Coordinated effects of microRNA-494 induce G2/M arrest in human cholangiocarcinoma

Sumitaka Yamanaka, Nathaniel R. Campbell, Fangmei An, Scot C. Kuo, James J. Potter, Esteban Mezey, Anirban Maitra, Florin M. Selaru

Research output: Contribution to journalArticlepeer-review

Abstract

MicroRNA (miRs) have emerged as salient regulators in cancer homeostasis and, recently, as putative therapeutics. Cholangiocarcinomas (CCA) are aggressive cancers with survival usually measured in months. mRNA arrays followed by pathway analysis revealed that miR-494 is a major modulator of the cell cycle progression from gap 2 (G2) to mitosis (M). We performed fluorescence activated cell sorting (FACS) as well as differential interference contrast (DIC) microscopy, and confirmed that miR-494 induces a significant arrest in G2/M in CCA cells. Furthermore, we verified that miR-494 modulates the protein level of six genes involved in the G2/M transition: Polo-like Kinase 1 (PLK1), pituitary tumor-transforming gene 1 (PTT G1), Cyclin B1 (CCNB1), cell-division cycle 2 (CDC2), cell-division cycle 20 (CDC20) and topoisomerase II α(TOP 2A). Next, we identified direct binding of miR-494 to the open reading frame (ORF) and downregulation of PTT G1 and TOP 2A. In summary, our findings suggest that miR-494 has a global regulatory role in cell cycle progression, exerted by concerted effects on multiple proteins involved in gap 1 (G1) to synthesis (S), as described previously, as well as G2 to M progression. Therefore, it appears that the simultaneous effects of a single miR species on multiple targets along the same canonical pathway is advantageous for the usage of miRs as therapeutics. In addition, our data suggest that miRs act within a narrow range. miR expression above the upper threshold does not appear to induce further effects, which is reassuring in terms of off-target effects of miR surrounding noncancerous tissue.

Original languageEnglish (US)
Pages (from-to)2729-2738
Number of pages10
JournalCell Cycle
Volume11
Issue number14
DOIs
StatePublished - Jul 15 2012

Keywords

  • Cholangiocarcinoma
  • Global regulation
  • miR-494

ASJC Scopus subject areas

  • Molecular Biology
  • Developmental Biology
  • Cell Biology

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