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1.
Mol Biol (Mosk) ; 53(5): 838-848, 2019.
Artigo em Russo | MEDLINE | ID: mdl-31661482

RESUMO

Lymphocyte phosphatase-associated phosphoprotein (LPAP) is a small transmembrane protein that is found in lymphocytes and is tightly associated with the phosphatase CD45. The function of LPAP is still unknown. Studies of the LPAP interactome may reveal new details of how C45 and lymphocyte signaling in general are regulated. LPAP binding partners were sought using coimmunoprecipitation coupled with mass spectrometry, stabilization of protein complexes with chemical crosslinkers, and Blue Native electrophoresis. In addition to CD45, several proteins were identified as LPAP partners, including CD71, CD98, cytoskeletal proteins, the amino acid transporter SLC1A4, and the cell signaling component HS1. It was confirmed that more than 70% of LPAP molecules were bound with CD45 in a 1 : 1 complex. The effect of CD45 on LPAP was studied in CEM and Jurkat cells with a CD45 knockout. The LPAP levels in the cells were 10% of the level in wild-type cells. In the absence of CD45, LPAP phosphorylation at Ser-153 and Ser-163 was not affected, whereas phosphorylation at Ser-99 and Ser-172 decreased significantly. Based on the results, CD45 was assumed to play a role in regulating LPAP expression and phosphorylation status.


Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Ativação Linfocitária , Proteínas de Membrana/metabolismo , Mapas de Interação de Proteínas , Sistema ASC de Transporte de Aminoácidos/metabolismo , Antígenos CD/metabolismo , Proteína-1 Reguladora de Fusão/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/química , Antígenos Comuns de Leucócito/metabolismo , Proteínas de Membrana/química , Fosforilação , Ligação Proteica , Receptores da Transferrina/metabolismo
2.
Zhonghua Gan Zang Bing Za Zhi ; 27(8): 628-633, 2019 Aug 20.
Artigo em Chinês | MEDLINE | ID: mdl-31594081

RESUMO

Objective: To investigate the change in expression of anti-senescence marker protein calmodulin (RGN) in liver tissues of rats with immune hepatic fibrosis, and to observe the effect of compound glutathione inosine injection (CGII) on it. Methods: Rat liver fibrosis model was induced by intraperitoneal injection of porcine serum, and CGII intervention was administered at the appropriate time. Rat liver tissues were stained with HE and Masson. RGN and protein expression at mRNA in liver tissues was detected by fluorescence quantitative PCR and immunohistochemistry. One-way Anova was used for measurement data. LDS test was used for two-way comparison, and pathological semi-quantitative results were analyzed by rank-sum test. Results: The relative expression of RGN mRNA and protein in liver tissue of fibrotic rats was 82.23 ± 15.21 and 12.52 ± 3.23, respectively, which were significantly lower than that of normal rats 176.39 ± 11.35 and 59.23 ± 9.13 (P < 0.01). The degree of liver fibrosis in fibrotic rats after CGII intervention was significantly lower than fibrotic rats. The relative expression of RGN mRNA and protein in the intervention group was 168.78 ± 21.31 and 46.42 ± 4.71, respectively, which were significantly higher than fibrosis and spontaneous recovery group. The difference was statistically significant (P < 0.01). The relative expression of RGN mRNA and protein in the spontaneous recovery group was 86.23 ± 17.16 and 14.34 ± 5.16, which was higher than model group. The difference was not statistically significant (P > 0.05). Conclusion: The expression of RGN in liver tissue of rats with hepatic fibrosis induced by porcine serum is decreased, while the expression of RGN increases with the decrease of fibrosis after CGII intervention, suggesting that the protein may play an important role in the development of liver fibrosis.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Glutationa/farmacologia , Inosina/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Cirrose Hepática Experimental/tratamento farmacológico , Animais , Hidrolases de Éster Carboxílico , Fígado/efeitos dos fármacos , Fígado/metabolismo , Cirrose Hepática Experimental/metabolismo , Ratos , Ratos Sprague-Dawley , Suínos
3.
Life Sci ; 235: 116799, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31472144

RESUMO

Colorectal cancer (CRC) is one of the most common malignancies in the world. Emerging evidence has shown that dysregulation of tripartite motif (TRIM) family proteins is strongly correlated with the tumorigenesis of CRC. Here, we evaluated the biological roles of TRIM66, a member of TRIM family, in the progression of CRC. The results demonstrated that TRIM66 was markedly up-regulated in both CRC tissues and cell lines. To further investigate the functions of TRIM66 in CRC, CRC cells were infected with lentivirus expressing anti-TRIM66 shRNA (sh-TRIM66) or control lentivirus (sh-con). We found that knockdown of TRIM66 significantly inhibited cell proliferation, migration, invasion of CRC cells. TRIM66 knockdown also suppressed epithelial-mesenchymal transition (EMT), as proved by the increased E-cadherin expression and decreased expressions of N-cadherin and vimentin. Furthermore, TRIM66 knockdown markedly inhibited tumor growth in a mouse xenograft model. Knockdown of TRIM66 reduced the activation of JAK2/STAT3 signaling pathway in CRC cells. Treatment with AG490, an inhibitor of JAK2/STAT3 signaling pathway, enhanced the inhibitory effects of TRIM66 knockdown on cell proliferation, migration and invasion. These findings suggested that knockdown of TRIM66 exhibited anti-tumor activity through inhibiting the JAK2/STAT3 signaling pathway in CRC cells.


