Hepatitis C virus (HCV) nonstructural protein 4B (NS4B) is a multi-transmembrane protein, but little is known about how NS4B contributes to HCV replication and tumorigenesis. Its C-terminal domain (CTD) has been shown to associate with intracellular membrane, and we have previously shown that NS4B CTD contains a class I PDZ-binding motif (PBM). Here, we demonstrated that NS4B PBM interacts with the PDZ-containing tumor suppressor protein, Scribble, using immunofluorescence and co-immunoprecipitation assays, and this interaction requires at least three contiguous PDZ domains of Scribble. In addition, NS4B PBM specifically induced Scribble degradation by activating the proteasome-ubiquitin pathway. Similar Scribble degradation was also observed in HCV-infected cells, suggesting NS4B could work in the context of HCV. Finally, NS4B PBM mutants showed reduced colony formation capacity compared with its wild-type counterpart, indicating that NS4B PBM plays important roles in NS4B-mediated cell transformation. Altogether, we provide a mechanism by which NS4B induces cell transformation through its PBM, which specifically interacts with the PDZ domains of Scribble and targets Scribble for degradation.
// Lina Wu 1, * , Bo Hu 2, * , Bingtian Zhao 3, * , Yinan Liu 2 , Yue Yang 2 , Lijian Zhang 2 and Jinfeng Chen 2 1 Central Laboratory, Key Laboratory of Carcinogenesis and Translational Research, Ministry of Education, Beijing Cancer Hospital & Institute, Peking University Cancer Hospital, Beijing, China 2 Key Laboratory of Carcinogenesis and Translational Research, Ministry of Education, Department of Thoracic Surgery II, Peking University School of Oncology, Beijing Cancer Hospital & Institute, Beijing, China 3 Department of Respiratory & Critical Care Medicine, Tianjin Chest Hospital, Tianjin, China * These authors have contributed equally to this work Correspondence to: Jinfeng Chen, email: chenjinfengdoctor@163.com Keywords: microRNA, miR-422a, lung cancer, lymphatic metastasis, biomarker Received: August 15, 2016 Accepted: January 10, 2017 Published: February 02, 2017 ABSTRACT To identify specific circulating microRNAs that were associated with the lymphatic metastasis in lung cancer, we performed miRNA microarray analysis of lymph node with and without metastasis from five lung cancer patients. Top six differentially expressed miRNAs were selected for further validation. A training cohort of 26 patients with lung cancer was firstly recruited and the selected miRNAs in the plasma samples were investigated. miRNA-422a, with highest diagnostic accuracy in lymphatic metastasis was identified (AUC, area under the receiver operating characteristic curve, 0.744; 95%CI, 0.570-0.918). The diagnostic value of miR-422a was also demonstrated by a validation cohort of 51 lung cancer patients (AUC, 0.880; 95%CI, 0.787-0.972). Moreover, a high diagnostic value was also observed after integrated analysis of training and validation cohorts (AUC, 0.792; 95%CI, 0.688-0.896). The odds ratio of high miR-422a expression for lymphatic metastasis in lung cancer was 13.645 (95%CI, 2.677-69.553) after adjustment of the potential confounding factors. Furthermore, we predicted the target genes of miR-422a by combining the online database, miRcords, and the data from GEO and TCGA. Sixty-one target genes of miR-422a that might be involved in lymphatic metastasis in lung cancer were identified. And GO analysis suggested multiple target genes relatively concentrated in the biological processes of apoptosis, transport, and protein phosphorylation.
Background: aFGF content in serum and cerebrospinal fluid is increased in Alzheimer's disease (AD) patients and attenuates the activation of astrocytes.Extracellular vesicles (EVs) are a major mediator in astrocyte-neuron communications.Since excessive or persistent reactive astrocytes lead to chronic inflammation and neuronal dysfunction, and the activation of astrocytes can be inhibited by aFGF, we proposed that the cargoes of astrocyte-derived EVs (AEVs) might be modified by aFGF stimulation, playing an important role in AD progression.However, the mechanisms underlying the role of aFGF remain unclear.Methods: AEVs were isolated from damaged astrocytes, treated with or without aFGF in Aβ-loading condition, and were intranasally administered to AD mice.We determined the ability of AEVs to enter the brain, ameliorate cognitive behavior deficits, alleviate the Aβ burden in the brain, and improve synapse ultrastructure.Subsequently, the miRNAs enriched in AEVs were sequenced to identify the key molecules specifically modified by aFGF.Finally, we explored the protective effects of miR-206-3p inhibition on cognitive deficiency and its regulatory mechanism and determined its role as a specific biomarker for potential AD diagnosis.Results: AEVs stimulated by aFGF (defined as AEVs-A β+H ) had favorable neuroprotection in AD pathology by enhancing neurite growth and reduction of Aβ loading on neurons in vitro.Following intranasal administration, AEVs-A β+H ameliorated cognitive behavior deficits, promoted synaptic plasticity, and alleviated brain Aβ burden in the APP/PS1 and Aβ brain-injected mice.AEVs-A β+H showed beneficial effects on AD similar to AEVs produced in normal situations (AEVs-Ctrl).aFGF stimulation modified the cargoes in EVs derived from Aβ damaged astrocytes, the most significant of which being the down-regulation of miR-206-3p.The miR-206-3p level was specifically high in the plasma of AD mice and patients, and miR-206-3p antagomir reversed the Alzheimer phenotype in AD mice.The brain-derived neurotrophic factor (BDNF) gene was negatively regulated by miR-206-3p and upregulated by AEVs-A β+H and miR-206-3p antagomir in AD mice.AEVs-A β+H inhibited δ-secretase (Asparagine endopeptidase, AEP) activation via the miR-206-3p/BDNF axis to alleviate Aβ burden in the AD brain.Conclusion: Our findings highlight the role of aFGF in the modification of AEVs cargoes, especially miR-206-3p that can potentially serve as a biomarker for AD diagnosis and therapeutic target.
Chronic liver inflammation caused by chronic hepatitis B virus (CHB) infection leads to liver cirrhosis and hepatocellular carcinoma. Recently, the role of alanine aminotransferase (ALT) as a predictor of liver inflammation has been questioned. The aim of this study was to investigate the utility of noninvasive fibrosis markers including hyaluronic acid (HA), collagen type IV (CIV), N‐terminal propeptide of type III procollagen (PIIINP), and laminin (LN) in identifying significant liver inflammation in patients with CHB, especially in patients with normal or near‐normal ALT. A total of 242 CHB patients who underwent liver biopsy were enrolled. The serum levels of ALT, aspartate aminotransferase, HA, CIV, PIIINP, and LN were quantified and the relationship between histological staging and serum markers was systematically analyzed. Serum CIV, PIIINP, HA, and LN levels increased significantly along with the increasing severity of liver inflammation. Multivariate analysis showed that CIV and LN were independently associated with significant inflammation. CIV, PIIINP, HA, and LN levels were found to have high diagnostic values for predicting significant inflammation in patients with CHB (area under the curve, AUC = 0.807, 0.795, 0.767, and 0.703, respectively). The combined index for the identification of significant inflammation, including CIV, PIIINP, HA, and LN levels, significantly improved diagnostic performance (AUC = 0.851). Moreover, the combined index also achieved excellent diagnostic accuracy (AUC = 0.861) in patients with CHB with normal or near‐normal ALT. In conclusion, the combined index may be a strong indicator for discriminating significant liver inflammation, especially in patients with CHB with normal or near‐normal ALT.
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