Zebrafish let-7b acts downstream of hypoxia-inducible factor-1α to assist in hypoxia-mediated cell proliferation and cell cycle regulation

Huang, C.X., Chen, N., Wu, X.J., He, Y., Huang, C.H., Liu, H., Wang, W.M., Wang, H.L.
Life sciences   171: 21-29 (Journal)
Registered Authors
He, Yan
Cell cycle, Hif-1α, Let-7b/7f, Proliferation, foxh1, miRNA profile, shRNA
MeSH Terms
  • Animals
  • Cell Cycle/physiology*
  • Cell Proliferation/physiology*
  • Gene Silencing
  • Hypoxia-Inducible Factor 1, alpha Subunit/genetics
  • Hypoxia-Inducible Factor 1, alpha Subunit/physiology*
  • MicroRNAs/physiology*
  • RNA, Messenger/genetics
  • Transcription, Genetic
  • Zebrafish
28077310 Full text @ Life Sci.
Hypoxia-inducible factor-1α (HIF-1α) is a transcriptional regulator of cellular responses to hypoxic stress. MicroRNAs (miRNAs) play an essential role in hypoxia-mediated cellular responses. Previous studies have identified some let-7 family members as hypoxia-regulated miRNAs (HRMs). In the present study, we aimed to investigate whether zebrafish let-7b/7f contribute cellular hypoxic response in a Hif-1α-dependent manner.
Stable suppression of zebrafish hif-1α was achieved by microinjection of an optimized short-hairpin RNA (shRNA) expression vector. Next-generation sequencing was conducted to characterize miRNA and mRNA expression profiles. MiRNA promoter analysis and target detection was performed by dual-luciferase assay. Quantitative real-time PCR (qRT-PCR) and western blot were used to determine the expression of let-7b/7f, Hif-1α and Foxh1. Proliferation of ZF4 cells was examined using Cell Counting Kit-8 (CCK-8) and cell cycle progression was analyzed by flow cytometry assay.
Correlation between 7 miRNAs and 76 putative targets was identified based on integrated analysis of miRNA-mRNA profiles. Let-7b and let-7f were further considered as potential HRMs, with let-7b further validated as Hif-1α up-regulated. In addition, Forkhead-box H1 (Foxh1) was confirmed as a bona fide downstream target of let-7b. Furthermore, overexpression of both let-7b and let-7f repressed cell proliferation through blocking cell cycle progression of the G1-S transition.
Our findings for the first time suggest zebrafish let-7b acts downstream of Hif-1α to assist in hypoxia-mediated cell proliferation and cell cycle regulation at least in part through the downregulation of foxh1. We also identified 4 novel potential HIF-1α-regulated miRNAs in zebrafish.
Genes / Markers
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Engineered Foreign Genes