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Dicer-dependent endothelial microRNAs are necessary for postnatal angiogenesis.

ABSTRACT: Posttranscriptional gene regulation by microRNAs (miRNAs) is important for many aspects of development, homeostasis, and disease. Here, we show that reduction of endothelial miRNAs by cell-specific inactivation of Dicer, the terminal endonuclease responsible for the generation of miRNAs, reduces postnatal angiogenic response to a variety of stimuli, including exogenous VEGF, tumors, limb ischemia, and wound healing. Furthermore, VEGF regulated the expression of several miRNAs, including the up-regulation of components of the c-Myc oncogenic cluster miR-17-92. Transfection of endothelial cells with components of the miR-17-92 cluster, induced by VEGF treatment, rescued the induced expression of thrombospondin-1 and the defect in endothelial cell proliferation and morphogenesis initiated by the loss of Dicer. Thus, endothelial miRNAs regulate postnatal angiogenesis and VEGF induces the expression of miRNAs implicated in the regulation of an integrated angiogenic response.

SUBMITTER: Suarez Y

PROVIDER: S-EPMC2544582 | biostudies-literature | 2008 Sep

REPOSITORIES: biostudies-literature

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Dicer-dependent endothelial microRNAs are necessary for postnatal angiogenesis.

Suárez Yajaira Y Fernández-Hernando Carlos C Yu Jun J Gerber Scott A SA Harrison Kenneth D KD Pober Jordan S JS Iruela-Arispe M Luisa ML Merkenschlager Matthias M Sessa William C WC

Proceedings of the National Academy of Sciences of the United States of America 20080908 37

Posttranscriptional gene regulation by microRNAs (miRNAs) is important for many aspects of development, homeostasis, and disease. Here, we show that reduction of endothelial miRNAs by cell-specific inactivation of Dicer, the terminal endonuclease responsible for the generation of miRNAs, reduces postnatal angiogenic response to a variety of stimuli, including exogenous VEGF, tumors, limb ischemia, and wound healing. Furthermore, VEGF regulated the expression of several miRNAs, including the up-r ...[more]

PMID: 18779589

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Project description:BackgroundVascular endothelial growth factor receptor-2 (VEGFR-2) signaling is an obligate requirement for normal development and pathological angiogenesis such as cancer and age-related macular degeneration. Although autophosphorylation of tyrosine 1173 (Y1173) of VEGFR-2 is considered a focal point for its angiogenic signal relay, however, the mechanism of phosphorylation of Y1173, signaling proteins that are recruited to this residue and their role in angiogenesis is not fully understood.Methodology/principal findingsIn this study we demonstrate that c-Src kinase directly through its Src homology 2 (SH2) domain and indirectly via c-Cbl binds to phospho-Y1057 of VEGFR-2. Activation of c-Src kinase by a positive feedback mechanism phosphorylates VEGFR-2 at multi-docking site, Y1173. c-Src also catalyzes tyrosine phosphorylation of IQGAP1 and acts as an adaptor to bridge IQGAP1 to VEGFR-2. In turn, IQGAP1 activates b-Raf and mediates proliferation of endothelial cells. Silencing expression of IQGAP1 and b-Raf revealed that their activity is essential for VEGF to stimulate angiogenesis in an in vivo angiogenesis model of chicken chorioallantoic membrane (CAM).Conclusions/significanceAngiogenesis contributes to the pathology of numerous human diseases ranging from cancer to age-related macular degeneration. Determining molecular mechanism of tyrosine phosphorylation of VEGFR-2 and identification of molecules that are relaying its angiogenic signaling may identify novel targets for therapeutic intervention against angiogenesis-associated diseases. Our study shows that recruitment and activation of c-Src by VEGFR-2 plays a pivotal role in relaying angiogenic signaling of VEGFR-2; it phosphorylates VEGFR-2 at Y1173, facilitates association and activation of IQGAP1 and other signaling proteins to VEGFR-2. IQGAP1-dependent signaling, in part, is critically required for endothelial cell proliferation, a key step in angiogenesis. Thus, Y1057 of VEGFR-2 serves to regulate VEGFR-2 function in a combinatorial manner by supporting both diversity of recruitment of angiogenic signaling proteins to VEGFR-2, and its ability to promote angiogenesis.

Project description:BackgroundVascular endothelial growth factor receptors (VEGFRs) are the major receptors involved in endothelial cell-dependent tumor angiogenesis. There are studies account for the effects of Hsp90 on angiogenesis, but the role and mechanism of Hsp90β isoforms and NVP-BEP800, a specific inhibitor of Hsp90β, in tumor angiogenesis is rarely mentioned.MethodsImmunohistochemistry and statistical analysis was used to evaluate the correlation between Hsp90β expression, CD31 endothelial cell-dependent vessel density, and VEGFRs expression in tissue samples of 96 HCCs. Kaplan-Meier survival analysis and COX proportional hazards analysis the relation of Hsp90β and prognosis. HUVEC cells were transfected with Hsp90β or treated with NVP-BEP800, and then cell proliferation, migration, invasion and tube formation were investigated. The VEGFR1 and VEGFR2 expression was determined by Western blot and immunofluorescence. The VEGFR1 and VEGFR2 promoter activities were detected by dual luciferase report system. In vivo, the angiogenesis promotion of Hsp90β and anti-angiogenesis efficacy of NVP-BEP800 was tested in HCC xenograft models. Histological analysis was performed on tumor samples to evaluate Hsp90β, VEGFRs expression and MVD.ResultsThis study investigated the correlation between Hsp90β expression and CD31+ endothelial cell-dependent vessel density. Hsp90β promoted VEGFRs expression by increasing their promoter activities. The proliferation, migration, invasion, and tube formation activities of human endothelial cells significantly increased when Hsp90β was overexpressed. NVP-BEP800 down-regulated VEGFRs expression to significantly reduce tubular differentiation, as well as endothelial cell proliferation, migration, and invasion. Furthermore, NVP-BEP800 decreased VEGFR1 and VEGFR2 promoter activities. In vivo, Hsp90β promoted VEGFRs and CD31 expression in human hepatocellular carcinoma tumor xenografts and was associated with increased tumor microvessel density. After 18 days of treatment with 30 mg/kg/day NVP-BEP800, VEGFRs and CD31 expression significantly decreased.ConclusionHsp90β induced endothelial cell-dependent tumor angiogenesis by activating VEGFRs transcription. NVP-BEP800 has potential as a therapeutic strategy for inhibiting tumor angiogenesis by decreasing endothelial cell progression and metastasis. It can help develop a therapeutic strategy for tumor treatment through the inhibition of endothelial cell progression and metastasis.

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Dicer-dependent endothelial microRNAs are necessary for postnatal angiogenesis.

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Dicer-dependent endothelial microRNAs are necessary for postnatal angiogenesis.

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