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The glycosyltransferase LARGE2 is repressed by Snail and ZEB1 in prostate cancer.

ABSTRACT: Reductions in both expression of the dystroglycan core protein and functional glycosylation of the ?-dystroglycan (?DG) subunit have been reported in a number of cancers and may contribute to disease progression. In the case of prostate cancer, one mechanism that contributes to ?DG hypoglycosylation is transcriptional down-regulation of LARGE2 (GYLTY1B), a glycosyltransferase that produces the functional (laminin-binding) glycan on ?DG, but the mechanism(s) underlying reduction of LARGE2 mRNA remain unclear. Here, we show that ?DG hypoglycosylation is associated with epithelial-to-mesenchymal transition (EMT)-like status. We examined immunoreactivity for both functionally-glycosylated ?DG and E-cadherin by flow cytometry and the relative expression of ZEB1 mRNA and the ?DG glycosyltransferase LARGE2 mRNA in prostate and other cancer cell lines by quantitative RT-PCR. To study the role of ZEB1 and other transcription factors in the regulation of LARGE2, we employed overexpression and knockdown approaches. Snail- or ZEB1-driven EMT caused ?DG hypoglycosylation by repressing expression of the LARGE2 mRNA, with both ZEB1-dependent and -independent mechanisms contributing to Snail-mediated LARGE2 repression. To examine the direct regulation of LARGE2 by Snail and ZEB1 we employed luciferase reporter and chromatin immunoprecipitation assays. Snail and ZEB1 were found to bind directly to the LARGE2 promoter, specifically to E/Z-box clusters. Furthermore, analysis of gene expression profiles of clinical samples in The Cancer Genome Atlas reveals negative correlation of LARGE2 and ZEB1 expression in various cancers. Collectively, our results suggest that LARGE2 is negatively regulated by Snail and/or ZEB1, revealing a mechanistic basis for ?DG hypoglycosylation during prostate cancer progression and metastasis.

SUBMITTER: Huang Q

PROVIDER: S-EPMC4623020 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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The glycosyltransferase LARGE2 is repressed by Snail and ZEB1 in prostate cancer.

Huang Qin Q Miller Michael R MR Schappet James J Henry Michael D MD

Cancer biology & therapy 20150101 1

Reductions in both expression of the dystroglycan core protein and functional glycosylation of the α-dystroglycan (αDG) subunit have been reported in a number of cancers and may contribute to disease progression. In the case of prostate cancer, one mechanism that contributes to αDG hypoglycosylation is transcriptional down-regulation of LARGE2 (GYLTY1B), a glycosyltransferase that produces the functional (laminin-binding) glycan on αDG, but the mechanism(s) underlying reduction of LARGE2 mRNA re ...[more]

PMID: 25455932

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Project description:BACKGROUND:Recent evidence indicates that metformin inhibits mammalian cancer growth and metastasis through the regulation of microRNAs. Metformin regulates miR-381 stability, which plays a vital role in tumor progression. Moreover, increased YAP expression and activity induce non-small cell lung cancer (NSCLC) tumor growth and metastasis. However, the molecular mechanism underpinning how metformin-induced upregulation of miR-381 directly targets YAP or its interactions with the epithelial-mesenchymal transition (EMT) marker protein Snail in NSCLC is still unknown. METHODS:Levels of RNA and protein were analyzed using qPCR, western blotting and immunofluorescence staining. Cellular proliferation was detected using a CCK8 assay. Cell migration and invasion were analyzed using wound healing and transwell assays. Promoter activity and transcription were investigated using the luciferase reporter assay. Chromatin immunoprecipitation was used to detect the binding of YAP to the promoter of Snail. The interaction between miR-381 and the 3'UTR of YAP mRNA was analyzed using the MS2 expression system and co-immunoprecipitation with biotin. RESULTS:We observed that miR-381 expression is negatively correlated with YAP expression and plays an opposite role to YAP in the regulation of cellular proliferation, invasion, migration, and EMT of NSCLC cells. The miR-381 function as a tumor suppressor was significantly downregulated in lung cancer tissue specimens and cell lines, which decreased the expression of its direct target YAP. In addition, metformin decreased cell growth, migration, invasion, and EMT via up-regulation of miR-381. Moreover, YAP, which functions as a co-transcription factor, enhanced NSCLC progression and metastasis by upregulation of Snail. Snail knockdown downregulated the mesenchymal marker vimentin and upregulated the epithelial marker E-cadherin in lung cancer cells. Furthermore, miR-381, YAP, and Snail constitute the miR-381-YAP-Snail signal axis, which is repressed by metformin, and enhances cancer cell invasiveness by directly regulating EMT. CONCLUSIONS:Metformin-induced repression of miR-381-YAP-Snail axis activity disrupts NSCLC growth and metastasis. Thus, we believe that the miR-381-YAP-Snail signal axis may be a suitable diagnostic marker and a potential therapeutic target for lung cancer.

Dataset Information

The glycosyltransferase LARGE2 is repressed by Snail and ZEB1 in prostate cancer.

Publications

The glycosyltransferase LARGE2 is repressed by Snail and ZEB1 in prostate cancer.

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