Assuntos
Movimento Celular , Proliferação de Células , Neoplasias Colorretais/patologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Janus Quinase 2/metabolismo , Fator de Transcrição STAT3/metabolismo , Animais , Apoptose , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Transição Epitelial-Mesenquimal , Feminino , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Peptídeos e Proteínas de Sinalização Intracelular/genética , Janus Quinase 2/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Invasividade Neoplásica , RNA Interferente Pequeno/genética , Fator de Transcrição STAT3/genética , Transdução de Sinais , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
4.
Nature ; 574(7777): 259-263, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31554973

RESUMO

Chikungunya virus (CHIKV) is a re-emerging alphavirus that is transmitted to humans by mosquito bites and causes musculoskeletal and joint pain1,2. Despite intensive investigations, the human cellular factors that are critical for CHIKV infection remain unknown, hampering the understanding of viral pathogenesis and the development of anti-CHIKV therapies. Here we identified the four-and-a-half LIM domain protein 1 (FHL1)3 as a host factor that is required for CHIKV permissiveness and pathogenesis in humans and mice. Ablation of FHL1 expression results in the inhibition of infection by several CHIKV strains and o'nyong-nyong virus, but not by other alphaviruses and flaviviruses. Conversely, expression of FHL1 promotes CHIKV infection in cells that do not normally express it. FHL1 interacts directly with the hypervariable domain of the nsP3 protein of CHIKV and is essential for the replication of viral RNA. FHL1 is highly expressed in CHIKV-target cells and is particularly abundant in muscles3,4. Dermal fibroblasts and muscle cells derived from patients with Emery-Dreifuss muscular dystrophy that lack functional FHL15 are resistant to CHIKV infection. Furthermore,  CHIKV infection  is undetectable in Fhl1-knockout mice. Overall, this study shows that FHL1 is a key factor expressed by the host that enables CHIKV infection and identifies the interaction between nsP3 and FHL1 as a promising target for the development of anti-CHIKV therapies.


Assuntos
Febre de Chikungunya/virologia , Vírus Chikungunya/patogenicidade , Fatores Celulares Derivados do Hospedeiro/metabolismo , Interações Hospedeiro-Patógeno , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas com Domínio LIM/metabolismo , Proteínas Musculares/metabolismo , Animais , Células Cultivadas , Febre de Chikungunya/tratamento farmacológico , Vírus Chikungunya/efeitos dos fármacos , Vírus Chikungunya/genética , Vírus Chikungunya/crescimento & desenvolvimento , Feminino , Fibroblastos/virologia , Células HEK293 , Fatores Celulares Derivados do Hospedeiro/genética , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/deficiência , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas com Domínio LIM/deficiência , Proteínas com Domínio LIM/genética , Masculino , Camundongos , Proteínas Musculares/deficiência , Proteínas Musculares/genética , Mioblastos/virologia , Vírus O'nyong-nyong/crescimento & desenvolvimento , Vírus O'nyong-nyong/patogenicidade , Ligação Proteica , RNA Viral/biossíntese , Proteínas não Estruturais Virais/genética , Proteínas não Estruturais Virais/metabolismo , Replicação Viral
5.
Nat Cell Biol ; 21(9): 1127-1137, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31481798

RESUMO

The inner centromere is a region on every mitotic chromosome that enables specific biochemical reactions that underlie properties, such as the maintenance of cohesion, the regulation of kinetochores and the assembly of specialized chromatin, that can resist microtubule pulling forces. The chromosomal passenger complex (CPC) is abundantly localized to the inner centromeres and it is unclear whether it is involved in non-kinase activities that contribute to the generation of these unique chromatin properties. We find that the borealin subunit of the CPC drives phase separation of the CPC in vitro at concentrations that are below those found on the inner centromere. We also provide strong evidence that the CPC exists in a phase-separated state at the inner centromere. CPC phase separation is required for its inner-centromere localization and function during mitosis. We suggest that the CPC combines phase separation, kinase and histone code-reading activities to enable the formation of a chromatin body with unique biochemical activities at the inner centromere.


