the E3 ligase ITCH. Impaired EGFR degradation caused BLBC cell proliferation and promoted invasive properties and self-renewal. To verify the role of the FGD5-EGFR interaction in the regulation of EGFR stability, we screened a cell-penetrating α-helical peptide PER3 binding with FGD5 to disrupt the interaction. Treatment of BLBC patient-derived xenograft-bearing mice with the peptide PER3 disrupting the FGD5-EGFR interaction either with or without chemotherapy reduced BLBC progression. Our study identified FGD5 as a positive modulator of tumor-initiating cells and suggests a potential therapeutic option for the BLBC subtype of breast cancer.


Assuntos
Fatores de Troca do Nucleotídeo Guanina/genética , Células-Tronco Neoplásicas , Neoplasias de Mama Triplo Negativas , Animais , Receptores ErbB , Feminino , Humanos , Camundongos , Neoplasias de Mama Triplo Negativas/genética
5.
BMC Surg ; 21(1): 103, 2021 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-33632188

RESUMO

BACKGROUNDS: Pilocytic astrocytomas (PAs) are World Health Organization (WHO) grade I tumors, which are relatively common, and are benign lesions in children. PAs could originate from the cerebellum, optic pathways, and third ventricular/hypothalamic region. Traditional various transcranial routes are used for hypothalamic PAs (HPAs). However, there are few studies on hypothalamic PAs treated through the endoscopic endonasal approach (EEA). This study reports the preliminary experience of the investigators and results with HPAs via expanded EEAs. METHODS: All patients with HPAs, undergone EEA in our hospital from 2017 to 2019, were retrospectively reviewed. The demographic data, clinical symptoms, complications, skull base reconstruction, prognosis, and endocrinological data were all recorded and analyzed in detail. RESULTS: Finally, five female patients were enrolled. The average age of patients was 28.6 ± 14.0. All patients had complaints about their menstrual disorder. One patient had severe bilateral visual impairment. Furthermore, only one patient suffered from severe headache due to acute hydrocephalus, although there were four patients with headache or dizziness. Four cases achieved gross-total resection, and one patient achieved subtotal resection. Furthermore, there was visual improvement in one patient (case 5), and postoperative worsening of vision in one patient (case 4). However, only one patient had postoperative intracranial infection. None of the patients experienced a postoperative CSF leak, and in situ bone flap (ISBF) techniques were used for two cases for skull base repair. In particular, ISBF combined with free middle turbinate mucosal flap was used for case 5. After three years of follow-up, three patients are still alive, two patients had no neurological or visual symptoms, or tumor recurrence, and one patient had severe hypothalamic dysfunction. Unfortunately, one patient died of severe postoperative hypothalamus reaction, which presented with coma, high fever, diabetes insipidus, hypernatremia and intracranial infection. The other patient died of recurrent severe pancreatitis at one year after the operation. CONCLUSION: Although the data is still very limited and preliminary, EEA provides a direct approach to HPAs with acceptable prognosis in terms of tumor resection, endocrinological and visual outcomes. ISBF technique is safe and reliable for skull base reconstruction.


Assuntos
Astrocitoma , Hipotálamo , Cirurgia Endoscópica por Orifício Natural , Adulto , Astrocitoma/cirurgia , Feminino , Humanos , Hipotálamo/cirurgia , Estudos Retrospectivos , Resultado do Tratamento
6.
Nat Commun ; 11(1): 6316, 2020 12 09.
Artigo em Inglês | MEDLINE | ID: mdl-33298911

