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01.12.2018 | Research | Ausgabe 1/2018 Open Access

Molecular Cancer 1/2018

The Jun/miR-22/HuR regulatory axis contributes to tumourigenesis in colorectal cancer

Zeitschrift:
Molecular Cancer > Ausgabe 1/2018
Autoren:
Yanqing Liu, Xiaorui Chen, Rongjie Cheng, Fei Yang, Mengchao Yu, Chen Wang, Shufang Cui, Yeting Hong, Hongwei Liang, Minghui Liu, Chihao Zhao, Meng Ding, Wu Sun, Zhijian Liu, Feng Sun, Chenyu Zhang, Zhen Zhou, Xiaohong Jiang, Xi Chen
Wichtige Hinweise

Electronic supplementary material

The online version of this article (https://​doi.​org/​10.​1186/​s12943-017-0751-3) contains supplementary material, which is available to authorized users.

Abstract

Background

Colorectal cancer (CRC) is a severe health problem worldwide. Clarifying the mechanisms for the deregulation of oncogenes and tumour suppressors in CRC is vital for its diagnosis, treatment, prognosis and prevention. Hu antigen R (HuR), which is highly upregulated in CRC, functions as a pivotal oncogene to promote CRC progression. However, the underlying cause of its dysregulation is poorly understood.

Methods

In CRC tissue sample pairs, HuR protein levels were measured by Western blot and immunohistochemical (IHC) staining, respectively. HuR mRNA levels were also monitored by qRT-PCR. Combining meta-analysis and microRNA (miRNA) target prediction software, we predicted miRNAs that targeted HuR. Pull-down assay, Western blot and luciferase assay were utilized to demonstrate the direct binding of miR-22 on HuR’s 3’-UTR. The biological effects of HuR and miR-22 were investigated both in vitro by CCK-8, EdU and Transwell assays and in vivo by a xenograft mice model. JASPAR and SABiosciences were used to predict transcriptional factors that could affect miR-22. Luciferase assay was used to explore the validity of putative Jun binding sites for miR-22 regulation. ChIP assay was performed to test the Jun’s occupancy on the C17orf91 promoter.

Results

We observed a significant upregulation of HuR in CRC tissue pairs and confirmed the oncogenic function of HuR both in vitro and in vivo. We found that an important tumour-suppressive miRNA, miR-22, was significantly downregulated in CRC tissues and inversely correlated with HuR in both CRC tissues and CRC cell lines. We demonstrated that miR-22 directly bound to the 3’-UTR of HuR and led to inhibition of HuR protein, which repressed CRC proliferation and migration in vitro and decelerated CRC xenografted tumour growth in vivo. Furthermore, we found that the onco-transcription factor Jun could inhibit the transcription of miR-22.

Conclusions

Our findings highlight the critical roles of the Jun/miR-22/HuR regulatory axis in CRC progression and may provide attractive potential targets for CRC prevention and treatment.
Zusatzmaterial
Additional file 1: Table S1. Clinical features of colorectal cancer patients. (DOCX 17 kb)
12943_2017_751_MOESM1_ESM.docx
Additional file 2: Table S2. Sequences of siRNAs, probe and primers. (DOCX 16 kb)
12943_2017_751_MOESM2_ESM.docx
Additional file 3: Figure S1. HuR protein is significantly upregulated in CRC tissues and negatively correlated with CRC patient survival. (a) HuR levels in normal colon, normal rectum, colon adenocarcinoma and rectal adenocarcinoma in the TCGA dataset analysed by Oncomine. (b) Kaplan-Meier curve showing the negative correlation of HuR level and CRC patients’ survival. (TIFF 167 kb)
12943_2017_751_MOESM3_ESM.tif
Additional file 4: Figure S2. The efficiencies of siRNA and overexpression vector of HuR (a) or Jun (b). (TIFF 377 kb)
12943_2017_751_MOESM4_ESM.tif
Additional file 5: Figure S3. HuR functions as an oncogene in CRC. (a-c) HuR promoted SW480 proliferation. a: CCK-8 assays; b and c: EdU assays. (d and e) HuR promoted SW480 migration. (f-h) HuR accelerated CRC xenografted tumour growth. f: Photos of CRC tumours; g: Tumour volume curves; h: Tumour weights. (i) Western blot analysis of HuR levels in CRC xenografted tumours. (j and k) HE staining and IHC staining for HuR and Ki-67 in xenografted tumours. **P < 0.01; ***P < 0.001. (TIFF 5991 kb)
12943_2017_751_MOESM5_ESM.tif
Additional file 6: Figure S4. (a and b) The Kaplan-Meier curve revealed the positive correlation of miR-22 (a) or miR-129 (b) level and CRC patients’ survival. (c and d) The transfection efficiencies of miR-22 (c) or miR-129 (d) mimics or inhibitors in 3 CRC cell lines. ***P < 0.001. (TIFF 1147 kb)
12943_2017_751_MOESM6_ESM.tif
Additional file 7: Figure S5. Pri-miR-22 is downregulated in CRC tissues compared with that in normal adjacent tissues. *P < 0.05. (TIFF 186 kb)
12943_2017_751_MOESM7_ESM.tif
Literatur
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