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Endothelial-specific intron-derived miR-126 is down-regulated in human breast cancer and targets both VEGFA and PIK3R2

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Abstract

Endothelial cells are the key components of vascular intima and play pivotal roles in vasculogenesis, angiogenesis, and tumor growth. Using Northern blot and real-time PCR, we confirmed that miR-126 and its host gene EGF-like domain 7 (EGFL7) were widely expressed in rat tissues but strictly expressed in endothelial cells. In mammals, miR-126 gene is embedded in intron7 of EGFL7. To explore the biogenesis of miR-126, plasmid EGFL7(126)-pEGFPc1 containing segment of exon7-intron7-exon8 of EGFL7 was constructed and expressed in 293T. Expression of spliced exon7–8 and excised mature miR-126 was detected by PCR and Northern blot. Knocking-down of endothelial endogenous miR-126 did not affect EGFL7 expression at mRNA or protein level. To investigate the possible roles of miR-126, PicTar, miRBase, miRanda, Bibiserv, and Targetscan were used to screen the targets. VEGFA and PIK3R2 were confirmed as the targets of miR-126 by luciferase reporter assay and Western blot. Interestingly, Northern blot and western blot showed that miR-126 was down-regulated in breast tumors where the VEGF/PI3K/AKT signaling pathway was activated. Introduction of miR-126 mimics into MCF-7 could effectively decrease VEGF/PI3K/AKT signaling activity. In summary, miR-126 was strictly expressed in endothelial cells and excised from EGFL7 pre-mRNA without affecting splicing and expression of its host gene. In addition, miR-126 could target both VEGFA and PIK3R2, and its expression was decreased in human breast cancer, implying that miR-126 may play a role in tumor genesis and growth by regulating the VEGF/PI3K/AKT signaling pathway.

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Acknowledgments

This study was supported by the grants from the Major State Basic Research Development Program of the People’s Republic of China (Nos. 2009CB521902, 2009CB521907, 2007CB947002, and 2006CB503807) and the National Natural Science Foundation of China (Nos. 30971231, 30800375, 30670760, and 30570397).

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Authors and Affiliations

  1. Department of Cardiology, Changhai Hospital, 168 Changhai Road, Shanghai, 200433, People’s Republic of China

    Ni Zhu, Yuan Bai, Qing Jing & Yongwen Qin

  2. Department of Physiology and Key lab of Ministry of Education in Fertility Preservation and Maintenance, Ningxia Medical University, 1160 Shengli Road, Yinchuan, 750004, People’s Republic of China

    Dongze Zhang, Haoping Xie, Zhe Zhou, Huyan Chen, Tiantian Hu & Wenjun Yuan

  3. Department of Breast Surgery, Changhai Hospital, 168 Changhai Road, Shanghai, 200433, People’s Republic of China

    Yuan Shen

  4. Department of Physiology, Second Military Medical University, 800 Xiangyin Road, Shanghai, 200433, People’s Republic of China

    Wenjun Yuan

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  1. Ni Zhu
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  2. Dongze Zhang
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  3. Haoping Xie
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  4. Zhe Zhou
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  8. Yuan Shen
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  10. Qing Jing
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Correspondence to Qing Jing or Yongwen Qin.

Electronic supplementary material

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11010_2011_723_MOESM1_ESM.ppt

Figure. S1 (A) The sequence of miR-126-3p and 5p are highly conserved in human, rat and mouse, fish and chicken. (B) Location of miR-126 in EGFL7 in human, mouse and rat. Figure. S2 Green fluorescence was observed in 293T which were transfected with pEGFPc1-EGFL7(126) (A) and pEGFPc1(B) after 24 h. (C) Luciferase activity was measured in 293T which were co-transfected of PGL3-VEGFA or PGL3-PIK3R2 with pEGFPc1-EGFL7(126) or pEGFPc1. Firefly luciferase was normalized to renilla luciferase. * P < 0.05 compares to controls. (PPT 343 kb)

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Zhu, N., Zhang, D., Xie, H. et al. Endothelial-specific intron-derived miR-126 is down-regulated in human breast cancer and targets both VEGFA and PIK3R2. Mol Cell Biochem 351, 157–164 (2011). https://doi.org/10.1007/s11010-011-0723-7

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  • DOI: https://doi.org/10.1007/s11010-011-0723-7

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