RESUMO

The transcription factor MYC is deregulated in almost all human cancers, especially in aggressive lymphomas, through chromosomal translocation, amplification, and transcription hyperactivation. Here, we report that high expression of tribbles homologue 3 (TRIB3) positively correlates with elevated MYC expression in lymphoma specimens; TRIB3 deletion attenuates the initiation and progression of MYC-driven lymphoma by reducing MYC expression. Mechanistically, TRIB3 interacts with MYC to suppress E3 ubiquitin ligase UBE3B-mediated MYC ubiquitination and degradation, which enhances MYC transcriptional activity, causing high proliferation and self-renewal of lymphoma cells. Use of a peptide to disturb the TRIB3-MYC interaction together with doxorubicin reduces the tumor burden in MycEµ mice and patient-derived xenografts. The pathophysiological relevance of UBE3B, TRIB3 and MYC is further demonstrated in human lymphoma. Our study highlights a key mechanism for controlling MYC expression and a potential therapeutic option for treating lymphomas with high TRIB3-MYC expression.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Linfoma não Hodgkin/patologia , Proteínas Proto-Oncogênicas c-myc/metabolismo , Proteínas Repressoras/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Autorrenovação Celular/efeitos dos fármacos , Autorrenovação Celular/genética , Sequenciamento de Cromatina por Imunoprecipitação , Progressão da Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Introdução de Genes , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , Linfoma não Hodgkin/tratamento farmacológico , Linfoma não Hodgkin/genética , Masculino , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Cultura Primária de Células , Complexo de Endopeptidases do Proteassoma/metabolismo , Inibidores de Proteassoma/farmacologia , /metabolismo , Proteólise/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-myc/antagonistas & inibidores , RNA-Seq , Proteínas Repressoras/genética , Ubiquitinação/efeitos dos fármacos , Ubiquitinação/genética , Ensaios Antitumorais Modelo de Xenoenxerto , Adulto Jovem
7.
J Food Sci ; 85(8): 2452-2460, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32691480

RESUMO

Owing to the strong hydrophobicity of zein, improved solubility is required to enhance the recovery of bioactive peptides. Using a zein suspension prepared by the antisolvent precipitation method, the impact of varying the voltage during dielectric barrier discharge (DBD) treatment on the physicochemical and conformational properties of zein in water was investigated. Analysis of the particle size, specific surface area, and free sulfhydryl content indicated that the protein solubility was maximized by treatment at 70 V for 70 s. DBD treatment destroyed covalent bonds and introduced some hydrophilic groups onto the zein surface, thus enhancing the contact area with water molecules and leading to a more uniform dispersion. A decrease in the hydrodynamic radius of zein micelles indicated that intermolecular interactions were disrupted, thus improving dispersion stability. A more hydrophilic microenvironment was formed owing to the reduction in hydrophobic interactions. Additionally, evaluation of the secondary structure demonstrated that DBD treatment broke hydrogen bonds, resulting in a loose conformation with more exposed sites of action for water. These results are expected to facilitate the development of technologies for improving utilization of zein. PRACTICAL APPLICATION: Strong hydrophobicity limits the application of zein in the food industry. The study indicated that DBD treatment could promote loose structure, and improve dispersion stability and hydrophilicity of zein suspension prepared by antisolvent precipitation method. This work revealed the potential of cold plasma treatment for modifying zein and other insoluble proteins, which would expand their scope of application.


Assuntos
Tecnologia de Alimentos/métodos , Zeína/química , Fenômenos Químicos , Eletricidade , Ligação de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Nanopartículas/química , Tamanho da Partícula , Conformação Proteica , Solubilidade , Compostos de Sulfidrila/análise , Suspensões/química , Água/química
8.
Nat Commun ; 11(1): 3660, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32694521

RESUMO

High expression or aberrant activation of epidermal growth factor receptor (EGFR) is related to tumor progression and therapy resistance across cancer types, including non-small cell lung cancer (NSCLC). EGFR tyrosine kinase inhibitors (TKIs) are first-line therapy for NSCLC. However, patients eventually deteriorate after inevitable acquisition of EGFR TKI-resistant mutations, highlighting the need for therapeutics with alternative mechanisms of action. Here, we report that the elevated tribbles pseudokinase 3 (TRIB3) is positively associated with EGFR stability and NSCLC progression. TRIB3 interacts with EGFR and recruits PKCα to induce a Thr654 phosphorylation and WWP1-induced Lys689 ubiquitination in the EGFR juxtamembrane region, which enhances EGFR recycling, stability, downstream activity, and NSCLC stemness. Disturbing the TRIB3-EGFR interaction with a stapled peptide attenuates NSCLC progression by accelerating EGFR degradation and sensitizes NSCLC cells to chemotherapeutic agents. These findings indicate that targeting EGFR degradation is a previously unappreciated therapeutic option in EGFR-related NSCLC.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Proteínas de Ciclo Celular/metabolismo , Neoplasias Pulmonares/patologia , Proteínas Repressoras/metabolismo , Adulto , Animais , Antineoplásicos/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Linhagem Celular Tumoral , Progressão da Doença , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Receptores ErbB/metabolismo , Feminino , Humanos , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/mortalidade , Masculino , Camundongos , Pessoa de Meia-Idade , Mutação , Fosforilação/efeitos dos fármacos , Proteína Quinase C-alfa/metabolismo , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Estabilidade Proteica/efeitos dos fármacos , Proteólise/efeitos dos fármacos , Taxa de Sobrevida , Ubiquitinação/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
9.
Comp Immunol Microbiol Infect Dis ; 72: 101514, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32634650