Assuntos
Centrômero/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Cinetocoros/metabolismo , Microtúbulos/metabolismo , Proteínas de Ciclo Celular/metabolismo , Segregação de Cromossomos/fisiologia , Citoesqueleto/metabolismo , Células HeLa , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Mitose
6.
Adv Exp Med Biol ; 1155: 113-118, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31468390

RESUMO

We previously showed that taurine administration contributed to the extension of time to exhaustion through exercise-induced hypoglycemia restraint, and we suggested that the activation of hepatic gluconeogenesis was initiated before the exercise with the taurine administration. We hypothesize that the extension effect of exercise duration with the taurine administration is restrained in the rats which inhibited hepatic gluconeogenesis. In this study, we aimed to produce a rat model that inhibited hepatic gluconeogenesis as a first step in testing our hypothesis.F344 male rats of 8 weeks after birth were purchased. The blood samples were collected via jugular vein catheter to perform the pyruvate tolerance test (PTT) with the intraperitoneal administration, and to determine the optimal time point of blood glucose measurement. 3-mercaptopicolinic acids (3MPA) was used as an inhibitor of PEPCK. The rats were divided into three groups, Non-dosage control (CON) group, 30 mg/kg・BW 3MPA (3MPA 30) group, and 300 mg/kg・BW 3MPA (3MPA 300) group.The blood glucose level showed a significant peak 15 min after pyruvate administration. The change of the blood glucose level after the PTT in 3MPA 300 group was significantly smaller than that of the CON group at this time point. These results show we could prepare the rat model that inhibited hepatic gluconeogenesis.


Assuntos
Modelos Animais de Doenças , Gluconeogênese , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Fígado/fisiopatologia , Fosfoenolpiruvato Carboxiquinase (GTP)/metabolismo , Animais , Glicemia , Masculino , Condicionamento Físico Animal , Ratos , Ratos Endogâmicos F344 , Taurina
7.
Yonsei Med J ; 60(9): 832-841, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31433581

RESUMO

PURPOSE: Epirubicin is one of the most effective drugs against osteosarcoma. miR-1301 is involved in the occurrence and development of osteosarcoma. Whether miR-1301 is responsible for the chemosensitivity of osteosarcoma cells to epirubicin remains largely unknown. MATERIALS AND METHODS: U2OS and SAOS-2 cells were treated with various concentrations of epirubicin. Flow cytometry was employed to evaluate cell apoptotic rate. Cell proliferation was measured by Cell Counting Kit-8 assay. Western blot and quantitative real-time polymerase chain reaction were utilized to detect the expressions of B-cell lymphoma-2 (Bcl-2), Bcl-2 assaciated X protein (Bax), cleaved-caspase-3, cleaved-poly (ADP-ribose) polymerases (PARP1), TP53-regulated inhibitor of apoptosis 1 (TRIAP1), and microRNA-1301 (miR-1301). The relationship between miR-1301 and TRIAP1 was determined by luciferase reporter assay. RESULTS: Epirubicin inhibited proliferation in a dose-dependent manner, induced apoptosis, decreased the expression of Bcl-2, and increased the expressions of Bax, cleaved-caspase-3, and cleaved-PARP1 in osteosarcoma cells. miR-1301 was downregulated in U2OS and SAOS-2 cells. Importantly, epirubicin significantly increased the levels of miR-1301. Overexpression of miR-1301 suppressed proliferation and promoted apoptosis. Interestingly, those effects were enhanced by epirubicin. In contrast, miR-1301 depletion attenuated the epirubicin-mediated anti-osteosarcoma effect. miR-1301 negatively regulated the expression of TRIAP1 in U2OS and SAOS-2 cells. Furthermore, epirubicin inhibited the mRNA and protein levels of TRIAP1 by upregulating miR-1301 levels. Epirubicin suppressed cell proliferation by downregulating TRIAP1. CONCLUSION: miR-1301 was implicated in the chemosensitivity of osteosarcoma to epirubicin by modulating TRIAP1.


Assuntos
Antibióticos Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Neoplasias Ósseas/tratamento farmacológico , Proliferação de Células/efeitos dos fármacos , Epirubicina/uso terapêutico , Peptídeos e Proteínas de Sinalização Intracelular/genética , MicroRNAs/genética , Osteossarcoma/tratamento farmacológico , Neoplasias Ósseas/genética , Neoplasias Ósseas/patologia , Caspase 3/biossíntese , Linhagem Celular Tumoral , Regulação para Baixo , Epirubicina/farmacologia , Citometria de Fluxo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , MicroRNAs/metabolismo , Osteossarcoma/genética , Osteossarcoma/patologia , Reação em Cadeia da Polimerase em Tempo Real , Regulação para Cima
8.
Cancer Sci ; 110(10): 3122-3131, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31369178

