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1.
Int J Biol Macromol ; 271(Pt 1): 132438, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38761906

RESUMO

Spider silk is the self-assembling product of silk proteins each containing multiple repeating units. Each repeating unit is entirely intrinsically disordered or contains a small disordered domain. The role of the disordered domain/unit in conferring silk protein storage and self-assembly is not fully understood yet. Here, we used biophysical and biochemical techniques to investigate the self-assembly of a miniature version of a minor ampullate spidroin (denoted as miniMiSp). miniMiSp consists of two identical intrinsically disordered domains, one folded repetitive domain, and two folded terminal domains. Our data indicated that miniMiSp self-assembles into oligomers and further into liquid droplets. The oligomerization is attributed to the aggregation-prone property of both the disordered domains and the folded repetitive domain. Our results support the model of micellar structure for silk proteins at high protein concentrations. The disordered domain is indispensable for liquid droplet formation via liquid-liquid phase separation, and tyrosine residues located in the disordered domain make dominant contributions to stability of the liquid droplets. As the same tyrosine residues are also critical to fibrillation, the liquid droplets are likely an intermediate state between the solution state and the fiber state. Additionally, the terminal domains contribute to the pH- and salt-dependent self-assembly properties.


Assuntos
Fibroínas , Proteínas Intrinsicamente Desordenadas , Aranhas , Aranhas/química , Animais , Proteínas Intrinsicamente Desordenadas/química , Fibroínas/química , Seda/química , Concentração de Íons de Hidrogênio , Domínios Proteicos , Multimerização Proteica , Sequência de Aminoácidos
2.
Medicine (Baltimore) ; 94(6): e502, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25674742

RESUMO

Our previous study indicated overexpression of metadherin (MTDH) is an adverse prognostic factor in squamous cell carcinoma of the head and neck (SCCHN) and promotes SCCHN cell proliferation and invasion. However, its mechanism remains unclear. Recent studies have indicated that MTDH is a cancer-metastasis-associated molecule that participates in the process of angiogenesis. Therefore, the study is aimed to investigate that whether vascular endothelial growth factor (VEGF), as one of the most potent proangiogenic cytokines, is regulated by MTDH and the role of the phosphatidylinositide 3-kinases/Protein Kinase B (PI3K/Akt) pathway in this process of regulation and the clinical significance of both MTDH and VEGF in SCCHN.Immunohistochemistry was used to assay the expression of MTDH and VEGF in a cohort of 189 SCCHN patients with intact follow-up information. The expression of MTDH was then upregulated or inhibited by lentivirus-mediated MTDH Complementary deoxyribonucleic acid or MTDH short hairpin ribonucleic acid (shRNA) to observe the resulting alterations in VEGF expression and the PI3K/Akt signaling pathway in SCCHN cell lines. In addition, the PI3K/Akt pathway was modulated to observe the resulting changes in the MTDH-mediated expression of VEGF.The immunohistochemistry data showed that MTDH expression is positively correlated with VEGF expression in SCCHN tissues. Moreover, the overexpression of MTDH in SCCHN Tu686 and 5-8F cells led to increases in the expression of VEGF, and this effect was accompanied by activation of the PI3K/Akt pathway. Conversely, shRNA-mediated knockdown of MTDH led to decreased VEGF expression. In addition, inhibition of the Akt signaling pathway reversed the upregulation of VEGF resulting from MTDH overexpression. Moreover, the survival analysis revealed that VEGF is an independent prognostic factor, and a combined survival analysis based on both MTDH and VEGF showed synergistic effects in the prognosis evaluation of SCCHN patients.The findings of the present study demonstrate that MTDH regulates the expression of VEGF via the PI3K/Akt signaling pathway, indicating the potential role of the MTDH-mediated activation of VEGF signaling pathway in SCCHN angiogenesis and metastasis.