RESUMO

Hepatozoon canis, transmitted by Rhipicephalus sanguineus, is a tick-borne pathogen and causes canine hepatozoonosis. Until now, only limited previous studies were conducted on the molecular detection and characterization of Hepatozoon sp. in dogs in China. Blood samples were collected from 93 sick dogs that were clinically diagnosed as babesiosis but tested negative for Babesia, and 103 apparently healthy dogs, as well as their infesting ticks in Xi'an and Hanzhong cities, Shaanxi province of China. PCR amplifying partial 18S rRNA gene was used to detect the DNA of Hepatozoon sp. Genetic and phylogenetic analysis were performed to determine the Hepatozoon species. Our results demonstrated that H. canis was identified from the sick dogs and the infested ticks in Hanzhong, with no significant differences of prevalence between both genders and ages. No positive blood or tick samples were found in Xi'an. Moreover, all the 18S rRNA gene sequences recovered from both dogs and the infested ticks showed a high genetic similarity with each other, and also presented a close relationship with other known sequences in and outside China. In conclusion, H. canis was identified in babesiosis-suspected dogs and ticks infesting them in Shaanxi, China, although the association between clinical signs and H. canis need further study.


Assuntos
Coccidiose/veterinária , Doenças do Cão , Eucoccidiida , Animais , China/epidemiologia , Coccidiose/epidemiologia , Doenças do Cão/diagnóstico , Doenças do Cão/epidemiologia , Doenças do Cão/parasitologia , Cães , Eucoccidiida/genética , Eucoccidiida/isolamento & purificação , Feminino , Masculino , Filogenia
10.
Int J Biol Macromol ; 2020 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-32371128

RESUMO

Cold plasma (CP) treatment was used to prepare the glycosylation conjugates of high-temperature peanut protein isolate (HPPI) and lactose to improve the solubility of HPPI. We observed that by increasing the CP treatment time to 3 min, the solubility of the conjugates increased to 1.34 mg/mL. An increase in the degree of glycosylation and a decrease in the degree of browning indicated that although CP treatment accelerated the glycosylation of HPPI and lactose, it interfered with the formation of melanoidin. The analysis of protein tertiary structure showed that tryptophan and tyrosine residues in proteins undergoing CP treatment were the primary sites for the Maillard reaction. The relative decrease in surface hydrophobicity and FT-IR analysis indicated that the increase in the -OH stretching vibration intensity on the protein surface represented the formation of the covalent bonds between HPPI and lactose during the CP treatment. An increase in the denaturation temperature of proteins was observed after grafting with lactose. Changes in the secondary structure and surface structure of proteins showed that lactose covalently bonded to the surface of HPPI during CP treatment, forming a more stable ordered structure.

11.
Cancer Lett ; 474: 23-35, 2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-31931029

RESUMO

Despite the success of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in the treatment of non-small cell lung cancer (NSCLC) harboring EGFR-activating mutations, intrinsic or acquired resistance remains the major obstacle to long-term disease remission. Defective autophagy has been reported as an EGFR-TKI resistance mechanism. However, how EGFR regulate autophagic flux are still not fully understood. Here we found that EGFR-stimulated phosphorylation of SQSTM1 at tyrosine 433 induces dimerization of its UBA domain, which disturbs the sequestration function of SQSTM1 and causes autophagic flux blocking. SAH-EJ2, a staple optimized EGFR-derived peptide, showed enhanced in vitro and in vivo antitumor activity against NSCLC than the prototype regardless of EGFR mutation status. Mechanistically, SAH-EJ2 disrupts the EGFR-SQSTM1 interaction and protects against EGFR-induced SQSTM1 phosphorylation, which hinders the dimerization of the SQSTM1 UBA domains and restores SQSTM1 cargo function. Moreover, SAH-EJ2 suppresses EGFR activity by blocking its dimerization and reducing its protein stability, which reciprocally activates the core autophagy machinery. Our observations reveal that disturbing the EGFR-SQSTM1 interaction by SAH-EJ2 confers a potential strategy in the treatment of NSCLC through suppressing EGFR signalling and activating autophagy simultaneously.