RESUMO

Delta-like 3 (DLL3) is a member of the Delta/Serrate/Lag2 (DSL) group of Notch receptor ligands. Five DSL ligands are known in mammals, among which DLL3 has a unique structure. In the last few years, DLL3 has attracted attention as a novel molecular targeting gene in neuroendocrine carcinoma of the lung due to its high expression. However, the expression pattern and functions of DLL3 in the gastrointestinal tract and gastrointestinal neuroendocrine carcinoma remain unclear. In this study, we examined the expression and role of DLL3 in the gastrointestinal tract, as well as in gastrointestinal neuroendocrine carcinoma. Immunohistochemical staining of the human normal gastrointestinal tract revealed that DLL3 localized in neuroendocrine cells. DLL3 showed intense staining in chromogranin A-positive gastric cancer specimens. Real-time quantitative RT-PCR and western blotting analyses showed considerable upregulation of DLL3 in gastrointestinal neuroendocrine carcinoma cell lines. Immuno-electron microscopy demonstrated abundant expression of DLL3 in neurosecretory granules in these cells. Furthermore, gene silencing of DLL3 caused significant growth inhibition through the induction of intrinsic apoptosis. Our findings suggest that DLL3 is expressed in neuroendocrine cells of the gastrointestinal tract and that it has a pivotal role in gastrointestinal neuroendocrine carcinoma cells. Based on these findings, further investigations are required to achieve a breakthrough in developing therapeutic strategies for gastrointestinal neuroendocrine carcinoma.


Assuntos
Carcinoma Neuroendócrino/metabolismo , Neoplasias Gastrointestinais/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Células Neuroendócrinas/metabolismo , Idoso , Apoptose , Carcinoma Neuroendócrino/genética , Linhagem Celular Tumoral , Neoplasias Gastrointestinais/genética , Trato Gastrointestinal/citologia , Trato Gastrointestinal/metabolismo , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Masculino , Regulação para Cima
9.
Physiol Rev ; 99(4): 1655-1699, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31313981

RESUMO

Integrins are heterodimeric cell surface receptors ensuring the mechanical connection between cells and the extracellular matrix. In addition to the anchorage of cells to the extracellular matrix, these receptors have critical functions in intracellular signaling, but are also taking center stage in many physiological and pathological conditions. In this review, we provide some historical, structural, and physiological notes so that the diverse functions of these receptors can be appreciated and put into the context of the emerging field of mechanobiology. We propose that the exciting journey of the exploration of these receptors will continue for at least another new generation of researchers.


Assuntos
Adesão Celular , Membrana Celular/metabolismo , Matriz Extracelular/metabolismo , Integrinas/metabolismo , Mecanotransdução Celular , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proliferação de Células , Humanos , Integrinas/química , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Modelos Moleculares , Fosfoproteínas/metabolismo , Conformação Proteica , Relação Estrutura-Atividade , Fatores de Transcrição
10.
Toxicol Lett ; 313: 130-136, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31276767

RESUMO

We previously demonstrated that based on their potency, contact allergens differently modulate Blimp-1/NLRP12 expression in human keratinocytes, with the extreme allergen 2,4-dinitrochlorobenzene (DNCB) more rapidly upregulating Blimp-1, leading to downregulation of NLRP12, and to the production of interleukin-18 (IL-18). The purpose of this study was to further investigate the effects of DNCB and para-phenylenediamine (PPD) on the expression of the proteins of the inflammasome, namely NLRP3, ASC and caspase 1 by western blot analysis; to define the intracellular localization and co-localization of NLRP3 and NLPR12 by immunoprecipitation and immunohistochemistry; and to define the role of NF-κB in Blimp-1 induction by pharmacological inhibition. The human keratinocyte cell line NCTC2544 was used for all experiments. Dose and time course experiments were performed to evaluate the effect of the selected contact allergens on the parameters investigated. Results indicate, that consistent with previous finding, DNCB more rapidly (3 h) induces NLRP3, ASC protein expression and caspase-1 activation compared to PPD. Immunoprecipitation studies show the recruitment of ASC to the inflammasome following exposure to both allergens, while high level of NLRP12 and less ASC protein were found associated in control cells. By immunohistochemistry, we found increased NLRP3 expression following exposure to contact allergens, and observed a nuclear co-localization of the two proteins, indicating the NLRP12 likely acts preventing the cytosolic localization of NLRP3 and inflammasome assembly. Finally, contact allergen-induced Blimp-1 mRNA and protein expression can be completely blocked by inhibiting NF-κB activation, confirming the central role of NF-κB in contact allergen-induced keratinocyte activation.


