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Functional Characterization and Clinical Significance of PKNOX2, a Novel Tumor Suppressor in Gastric Cancer
Başlık:
Functional Characterization and Clinical Significance of PKNOX2, a Novel Tumor Suppressor in Gastric Cancer
Yazar:
Zhang, Li, author.
ISBN:
9780438106369
Yazar Ek Girişi:
Fiziksel Tanımlama:
1 electronic resource (134 pages)
Genel Not:
Source: Dissertation Abstracts International, Volume: 79-10(E), Section: B.
Özet:
Backgrounds: Using 450K DNA methylation array, we identified PBX/Knotted Homeobox 2 (PKNOX2) as a novel gene differentially methylated in gastric cancer (GC). PKNOX2 belongs to the TALE (three-amino acid loop extension) family of homedomain transcription factors, which play fundamental roles in cell proliferation, differentiation and death. However, the role of PKNOX2 in GC remains largely unknown.
Aims & Methods: We aimed to investigate the epigenetic regulation, biological function, molecular mechanism and clinical implication of PKNOX2 in GC.PKNOX2 promoter methylation was evaluated by bisulfite sequencing. PKNOX2 expression was determined by RT-PCR and qPCR.Biological functions of PKNOX2 were explored in GC cells by MTT, colony formation, apoptosis, cell cycle, cell migration/invasion assays, and in vivo using nude mice tumorigenicity assay. Pathways targeted by PKNOX2 were identified by Cancer pathway PCR array and dual luciferase reporter assay as well as human p53 signaling pathway array. The binding targets of PKNOX2 were predicted with the JASPAR database and confirmed by ChIP-PCR.
Results: PKNOX2 was silenced in 15 out of 18 GC cell lines through promoter methylation, and 5'-Azadeoxycytidine treatment restored PKNOX2 expression. PKNOX2 mRNA expression was down-regulated in GC compared to adjacent normal tissues (P<0.001). Moreover, promoter methylation of PKNOX2 was associated with poor survival in GC patients (P<0.05), suggesting that PKNOX2 may function as a tumor suppressor in GC. We therefore examined its functional effect in GC. Ectopic expression of PKNOX2 in GC cell lines (AGS and MKN45) suppressed cell proliferation, induced cell apoptosis and cell cycle arrest, concomitant with increased expression of apoptosis markers cleaved caspase-9,-7,-3 and PARP and cell cycle inhibitors p53, p21 and p27. PKNOX2 also attenuated cell migration and invasion by inhibiting epithelial-mesenchymal transition. PKNOX2 knockdown in GES1 cell resulted in opposite effects. Tumorigenicity assay in nude mice showed that stable PKNOX2 expression significantly suppressed tumor growth in vivo. To probe the mechanism of action of PKNOX2 in GC, we performed Human Cancer pathway PCR array, which unveiled profound up-regulation (>6-fold) of Insulin like Growth Factor Binding Protein 5 (IGFBP5) in PKNOX2-overexpressing GC cells. IGFBP5 knockdown partly abolished the growth inhibitory effect of PKNOX2 in GC cells, indicating that IGFBP5 mediated the tumor suppressive function of PKNOX2. Chromatin immunoprecipitation (ChIP)-PCR assay showed that PKNOX2 bind directly to IGFBP5 promoter to mediate transcription. Consistent with our data, PKNOX2 expression was positively correlated with IGFBP5 expression in the TCGA GC dataset. IGFBP5 mediated the tumor suppressive effect of PKNOX2 via activating p53 signaling pathway, as determined by western blot and luciferase reporter assays. PKNOX2 can also directly bind to p53 promoter region as confirmed by ChIP-PCR. Consequently, p53 transcription target genes were coordinately up-regulated in PKNOX2 overexpressing cells, leading to tumor suppression.
Conclusions: PKNOX2 functions as a novel tumor suppressor silenced in GC by promoter methylation. Its tumor suppressive effect is mediated via IGFBP5 and the activation of p53 signaling pathway. Promoter methylation of PKNOX2 may be a useful biomarker for predicting patient prognosis.
Notlar:
School code: 1307
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Yer Numarası | Demirbaş Numarası | Shelf Location | Lokasyon / Statüsü / İade Tarihi |
---|---|---|---|
XX(687335.1) | 687335-1001 | Proquest E-Tez Koleksiyonu | Arıyor... |
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