Assuntos
Autofagia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/tratamento farmacológico , Fragmentos de Peptídeos/farmacologia , Proteína Sequestossoma-1/antagonistas & inibidores , Animais , Antineoplásicos/farmacologia , Apoptose , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Proliferação de Células , Resistencia a Medicamentos Antineoplásicos , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Receptores ErbB/metabolismo , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Mutação , Inibidores de Proteínas Quinases/farmacologia , Multimerização Proteica , Proteína Sequestossoma-1/metabolismo , Transdução de Sinais , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
12.
Cancer Med ; 9(3): 1131-1140, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31823521

RESUMO

Epidermal growth factor receptor (EGFR) is highly expressed in head and neck squamous cell carcinoma (HNSCC) and correlates with poor prognosis. EGFR has been demonstrated to be associated with cancer stem cell traits in HNSCC. However, the underlying molecular mechanism is far from elucidated. Here, SOX2, one of the most important stem cell markers, was identified as a binding partner and substrate of EGFR. EGFR signaling inhibition decreases SOX2 expression by promoting its autophagic degradation. Mechanistically, EGFR activation induces SOX2 phosphorylation at the Y277 site and reduces its ubiquitination, which inhibits its association with p62 and subsequent autophagic degradation. Gefitinib, an EGFR tyrosine kinase inhibitor, shows in vitro and in vivo protective effects against oral cancer cells that can be reversed through autophagy inhibition. Our study suggests that EGFR plays an important role in the development of cancer stem cells by stabilizing SOX2. Targeting EGFR in combination with conventional chemotherapy might be a promising strategy for the treatment of HNSCC through elimination of cancer stem cells.


Assuntos
Autofagia/fisiologia , Gefitinibe/farmacologia , Neoplasias Bucais/patologia , Fatores de Transcrição SOXB1/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/patologia , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Autofagia/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/metabolismo , Gefitinibe/uso terapêutico , Humanos , Leupeptinas/farmacologia , Macrolídeos/farmacologia , Masculino , Camundongos , Neoplasias Bucais/tratamento farmacológico , Mutagênese , Células-Tronco Neoplásicas/patologia , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Estabilidade Proteica/efeitos dos fármacos , Proteólise/efeitos dos fármacos , Fatores de Transcrição SOXB1/genética , Carcinoma de Células Escamosas de Cabeça e Pescoço/tratamento farmacológico , Ensaios Antitumorais Modelo de Xenoenxerto
13.
Autophagy ; 16(5): 782-796, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-31286822

RESUMO

Impaired macroautophagy/autophagy is involved in the pathogenesis of hepatic fibrosis. However, how aberrant autophagy promotes fibrosis is far from understood. Here, we aimed to define a previously unrevealed pro-fibrotic mechanism for the stress protein TRIB3 (tribbles pseudokinase 3)-mediated autophagy dysfunction. Human fibrotic liver tissues were obtained from patients with cirrhosis who underwent an open surgical repair process. The functional implications of TRIB3 were evaluated in mouse models of hepatic fibrosis induced by bile duct ligation (BDL) or thioacetamide (TAA) injection. Human fibrotic liver tissues expressed higher levels of TRIB3 and selective autophagic receptor SQSTM1/p62 (sequestosome 1) than nonfibrotic tissues and the elevated expression of TRIB3 and SQSTM1 was positively correlated in the fibrotic tissues. Silencing Trib3 protected against experimentally induced hepatic fibrosis, accompanied by restored autophagy activity in fibrotic liver tissues. Furthermore, TRIB3 interacted with SQSTM1 and hindered its binding to MAP1LC3/LC3, which caused the accumulation of SQSTM1 aggregates and obstructed autophagic flux. The TRIB3-mediated autophagy impairment not only suppressed autophagic degradation of late endosomes but also promoted hepatocellular secretion of INHBA/Activin A-enriched exosomes which caused migration, proliferation and activation of hepatic stellate cells (HSCs), the effector cells of liver fibrosis. Disrupting the TRIB3-SQSTM1 interaction with a specific helical peptide exerted potent protective effects against hepatic fibrosis by restoring autophagic flux in hepatocytes and HSCs. Together, stress-elevated TRIB3 expression promotes hepatic fibrosis by interacting with SQSTM1 and interfering with its functions in liver-parenchymal cells and activating HSCs. Targeting this interaction is a promising strategy for treating fibroproliferative liver diseases.Abbreviations: 3-MA: 3-methyladenine; AAV: adeno-associated virus; ACTA2/α-SMA: actin, alpha 2, smooth muscle, aorta; BDL: bile duct ligation; BECN1/Beclin 1: beclin 1, autophagy related; CHX: cycloheximide; CQ: chloroquine; Edu: 5-ethynyl-2-deoxyuridine; ESCRT: endosomal sorting complexes required for transport; HSC: hepatic stellate cell; ILV: intralumenal vesicle; LAMP1: lysosomal-associated membrane protein 1; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MVB: multivesicular body; PIK3C3: phosphatidylinositol 3-kinase, catalytic subunit type 3; PPI: protein-protein interaction; SQSTM1/p62: sequestosome 1; TAA: thioacetamide; TEM: transmission electron microscopy; TGFB1/TGFß1: transforming growth factor, beta 1; TLR2: toll-like receptor 2; TRIB3: tribbles pseudokinase 3.