Assuntos
Alérgenos/toxicidade , Dermatite Alérgica de Contato/etiologia , Dinitroclorobenzeno/toxicidade , Inflamassomos/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Queratinócitos/efeitos dos fármacos , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Fenilenodiaminas/toxicidade , Proteínas Adaptadoras de Sinalização CARD/metabolismo , Caspase 1/metabolismo , Linhagem Celular , Dermatite Alérgica de Contato/genética , Dermatite Alérgica de Contato/metabolismo , Relação Dose-Resposta a Droga , Humanos , Inflamassomos/genética , Inflamassomos/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Queratinócitos/metabolismo , NF-kappa B/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Fatores de Tempo
11.
Anticancer Res ; 39(6): 2689-2697, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31177103

RESUMO

Colorectal cancer (CRC) is the second most prevalent type of cancer among males and the third among females. CRC recurrence and poor prognosis may be related to the prevalence of chemotherapy-resistant cancer stem cells (CSCs). Recent studies have indicated the role of doublecortin-like kinase 1 (DCLK1) protein as a marker of CSC in CRC. This review focuses on the role of DCLK1 in CRC. Long-lived DCLK1-positive tuft cells can function as cancer-initiating cells. Numerous studies have shown DCLK1 overexpression to be significantly correlated with the stage of disease, the presence of metastasis and poor survival rate. DCLK1 may also be used to identify patients at high risk and those with chemotherapy-resistant tumors. DCLK1-specific drugs are examined as potential cancer treatments.


Assuntos
Neoplasias Colorretais/patologia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Regulação para Cima , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Metástase Neoplásica , Estadiamento de Neoplasias , Células-Tronco Neoplásicas/metabolismo , Prognóstico , Análise de Sobrevida
12.
Nat Commun ; 10(1): 2882, 2019 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-31253783

RESUMO

NLR Family CARD Domain Containing 5 (NLRC5), an important immune regulator in innate immunity, is involved in regulating inflammation and antigen presentation. However, the role of NLRC5 in vascular remodeling remains unknown. Here we report the role of NLRC5 on vascular remodeling and provide a better understanding of its underlying mechanism. Nlrc5 knockout (Nlrc5-/-) mice exhibit more severe intimal hyperplasia compared with wild-type mice after carotid ligation. Ex vivo data shows that NLRC5 deficiency leads to increased proliferation and migration of human aortic smooth muscle cells (HASMCs). NLRC5 binds to PPARγ and inhibits HASMC dedifferentiation. NACHT domain of NLRC5 is essential for the interaction with PPARγ and stimulation of PPARγ activity. Pioglitazone significantly rescues excessive intimal hyperplasia in Nlrc5-/- mice and attenuates the increased proliferation and dedifferentiation in NLRC5-deficient HASMCs. Our study demonstrates that NLRC5 regulates vascular remodeling by directly inhibiting SMC dysfunction via its interaction with PPARγ.


Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Túnica Íntima/metabolismo , Animais , Aorta , Apoptose , Pressão Sanguínea , Transplante de Medula Óssea , Proliferação de Células , Células Cultivadas , Citocinas/genética , Citocinas/metabolismo , Frequência Cardíaca , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Camundongos , Camundongos Knockout , Miócitos de Músculo Liso/metabolismo , Plasmídeos , Transcriptoma , Remodelação Vascular
13.
Nat Commun ; 10(1): 2850, 2019 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-31253801

RESUMO

Cancer cells secrete matrix metalloproteinases to remodel the extracellular matrix, which enables them to overcome tissue barriers and form metastases. The membrane-bound matrix metalloproteinase MT1-MMP (MMP14) is internalized by endocytosis and recycled in endosomal compartments. It is largely unknown how endosomal sorting and recycling of MT1-MMP are controlled. Here, we show that the endosomal protein WDFY2 controls the recycling of MT1-MMP. WDFY2 localizes to endosomal tubules by binding to membranes enriched in phosphatidylinositol 3-phosphate (PtdIns3P). We identify the v-SNARE VAMP3 as an interaction partner of WDFY2. WDFY2 knockout causes a strong redistribution of VAMP3 into small vesicles near the plasma membrane. This is accompanied by increased, VAMP3-dependent secretion of MT1-MMP, enhanced degradation of extracellular matrix, and increased cell invasion. WDFY2 is frequently lost in metastatic cancers, most predominantly in ovarian and prostate cancer. We propose that WDFY2 acts as a tumor suppressor by serving as a gatekeeper for VAMP3 recycling.


Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Metaloproteinases da Matriz/metabolismo , Invasividade Neoplásica , Proteína 3 Associada à Membrana da Vesícula/metabolismo , Actinas/fisiologia , Linhagem Celular Tumoral , Membrana Celular , Exocitose/fisiologia , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Metaloproteinases da Matriz/genética , Microtúbulos , Fosfatos de Fosfatidilinositol/fisiologia , Transporte Proteico , Proteína 3 Associada à Membrana da Vesícula/genética , Proteínas rab4 de Ligação ao GTP/genética , Proteínas rab4 de Ligação ao GTP/metabolismo
14.
J Exp Clin Cancer Res ; 38(1): 273, 2019 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-31228948

RESUMO

BACKGROUND: DEPTOR is an endogenous inhibitor of mTORC1 and mTORC2 that plays a vital role in the progression of human malignances. However, the biological function of DEPTOR in HCC metastasis and the underlying molecular mechanisms are still unclear. METHODS: Western blot analysis and immunohistochemistry(IHC) were employed to examine DEPTOR expression in HCC cell lines and tissues. A series of in vivo and in vitro assays were performed to determine the function of DEPTOR and the possible mechanisms underlying its role in HCC metastasis. RESULTS: We found that DEPTOR was frequently overexpressed in HCC tissues, and its high expression was associated with high serum AFP levels, increased tumor size, vascular invasion and more advanced TMN and BCLC stage, as well as an overall poor prognosis. Functional experiments demonstrated that DEPTOR silencing inhibited the proliferation and mobility of HCC cells in vitro and suppressed tumor growth and metastasis of HCC cells in vivo. Accordingly, DEPTOR overexpression promoted the invasion and metastasis of HCC cells in vitro and in vivo, but had no effect on cell proliferation in vitro. Overexpression of DEPTOR induced EMT by snail induction. Conversely, knockdown of snail expression impaired the DEPTOR-induced migration, invasion and EMT of HCC cells. Furthermore, we found that the increase of snail expression by DEPTOR overexpression was due to an activation of TGF-ß1-smad3/smad4 signaling possibly through feedback inhibition of mTOR. CONCLUSION: DEPTOR promotes the EMT and metastasis of HCC cells by activating the TGF-ß1-smad3/smad4-snail pathway via mTOR inhibition. Therefore, targeting DEPTOR may be an ideal treatment strategy for inhibiting the growth and metastasis of HCC.


Assuntos
Carcinoma Hepatocelular/patologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Hepáticas/patologia , Transdução de Sinais , Regulação para Cima , Adulto , Idoso , Animais , Comunicação Autócrina , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Transição Epitelial-Mesenquimal , Feminino , Regulação Neoplásica da Expressão Gênica , Células Hep G2 , Humanos , Neoplasias Hepáticas/metabolismo , Masculino , Camundongos , Pessoa de Meia-Idade , Metástase Neoplásica , Estadiamento de Neoplasias , Prognóstico , Análise de Sobrevida , Serina-Treonina Quinases TOR/metabolismo , Fator de Crescimento Transformador beta1/metabolismo
15.
Nat Commun ; 10(1): 2761, 2019 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-31235698

RESUMO

Cerebral cavernous malformation (CCM) is a neurovascular familial or sporadic disease that is characterised by capillary-venous cavernomas, and is due to loss-of-function mutations to any one of three CCM genes. Familial CCM follows a two-hit mechanism similar to that of tumour suppressor genes, while in sporadic cavernomas only a small fraction of endothelial cells shows mutated CCM genes. We reported that in mouse models and in human patients, endothelial cells lining the lesions have different features from the surrounding endothelium, as they express mesenchymal/stem-cell markers. Here we show that cavernomas originate from clonal expansion of few Ccm3-null endothelial cells that express mesenchymal/stem-cell markers. These cells then attract surrounding wild-type endothelial cells, inducing them to express mesenchymal/stem-cell markers and to contribute to cavernoma growth. These characteristics of Ccm3-null cells are reminiscent of the tumour-initiating cells that are responsible for tumour growth. Our data support the concept that CCM has benign tumour characteristics.


Assuntos
Proteínas Reguladoras de Apoptose/genética , Neoplasias do Sistema Nervoso Central/patologia , Células Endoteliais/patologia , Hemangioma Cavernoso do Sistema Nervoso Central/patologia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas de Membrana/genética , Proteínas Proto-Oncogênicas/genética , Animais , Proteínas Reguladoras de Apoptose/metabolismo , Biomarcadores/metabolismo , Encéfalo/irrigação sanguínea , Encéfalo/citologia , Encéfalo/patologia , Diferenciação Celular/genética , Linhagem Celular , Neoplasias do Sistema Nervoso Central/genética , Modelos Animais de Doenças , Células Endoteliais/metabolismo , Endotélio Vascular/citologia , Endotélio Vascular/patologia , Feminino , Técnicas de Inativação de Genes , Hemangioma Cavernoso do Sistema Nervoso Central/genética , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Mutação com Perda de Função , Proteínas de Membrana/metabolismo , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Knockout , Proteínas Proto-Oncogênicas/metabolismo
16.
Nat Commun ; 10(1): 2532, 2019 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-31182717

