MiR-124 acts as a tumor suppressor by inhibiting the expression of sphingosine kinase 1 and its downstream signaling in head and neck squamous cell carcinoma

Yuan Zhao, Zhiqiang Ling, Yubin Hao, Xiaowu Pang, Xianlin Han, Joseph A. Califano, Liang Shan, Xinbin Gu

Research output: Contribution to journalArticlepeer-review

31 Scopus citations

Abstract

By analyzing the expression profile of microRNAs in head and neck squamous cell carcinomas (HNSCC), we found that the expression level of miR-124 was 4.59-fold lower in tumors than in normal tissues. To understand its functions, we generated a miR-124-expressing subline (JHU-22miR124) and a mock vector-transfected subline (JHU-22vec) by transfecting the mimic of miR-124 into JHU-22 cancer cells. Restored expression of miR-124 in JHU-22miR124 cells led to reduced cell proliferation, delayed colony formation, and decreased tumor growth, indicating a tumor-suppressive effect of miR-124. Subsequent target search revealed that the 3'-UTR of SphK1 mRNA carries a complementary site for the seed region of miR-124. SphK1 was also detected to be overexpressed in HNSCC cell lines, but down-expressed in JHU-22miR124 cells and tumor xenografts. These results suggest that SphK1 is a target of miR-124. To confirm this finding, we constructed a 3'-UTR-Luc-SphK1 vector and a binding sitemutated luciferase reporter vector. Co-transfection of 3'-UTR-Luc-SphK1 with miR-124 expression vector exhibited a 9-fold decrease in luciferase activity compared with mutated vector, suggesting that miR-124 inhibits SphK1 activity directly. Further studies on downstream signaling demonstrated accumulation of ceramide, increased expression of the pro-apoptotic Bax, BAD and PARP, decreased expression of the antiapoptotic Bcl-2 and Bcl-xL, and enhanced expression of cytochrome c and caspase proteins in JHU-22miR124 compared with JHU-22vec cells and tumor xenografts. We conclude that miR-124 acts as a tumor suppressor in HNSCC by directly inhibiting SphK1 activity and its downstream signals.

Original languageEnglish (US)
Pages (from-to)25005-25020
Number of pages16
JournalOncotarget
Volume8
Issue number15
DOIs
StatePublished - 2017

Keywords

  • BcL-2 family
  • Ceramide
  • Head and neck cancer
  • MiR-124
  • Sphingosine kinase 1

ASJC Scopus subject areas

  • Oncology

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