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01.12.2018 | Research | Ausgabe 1/2018 Open Access

Journal of Experimental & Clinical Cancer Research 1/2018

SP1-induced upregulation of lncRNA SPRY4-IT1 exerts oncogenic properties by scaffolding EZH2/LSD1/DNMT1 and sponging miR-101-3p in cholangiocarcinoma

Journal of Experimental & Clinical Cancer Research > Ausgabe 1/2018
Yi Xu, Yue Yao, Xingming Jiang, Xiangyu Zhong, Zhidong Wang, Chunlong Li, Pengcheng Kang, Kaiming Leng, Daolin Ji, Zhenglong Li, Lining Huang, Wei Qin, Yunfu Cui
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Electronic supplementary material

The online version of this article (https://​doi.​org/​10.​1186/​s13046-018-0747-x) contains supplementary material, which is available to authorized users.



Accumulating evidence has indicated that long non-coding RNAs (lncRNAs) behave as a novel class of transcription products during multiple cancer processes. However, the mechanisms responsible for their alteration in cholangiocarcinoma (CCA) are not fully understood.


The expression of SPRY4-IT1 in CCA tissues and cell lines was determined by RT-qPCR, and the association between SPRY4-IT1 transcription and clinicopathologic features was analyzed. Luciferase reporter and chromatin immunoprecipitation (ChIP) assays were performed to explore whether SP1 could bind to the promoter region of SPRY4-IT1 and activate its transcription. The biological function of SPRY4-IT1 in CCA cells was evaluated both in vitro and in vivo. ChIP, RNA binding protein immunoprecipitation (RIP) and luciferase reporter assays were performed to determine the molecular mechanism of SPRY4-IT1 in cell proliferation, apoptosis and invasion.


SPRY4-IT1 was abnormally upregulated in CCA tissues and cells, and this upregulation was correlated with tumor stage and tumor node metastasis (TNM) stage in CCA patients. SPRY4-IT1 overexpression was also an unfavorable prognostic factor for patients with CCA. Additionally, SP1 could bind directly to the SPRY4-IT1 promoter region and activate its transcription. Furthermore, SPRY4-IT1 silencing caused tumor suppressive effects via reducing cell proliferation, migration and invasion; inducing cell apoptosis and reversing the epithelial-to-mesenchymal transition (EMT) process in CCA cells. Mechanistically, enhancer of zeste homolog 2 (EZH2) along with the lysine specific demethylase 1 (LSD1) or DNA methyltransferase 1 (DNMT1) were recruited by SPRY4-IT1, which functioned as a scaffold. Importantly, SPRY4-IT1 positively regulated the expression of EZH2 through sponging miR-101-3p.


Our data illustrate how SPRY4-IT1 plays an oncogenic role in CCA and may offer a potential therapeutic target for treating CCA.
Additional file 5: Figure S2. A The interaction of SPRY4-IT1 and potential RNA binding proteins was predicted on https://​pridb.​gdcb.​iastate.​edu/​RPISeq/​. B U1 binding with SNRNP70 was used as a positive control in RIP assays using HuCCT1 and RBE cell extracts. C-D Association analysis of the relationship between SPRY4-IT1 and KLF2 /LATS2 expression levels in 20 paired CCA tissues. E-G EZH2, LSD1 and DNMT1 levels were detected in cells transfected with EZH2, LSD1 or DNMT1 siRNA or si-NC. **P < 0.01. (TIFF 770 kb)
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