Hypoxia-responsive miRNAs target argonaute 1 to promote angiogenesis
Zhen Chen, … , Michael Hsiao, John Y-J. Shyy
Zhen Chen, … , Michael Hsiao, John Y-J. Shyy
Published February 15, 2013
Citation Information: J Clin Invest. 2013;123(3):1057-1067. https://doi.org/10.1172/JCI65344.
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Research Article Oncology

Hypoxia-responsive miRNAs target argonaute 1 to promote angiogenesis

Abstract

Despite a general repression of translation under hypoxia, cells selectively upregulate a set of hypoxia-inducible genes. Results from deep sequencing revealed that Let-7 and miR-103/107 are hypoxia-responsive microRNAs (HRMs) that are strongly induced in vascular endothelial cells. In silico bioinformatics and in vitro validation showed that these HRMs are induced by HIF1α and target argonaute 1 (AGO1), which anchors the microRNA-induced silencing complex (miRISC). HRM targeting of AGO1 resulted in the translational desuppression of VEGF mRNA. Inhibition of HRM or overexpression of AGO1 without the 3′ untranslated region decreased hypoxia-induced angiogenesis. Conversely, AGO1 knockdown increased angiogenesis under normoxia in vivo. In addition, data from tumor xenografts and human cancer specimens indicate that AGO1-mediated translational desuppression of VEGF may be associated with tumor angiogenesis and poor prognosis. These findings provide evidence for an angiogenic pathway involving HRMs that target AGO1 and suggest that this pathway may be a suitable target for anti- or proangiogenesis strategies.

Authors

Zhen Chen, Tsung-Ching Lai, Yi-Hua Jan, Feng-Mao Lin, Wei-Chi Wang, Han Xiao, Yun-Ting Wang, Wei Sun, Xiaopei Cui, Ying-Shiuan Li, Tzan Fang, Hongwei Zhao, Chellappan Padmanabhan, Ruobai Sun, Danny Ling Wang, Hailing Jin, Gar-Yang Chau, Hsien-Da Huang, Michael Hsiao, John Y-J. Shyy

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Figure 3

Posttranscriptional targeting of AGO1 mRNA in AGO1-mediated miRISC.

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Posttranscriptional targeting of AGO1 mRNA in AGO1-mediated miRISC. (A...
(A) HEK293 cells were transfected with the WT Luc-AGO1–3′ UTR (WT), Luc-AGO1–3′ UTR with miR-103/107 or Let-7 target sites mutated (mut), or Luc-AGO2–3′ UTR, together with pre–Let-7e (40 nM), pre-103 (40 nM), pre–Let-7e and pre-103 (20 nM each), or control RNA (40 nM). (B) Bovine aortic ECs (BAECs) transfected with Luc-AGO1–3′ UTR or -AGO2–3′ UTR were subjected to normoxia (Nx) or hypoxia (Hx). CMV–β-gal was cotransfected in all groups as a transfection control. Luciferase activity was normalized to that of β-gal. (C–G) HUVECs were subjected to normoxia or hypoxia. (C and D) Western blot and qPCR analyses of protein and mRNA levels of AGO1–3. (E–G) AGO1 was immunoprecipitated from cell lysates. The immunoprecipitates were subjected to AGO1 immunoblotting (F) and the AGO1-associated miRNAs and AGO1 mRNA were quantified by qPCR (E and G). Data represent mean ± SD from 3 independent experiments. *P < 0.05 compared with control RNA group in A or normoxia group in B–G.