Assuntos
Autofagia/fisiologia , Proteínas de Ciclo Celular/metabolismo , Cirrose Hepática/patologia , Proteínas Repressoras/metabolismo , Proteína Sequestossoma-1/metabolismo , Animais , Autofagia/genética , Proteínas de Ciclo Celular/genética , Hepatócitos/metabolismo , Humanos , Cirrose Hepática/genética , Cirrose Hepática/metabolismo , Camundongos Transgênicos , /metabolismo , Proteínas Repressoras/genética , Proteína Sequestossoma-1/genética
14.
Nat Commun ; 10(1): 5720, 2019 12 16.
Artigo em Inglês | MEDLINE | ID: mdl-31844113

RESUMO

The existence of breast cancer stem cells (BCSCs) is a major reason underlying cancer metastasis and recurrence after chemotherapy and radiotherapy. Targeting BCSCs may ameliorate breast cancer relapse and therapy resistance. Here we report that expression of the pseudokinase Tribble 3 (TRIB3) positively associates with breast cancer stemness and progression. Elevated TRIB3 expression supports BCSCs by interacting with AKT to interfere with the FOXO1-AKT interaction and suppress FOXO1 phosphorylation, ubiquitination, and degradation by E3 ligases SKP2 and NEDD4L. The accumulated FOXO1 promotes transcriptional expression of SOX2, a transcriptional factor for cancer stemness, which in turn, activates FOXO1 transcription and forms a positive regulatory loop. Disturbing the TRIB3-AKT interaction suppresses BCSCs by accelerating FOXO1 degradation and reducing SOX2 expression in mouse models of breast cancer. Our study provides insights into breast cancer development and confers a potential therapeutic strategy against TRIB3-overexpressed breast cancer.


Assuntos
Neoplasias da Mama/genética , Proteínas de Ciclo Celular/metabolismo , Proteína Forkhead Box O1/metabolismo , Células-Tronco Neoplásicas/patologia , Proteínas Repressoras/metabolismo , Fatores de Transcrição SOXB1/genética , Animais , Mama/patologia , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Progressão da Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos , Pessoa de Meia-Idade , Ligação Proteica , Proteólise , Proteínas Proto-Oncogênicas c-akt/metabolismo , Análise Serial de Tecidos , Transcrição Genética , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Immunity ; 51(3): 522-534.e7, 2019 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-31471107

RESUMO

Although recent progress provides mechanistic insights into the pathogenesis of pulmonary fibrosis (PF), rare anti-PF therapeutics show definitive promise for treating this disease. Repeated lung epithelial injury results in injury-repairing response and inflammation, which drive the development of PF. Here, we report that chronic lung injury inactivated the ubiquitin-editing enzyme A20, causing progressive accumulation of the transcription factor C/EBPß in alveolar macrophages (AMs) from PF patients and mice, which upregulated a number of immunosuppressive and profibrotic factors promoting PF development. In response to chronic lung injury, elevated glycogen synthase kinase-3ß (GSK-3ß) interacted with and phosphorylated A20 to suppress C/EBPß degradation. Ectopic expression of A20 or pharmacological restoration of A20 activity by disturbing the A20-GSK-3ß interaction accelerated C/EBPß degradation and showed potent therapeutic efficacy against experimental PF. Our study indicates that a regulatory mechanism of the GSK-3ß-A20-C/EBPß axis in AMs may be a potential target for treating PF and fibroproliferative lung diseases.