Assuntos
Carcinoma de Células Escamosas/metabolismo , Moléculas de Adesão Celular/fisiologia , Neoplasias de Cabeça e Pescoço/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Western Blotting , Carcinoma de Células Escamosas/mortalidade , Células Cultivadas , Ensaio de Imunoadsorção Enzimática , Neoplasias de Cabeça e Pescoço/mortalidade , Humanos , Imuno-Histoquímica , Proteínas de Membrana , Prognóstico , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , RNA Interferente Pequeno/fisiologia , Proteínas de Ligação a RNA , Transdução de Sinais/fisiologia , Transfecção , Regulação para Cima
3.
Artigo em Chinês | MEDLINE | ID: mdl-23755789

RESUMO

OBJECTIVE: To investigate the regulatory effect of erythropoietin-producing hepatocellular receptor (EphA2) on the expression of VEGF protein, a pro-angiogenic factor, via p38 mitogen-activated protein kinase (p38 MAPK) signaling pathway in squamous cell carcinoma of the head and neck(SCCHN) in vitro. METHODS: SCCHN Tu686 cells were transfected with EphA2 overexpression vector pEGFP-N1-EphA2. Western blot was used to detect the expression of p38 MAPK and enzyme-linked immunosorbent assay (ELISA) was applied to assay of VEGF. SB203580 as a inhibitor of p38 MAPK signaling pathway was used. RESULTS: The expression of VEGF protein was significantly up-regulated in Tu686 cells transfected with EphA2 overexpression vector (535.31 ± 45.71) pg/ml, when compared with Tu686 cells transfected with empty vector (400.99 ± 33.50) pg/ml and Tu686 cells with no transfection (385.30 ± 33.50) pg/ml (F = 17.091, P < 0.01). The expression of phosphorylated p38 MAPK was obviously increased in Tu686 cells with EphA2 overexpression. SB203580 inhibited the expressions of VEGF and phosphorylated p38 MAPK proteins in Tu686 cells with EphA2 overexpression. CONCLUSION: EphA2 can regulate the expression of VEGF protein and stimulate p38 MAPK signaling pathway.


Assuntos
Carcinoma de Células Escamosas/metabolismo , Neoplasias de Cabeça e Pescoço/metabolismo , Sistema de Sinalização das MAP Quinases , Receptor EphA2/fisiologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Linhagem Celular Tumoral , Inibidores Enzimáticos/farmacologia , Humanos , Imidazóis/farmacologia , Piridinas/farmacologia , Carcinoma de Células Escamosas de Cabeça e Pescoço , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
4.
J Proteome Res ; 11(2): 1100-7, 2012 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-22070577

RESUMO

The present study was to identify and quantitate differentially expressed proteins in laryngeal squamous cell carcinoma (LSCC) tissues with or without lymph node metastasis and to explore transcriptional factors and regulation networks associated with the process. Tissue specimens were taken from 20 patients with LSCC, including 10 cases of LSCC without metastasis LSCC (N0) and 10 cases of LSCC with metastasis LSCC (Nx). Among the 643 unique proteins identified by using iTRAQ labeling and quantitative proteomic technology, 389 proteins showed an abundance change in LSCC (Nx) as compared to LSCC (N0). Cytoskeleton remodeling, cell adhesion, and immune response activation were found to be the main processes in LSCC metastasis. The construction of transcription regulation networks identified key transcription regulators for lymph node metastasis of LSCC, including Sp1, c-myc, and p53, which may affect LSCC metastasis through the epithelial-mesenchymal transition. Furthermore, our results suggest that ubiquitination may be a critical factor in the networks. The present study provides insights into transcriptional factors and regulation networks involved in LSCC metastasis, which may lead to new strategies for treatment of LSCC metastasis.


Assuntos
Carcinoma de Células Escamosas/metabolismo , Neoplasias de Cabeça e Pescoço/metabolismo , Neoplasias Laríngeas/metabolismo , Fatores de Transcrição/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Escamosas/patologia , Adesão Celular , Citoesqueleto/metabolismo , Regulação para Baixo , Feminino , Neoplasias de Cabeça e Pescoço/patologia , Humanos , Neoplasias Laríngeas/patologia , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Mapas de Interação de Proteínas , Proteínas Proto-Oncogênicas c-myc/metabolismo , Fator de Transcrição Sp1/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço , Proteína Supressora de Tumor p53/metabolismo , Regulação para Cima
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