RESUMO

Targeted inhibition of the ERK-MAPK pathway, upregulated in a majority of human cancers, has been hindered in the clinic by drug resistance and toxicity. The MRAS-SHOC2-PP1 (SHOC2 phosphatase) complex plays a key role in RAF-ERK pathway activation by dephosphorylating a critical inhibitory site on RAF kinases. Here we show that genetic inhibition of SHOC2 suppresses tumorigenic growth in a subset of KRAS-mutant NSCLC cell lines and prominently inhibits tumour development in autochthonous murine KRAS-driven lung cancer models. On the other hand, systemic SHOC2 ablation in adult mice is relatively well tolerated. Furthermore, we show that SHOC2 deletion selectively sensitizes KRAS- and EGFR-mutant NSCLC cells to MEK inhibitors. Mechanistically, SHOC2 deletion prevents MEKi-induced RAF dimerization, leading to more potent and durable ERK pathway suppression that promotes BIM-dependent apoptosis. These results present a rationale for the generation of SHOC2 phosphatase targeted therapies, both as a monotherapy and to widen the therapeutic index of MEK inhibitors.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Pulmonares/genética , Inibidores de Proteínas Quinases/farmacologia , Quinases raf/metabolismo , Animais , Apoptose , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Feminino , Células HEK293 , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Sistema de Sinalização das MAP Quinases , Masculino , Camundongos Knockout , Camundongos Nus , Mutação , Transplante de Neoplasias , Multimerização Proteica , Quinases raf/antagonistas & inibidores , Quinases raf/genética , Proteínas ras/metabolismo
17.
Int J Oncol ; 55(1): 81-92, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31180521

RESUMO

Renal cell carcinoma (RCC) is the most common type of kidney cancer. By analysing The Cancer Genome Atlas (TCGA) database, 16 genes were identified to be consistently highly expressed in RCC tissues compared with the matched para­tumour tissues. Using a high­throughput cell viability screening method, it was found that downregulation of only two genes significantly inhibited the viability of 786­O cells. Among the two genes, pleckstrin homology domain containing O1 (PLEKHO1) has never been studied in RCC, to the best of our knowledge, and its expression level was shown to be associated with the prognosis of patients with RCC in TCGA dataset. The upregulation of PLEKHO1 in RCC was first confirmed in 30 paired tumour and para­tumour tissues. Then, the effect of PLEKHO1 on cell proliferation and apoptosis was assessed in vitro. Additionally, xenograft tumour models were established to investigate the function of PLEKHO1 in vivo. The results showed that PLEKHO1 knockdown significantly inhibited cell viability and facilitated apoptosis in vitro and impaired tumour formation in vivo. Thus, PLEKHO1 is likely to be associated with the viability of RCC cells in vitro and in vivo. Further gene expression microarray and co­expression analyses showed that PLEKHO1 may be involved in the serine/threonine­protein kinase hippo and JNK signalling pathways. Together, the results of the present study suggest that PLEKHO1 may contribute to the development of RCC, and therefore, further study is needed to explore its potential as a therapeutic target.


Assuntos
Carcinoma de Células Renais/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Renais/metabolismo , Sistema de Sinalização das MAP Quinases , Proteínas Serina-Treonina Quinases/metabolismo , Adulto , Idoso , Animais , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/patologia , Linhagem Celular Tumoral , Sobrevivência Celular/fisiologia , Feminino , Técnicas de Silenciamento de Genes , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/biossíntese , Peptídeos e Proteínas de Sinalização Intracelular/genética , Neoplasias Renais/genética , Neoplasias Renais/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Prognóstico , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Regulação para Cima
18.
Food Chem Toxicol ; 131: 110533, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31150783

RESUMO

Hepatocellular carcinoma is the fifth most common and the third most lethal cancer worldwide. In recent years, natural flavonoids have drawn great attention as repository for the exploitation of novel antineoplastic agents. 2',4'-Dihydroxy-6'-methoxy-3',5'-dimethylchalcone (DMC), a functional chalcone isolated from the buds of Cleistocalyx operculatus, has been reported to exert potent cytotoxicity against multi-drug resistant BEL-7402/5-FU cells. In this study, the precise mechanisms of DMC-mediated growth inhibition in BEL-7402/5-FU cells were further investigated. DMC was found to trigger apoptosis predominantly via the mitochondria-dependent pathway and the enhancement of reactive oxygen species (ROS) generation. Meanwhile, DMC induced G1 cell cycle arrest through downregulation of cyclin D1 and CDK4. Furthermore, DMC increased p53 level and inhibited NF-κB nuclear-localization via suppression of PI3K/AKT signaling axis, which might be the underlying mechanism of DMC-induced apoptosis and cell cycle arrest in BEL-7402/5-FU cells. Collectively, the study elucidated the mechanisms by which DMC may inhibit the growth of BEL-7402/5-FU cells and suggested the possibility that DMC might be a promising candidate therapeutic agent for hepatoma treatment in the future.


Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Chalconas/farmacologia , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Transdução de Sinais/efeitos dos fármacos , Linhagem Celular Tumoral , Flores/química , Humanos , NF-kappa B/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Syzygium/química , Proteína Supressora de Tumor p53/metabolismo
19.
Biochim Biophys Acta Rev Cancer ; 1872(1): 24-36, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31152822

RESUMO

Cancer cells constantly face a fluctuating nutrient supply and interference with adaptive responses might be an effective therapeutic approach. It has been discovered that in the absence of glucose, cancer cells can synthesize crucial metabolites by expressing phosphoenolpyruvate carboxykinase (PEPCK, PCK1 or PCK2) using abbreviated forms of gluconeogenesis. Gluconeogenesis, which in essence is the reverse pathway of glycolysis, uses lactate or amino acids to feed biosynthetic pathways branching from glycolysis. PCK1 and PCK2 have been shown to be critical for the growth of certain cancers. In contrast, fructose-1,6-bisphosphatase 1 (FBP1), a downstream gluconeogenesis enzyme, inhibits glycolysis and tumor growth, partly by non-enzymatic mechanisms. This review sheds light on the current knowledge of cancer cell gluconeogenesis and its role in metabolic reprogramming, cancer cell plasticity, and tumor growth.


Assuntos
Proliferação de Células/genética , Gluconeogênese/genética , Redes e Vias Metabólicas/genética , Neoplasias/genética , Aminoácidos/metabolismo , Frutose-Bifosfatase/genética , Frutose-Bifosfatase/metabolismo , Glucose/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Fosfoenolpiruvato Carboxiquinase (ATP)/genética , Fosfoenolpiruvato Carboxiquinase (ATP)/metabolismo , Fosfoenolpiruvato Carboxiquinase (GTP)/genética , Fosfoenolpiruvato Carboxiquinase (GTP)/metabolismo
20.
Cell Prolif ; 52(5): e12654, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31222857

RESUMO

OBJECTIVES: Despite of the aberrant expression of 14-3-3ζ in head and neck squamous cell carcinoma (HNSCC), little is known about the role of 14-3-3ζ in the regulation of senescence in HNSCC. This study was performed to investigate whether 14-3-3ζ is implicated in senescence evasion of Hep-2 laryngeal cancer cells. METHODS: The expression of 14-3-3ζ was suppressed using RNA interference strategy. Senescence induction was determined by senescence-associated ß-galactosidase staining and the numbers of promyelocytic leukaemia nuclear body. Real-time PCR, western blotting and immunohistochemistry were applied for the expression of corresponding proteins. Xenograft experiment was performed to show in vivo effect of 14-3-3ζ silencing on tumour growth. RESULTS: 14-3-3ζ silencing significantly induced senescence phenotypes via 27 accumulations. Subsequently, we demonstrated that p27 accumulation is linked to inactivation of SCFSkp2 complex activity, probably due to the deneddylation of cullin-1 (Cul-1) as follows. (a) Neddylated Cul-1 is decreased by 14-3-3ζ silencing. (b) Blocking neddylation using MLN4924 reproduces senescence phenotypes. (c) Knockdown of CSN5, which functions as a deneddylase, was shown to restore the senescence phenotypes induced by 14-3-3ζ depletion. Finally, we demonstrated that 14-3-3ζ depletion effectively hindered the proliferation of Hep-2 cells implanted into nude mice. CONCLUSION: 14-3-3ζ negatively regulates senescence in Hep-2 cells, suggesting that 14-3-3ζ targeting may serve to suppress the expansion of laryngeal cancer via induction of senescence through the Cul-1/SCFSkp2 /p27 axis.


Assuntos
Proteínas 14-3-3/metabolismo , Proteínas Culina/metabolismo , Proteínas F-Box/metabolismo , Proteínas Quinases Associadas a Fase S/metabolismo , Proteínas 14-3-3/antagonistas & inibidores , Proteínas 14-3-3/genética , Animais , Complexo do Signalossomo COP9/antagonistas & inibidores , Complexo do Signalossomo COP9/genética , Complexo do Signalossomo COP9/metabolismo , Pontos de Checagem do Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Senescência Celular , Inibidor de Quinase Dependente de Ciclina p27/genética , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Laríngeas/metabolismo , Neoplasias Laríngeas/patologia , Masculino , Camundongos , Camundongos Nus , Peptídeo Hidrolases/genética , Peptídeo Hidrolases/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Proteínas Quinases Associadas a Fase S/genética
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