Assuntos
Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Macrófagos/metabolismo , Fibrose Pulmonar/metabolismo , Fatores de Transcrição/metabolismo , Ubiquitina/metabolismo , Animais , Linhagem Celular , Glicogênio Sintase Quinase 3 beta/metabolismo , Células HEK293 , Humanos , Inflamação/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Fosforilação/fisiologia , Transdução de Sinais/fisiologia , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação/fisiologia , Regulação para Cima/fisiologia
16.
Reprod Biol ; 19(2): 173-178, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31151753

RESUMO

The tumor suppressor gene KCTD11 plays a critical role in cell proliferation, differentiation and invasion. The current study investigated the regulation and the spatiotemporal expression pattern of Kctd11 in the rat ovary during the periovulatory period. Ovaries, granulosa cells, or theca-interstitial cells were collected at various times after hCG administration using an established gonadotropin-primed immature rat model that induces follicular development and ovulation. Real-time quantitative PCR analysis revealed that mRNA for Kctd11 was significantly induced both in theca-intersititial and granulosa cells after hCG treatment although their temporal expression patterns differed. In situ hybridization analysis demonstrated that Kctd11 mRNA expression was induced in theca-intersititial cells at 6 h after hCG, and the expression remained elevated until 12 h after hCG. Kctd11 mRNA was stimulated in granulosa cells at 6 h and reached the highest expression at 12 h. There was negligible Kctd11 mRNA signal observed in newly forming corpora lutea. In addition, the data indicate that both the protein kinase A and the protein kinase C pathway regulate the expression of Kctd11 mRNA in granulosa cells. Either forskolin or phorbol 12 myristate 13-acetate can mimic hCG induction of Kctd11 expression. Furthermore, the stimulation of Kctd11 by hCG requires new protein synthesis. Inhibition of progesterone action and the EGF pathway blocked Kctd11 mRNA expression, whereas inhibition of prostaglandin synthesis had no effect. Our finding suggest that the induction of the Kctd11 may be important for theca and granulosa cell differentiation into luteal cells.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Ovário/metabolismo , Ovulação/fisiologia , Transferases/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Animais , Proteínas de Ciclo Celular/genética , Gonadotropina Coriônica/farmacologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Células da Granulosa/fisiologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Células Tecais/fisiologia , Transferases/genética , Proteínas Supressoras de Tumor/genética
17.
Reprod Fertil Dev ; 31(4): 698-704, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30414623

RESUMO

High mobility group AT-hook 1 (HMGA1) is able to regulate gene expression and function as a tumour suppressor. The spatiotemporal expression pattern of HMGA1 was investigated in this study. Immature female rats (22-23 days old) were treated with 10IU, s.c., pregnant mare's serum gonadotrophin to stimulate follicular development, followed 48h later by injection with 5IU, s.c., human chorionic gonadotrophin (hCG). Whole ovaries or granulosa cells were collected at various times after hCG administration (n=3 per time point). Real-time polymerase chain reaction and western blot analysis revealed that HMGA1 was highly stimulated in the ovary by 4-12h after hCG treatment. In situ hybridisation analysis demonstrated that Hmga1 mRNA expression was induced in granulosa cells between 8 and 12h after hCG treatment. There was negligible Hmga1 mRNA signal observed in newly forming corpora lutea. In addition, the data indicated that both the protein kinase (PK) A and PKC pathways regulated Hmga1 expression in rat granulosa cells. In rat granulosa cell cultures, upregulation of Hmga1 was dependent on new protein synthesis because Hmga1 was inhibited by cycloheximide. Furthermore, Hmga1 mRNA expression in rat granulosa cell cultures was inhibited by AG1478, whereas NS398 and RU486 had no effect, suggesting that Hmga1 expression was regulated, in part, by the epidermal growth factor pathway. In summary, the findings of this study suggest that induction of Hmga1 may be important for theca and granulosa cell differentiation into luteal cells.


Assuntos
Diferenciação Celular/fisiologia , Proteína HMGA1a/metabolismo , Luteinização/metabolismo , Ovário/metabolismo , Ovulação/metabolismo , Animais , Diferenciação Celular/efeitos dos fármacos , Corpo Lúteo/efeitos dos fármacos , Corpo Lúteo/metabolismo , Feminino , Regulação da Expressão Gênica , Gonadotropinas Equinas/farmacologia , Células da Granulosa/efeitos dos fármacos , Células da Granulosa/metabolismo , Proteína HMGA1a/genética , Luteinização/efeitos dos fármacos , Luteinização/genética , Ovulação/efeitos dos fármacos , Ovulação/genética , Ratos , Ratos Sprague-Dawley , Células Tecais/efeitos dos fármacos , Células Tecais/metabolismo
18.
Gastroenterology ; 156(3): 708-721.e15, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30365932

RESUMO

BACKGROUND & AIMS: Activation of Wnt signaling to ß-catenin contributes to the development of colorectal cancer (CRC). Expression of tribbles pseudo-kinase 3 (TRIB3) is increased in some colorectal tumors and associated with poor outcome. We investigated whether increased TRIB3 expression promotes stem cell features of CRC cells and tumor progression by interacting with the Wnt signaling pathway. METHODS: We performed studies with C57BL/6J-ApcMin/J mice injected with an adeno-associated virus vector that expresses a small hairpin RNA against Trib3 mRNA (ApcMin/J-Trib3KD) or a control vector (ApcMin/J-Ctrl). We created BALB/c mice that overexpress TRIB3 from an adeno-associated virus vector and mice with small hairpin RNA-mediated knockdown of ß-catenin. The mice were given azoxymethane followed by dextran sodium sulfate to induce colitis-associated cancer. Intestinal tissues were collected and analyzed by histology, gene expression profiling, immunohistochemistry, and immunofluorescence. Leucine-rich repeat-containing G-protein-coupled receptor 5 (LGR5)-positive (LGR5Pos) and LGR5-negative (LGR5Neg) HCT-8 CRC cells, with or without knockdown or transgenic expression of TRIB3, were sorted and analyzed in sphere-formation assays. We derived organoids from human and mouse colorectal tumors to analyze the function of TRIB3 and test the effect of a peptide inhibitor. Wnt signaling to ß-catenin was analyzed in dual luciferase reporter, chromatin precipitation, immunofluorescence, and immunoblot assays. Proteins that interact with TRIB3 were identified by immunoprecipitation. CRC cell lines were grown in nude mice as xenograft tumors. RESULTS: At 10 weeks of age, more than half the ApcMin/J-Ctrl mice developed intestinal high-grade epithelial neoplasia, whereas ApcMin/J-Trib3KD mice had no intestinal polyps and normal histology. Colon tissues from ApcMin/J-Trib3KD mice expressed lower levels of genes regulated by ß-catenin and genes associated with cancer stem cells. Mice with overexpression of Trib3 developed more tumors after administration of azoxymethane and dextran sodium sulfate than BALB/c mice. Mice with knockdown of ß-catenin had a lower tumor burden after administration of azoxymethane and dextran sodium sulfate, regardless of Trib3 overexpression. Intestinal tissues from mice with overexpression of Trib3 and knockdown of ß-catenin did not have activation of Wnt signaling or expression of genes regulated by ß-catenin. LGR5Pos cells sorted from HCT-8 cells expressed higher levels of TRIB3 than LGR5Neg cells. CRC cells that overexpressed TRIB3 had higher levels of transcription by ß-catenin and formed larger spheroids than control CRC cells; knockdown of ß-catenin prevented the larger organoid size caused by TRIB3 overexpression. TRIB3 interacted physically with ß-catenin and transcription factor 4 (TCF4). TRIB3 overexpression increased, and TRIB3 knockdown decreased, recruitment of TCF4 and ß-catenin to the promoter region of genes regulated by Wnt. Activated ß-catenin increased expression of TRIB3, indicating a positive-feedback loop. A peptide (P2-T3A6) that bound ß-catenin disrupted its interaction with TRIB3 and TCF4. In primary CRC cells and HCT-8 cells, P2-T3A6 decreased expression of genes regulated by ß-catenin and genes associated with cancer stem cells and decreased cell viability and migration. Injection of C57BL/6J-ApcMin/J mice with P2-T3A6 decreased the number and size of tumor nodules and colon expression of genes regulated by ß-catenin. P2-T3A6 increased 5-fluorouracil-induced death of CRC cells and survival times of mice with xenograft tumors. CONCLUSION: TRIB3 interacts with ß-catenin and TCF4 in intestine cells to increase expression of genes associated with cancer stem cells. Knockdown of TRIB3 decreases colon neoplasia in mice, migration of CRC cells, and their growth as xenograft tumors in mice. Strategies to block TRIB3 activity might be developed for treatment of CRC.


Assuntos
Carcinogênese/genética , Proteínas de Ciclo Celular/genética , Transformação Celular Neoplásica/genética , Neoplasias Colorretais/genética , beta Catenina/metabolismo , Animais , Comunicação Celular/genética , Neoplasias Colorretais/patologia , Modelos Animais de Doenças , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Nus , Células-Tronco Neoplásicas/metabolismo , Distribuição Aleatória , Sensibilidade e Especificidade , Regulação para Cima , Via de Sinalização Wnt/genética , Ensaios Antitumorais Modelo de Xenoenxerto
19.
Oncogene ; 37(22): 2967-2981, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29520103

RESUMO

Metformin has beneficial effects of preventing and treating cancers on type 2 diabetic patients. However, the role of metformin in non-diabetic cancer patients and the precise molecular mechanisms against cancer have not yet been sufficiently elucidated. We recently reported that the pseudokinase protein TRIB3 acts as a stress sensor linking metabolic stressors to cancer promotion by inhibiting autophagy and ubiquitin-proteasomal degradation systems; genetically abrogating of TRIB3 expression reduces tumourigenesis and cancer progression. Thus, TRIB3 is a potential therapeutic target for diverse cancers. In this study, we found that metformin attenuates melanoma growth and metastasis by reducing TRIB3 expression in non-diabetic C57BL/6 mice and diabetic KK-Ay mice; overexpression of TRIB3 protects metformin from the activation of autophagic flux, the clearance of accumulated tumour-promoting factors and the attenuation of tumour progression. We further elucidated that TRIB3 acts as an adaptor to recruit lysine acetyltransferase 5 (KAT5) to SMAD3 and induce a phosphorylation-dependent K333 acetylation of SMAD3, which sustains transcriptional activity of SMAD3 and subsequently enhances TRIB3 transcription. Metformin suppresses SMAD3 phosphorylation and decreases the KAT5/SMAD3 interaction, to attenuate the KAT5-mediated K333 acetylation of SMAD3, reduce the SMAD3 transcriptional activity and subsequent TRIB3 expression, thereby antagonizes melanoma progression. Together, our study not only defines a molecular mechanism by which metformin protects against melanoma progression by disturbing the KAT5/TRIB3/SMAD3 positive feedback loop in diabetes and non-diabetes mice, but also suggests a candidate diverse utility of metformin in tumour prevention and therapy because of suppressing stress protein TRIB3 expression.


Assuntos
Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Lisina Acetiltransferase 5/metabolismo , Melanoma/tratamento farmacológico , Metformina/administração & dosagem , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Proteína Smad3/metabolismo , Acetilação/efeitos dos fármacos , Animais , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Progressão da Doença , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Melanoma/genética , Melanoma/metabolismo , Metformina/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , /metabolismo , Proteína Smad3/genética , Ensaios Antitumorais Modelo de Xenoenxerto
20.
Cancer Cell ; 31(5): 697-710.e7, 2017 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-28486108

RESUMO

Acute promyelocytic leukemia (APL) is driven by the oncoprotein PML-RARα, which antagonizes myeloid differentiation and promotes APL-initiating cell self-renewal. Combined all-trans retinoic acid (ATRA) with arsenic trioxide (As2O3) or chemotherapy dramatically improves the prognosis of APL patients. Here we report that expression of pseudokinase Tribble 3 (TRIB3) associates positively with APL progression and therapeutic resistance. The elevated TRIB3 expression promotes APL by interacting with PML-RARα and suppressing its sumoylation, ubiquitylation, and degradation. This represses PML nuclear body assembly, p53-mediated senescence, and cell differentiation, and supports cellular self-renewal. Genetically inhibiting TRIB3 expression or combination of a peptide disturbing TRIB3/PML-RARα interaction with ATRA/As2O3 eradicates APL by accelerating PML-RARα degradation. Our study provides insight into APL pathogenesis and a potential therapeutic option against APL.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Proliferação de Células , Senescência Celular , Leucemia Promielocítica Aguda/metabolismo , Proteínas de Fusão Oncogênica/metabolismo , Proteínas Repressoras/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Trióxido de Arsênio , Arsenicais/farmacologia , Proteínas de Ciclo Celular/deficiência , Proteínas de Ciclo Celular/genética , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Progressão da Doença , Resistencia a Medicamentos Antineoplásicos , Feminino , Regulação da Expressão Gênica , Fusão Gênica , Células HEK293 , Humanos , Leucemia Promielocítica Aguda/tratamento farmacológico , Leucemia Promielocítica Aguda/genética , Leucemia Promielocítica Aguda/patologia , Masculino , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos SCID , Camundongos Transgênicos , Proteínas de Fusão Oncogênica/genética , Óxidos/farmacologia , Peptídeos/farmacologia , /metabolismo , Estabilidade Proteica , Proteólise , Proteínas Repressoras/genética , Transdução de Sinais , Sumoilação , Fatores de Tempo , Transfecção , Tretinoína/farmacologia , Proteína Supressora de Tumor p53/genética , Ubiquitinação , Ensaios Antitumorais Modelo de Xenoenxerto
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