Requirement of the nuclear localization of transcription enhancer factor 3 for proliferation, migration, tube formation, and angiogenesis induced by vascular endothelial growth factor.
ABSTRACT Transcription enhancer factor 3 (TEF3) is known to regulate the expression of muscle-specific genes and to play important roles in muscle development and diseases. However, little is known about its role in vascular endothelial growth factor (VEGF)-induced angiogenesis. Most recently, we discovered a novel function of TEF3, in which TEF3 is required for the up-regulation of a proangiogenic factor, Down syndrome candidate region 1 isoform 1L (DSCR1-1L), induced by VEGF-A(165) in endothelial cells. Overexpression of TEF3 isoform 1 (TEF3-1) is sufficient to induce DSCR1-1L expression. Here, we report that knocking down the expression of TEF3 almost completely inhibits VEGF-A(165)-induced proliferation, migration, tube formation, formation of F-actin stress fiber, and in vivo Matrigel angiogenesis. This inhibition cannot be rescued by DSCR1-1L overexpression. Further, overexpression of TEF3-1, but not its nuclear localization signal-deletion mutant (TEF3-ΔNLS), induces human umbilical vein endothelial cell proliferation, migration, tube formation, and formation of F-actin stress fiber, even in the absence of VEGF-A(165) stimulation, which is partially inhibited by DSCR1-1L silencing. Our data demonstrate that TEF3, mainly its nuclear localization, is required for VEGF-A(165)-induced endothelial proliferation, migration, tube formation, and in vivo Matrigel angiogenesis.
- [Show abstract] [Hide abstract]
ABSTRACT: Histone modifications are now well-established mediators of transcriptional programs that distinguish cell states. However, the kinetics of histone modification and their role in mediating rapid, signal-responsive gene expression changes has been little studied on a genome-wide scale. Vascular endothelial growth factor A (VEGFA), a major regulator of angiogenesis, triggers changes in transcriptional activity of human umbilical vein endothelial cells (HUVECs). Here we used chromatin immunoprecipitation followed by high throughput sequencing (ChIP-seq) to measure genome-wide changes in histone H3 acetylation at lysine 27 (H3K27ac), a marker of active enhancers, in unstimulated HUVECs and HUVECs stimulated with VEGFA for 1, 4, and 12 hours. We show that sites with the greatest H3K27ac change upon stimulation were associated tightly with EP300, a histone acetyltransferase. Using the variation of H3K27ac as a novel epigenetic signature, we identified transcriptional regulatory elements that are functionally linked to angiogenesis, participate in rapid VEGFA-stimulated changes in chromatin conformation, and mediate VEGFA-induced transcriptional responses. Dynamic H3K27ac deposition and associated changes in chromatin conformation required EP300 activity instead of altered nucleosome occupancy or changes in DNase I hypersensitivity. EP300 activity was also required for a subset of dynamic H3K27ac sites to loop into proximity of promoters. Our study identified thousands of endothelial, VEGFA-responsive enhancers, demonstrating that an epigenetic signature based on the variation of a chromatin feature is a productive approach to define signal-responsive genomic elements. Further, our study implicates global epigenetic modifications in rapid, signal-responsive transcriptional regulation.Genome Research 04/2013; · 14.40 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: In the preimplantation mouse embryo, TEAD4 is critical to establishing the trophectoderm (TE)-specific transcriptional program and segregating TE from the inner cell mass (ICM). However, TEAD4 is expressed in the TE and the ICM. Thus, differential function of TEAD4 rather than expression itself regulates specification of the first two cell lineages. We used ChIP sequencing to define genomewide TEAD4 target genes and asked how transcription of TEAD4 target genes is specifically maintained in the TE. Our analyses revealed an evolutionarily conserved mechanism, in which lack of nuclear localization of TEAD4 impairs the TE-specific transcriptional program in inner blastomeres, thereby allowing their maturation toward the ICM lineage. Restoration of TEAD4 nuclear localization maintains the TE-specific transcriptional program in the inner blastomeres and prevents segregation of the TE and ICM lineages and blastocyst formation. We propose that altered subcellular localization of TEAD4 in blastomeres dictates first mammalian cell fate specification.Proceedings of the National Academy of Sciences 04/2012; 109(19):7362-7. · 9.81 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Choroidal neovascularization (CNV) is one of the most common causes of severe vision loss. Decorin, a multiple receptor tyrosine kinase inhibitor, has been recently shown to play an important regulatory role in angiogenic response. This study aims to investigate whether the overexpression of decorin in retinal pigment epithelial (RPE) cells under hypoxia alters the in vitro angiogenic ability of cocultured choroid-retinal endothelial cells and to explore the possible mechanisms involved. Human RPE cells (ARPE-19) were subjected to hypoxia with or without decorin pretreatment, and RNA interference technique was used to knock down the Met gene in ARPE-19 cells. Cell viability was determined using the Cell Counting Kit-8 assay. Expression of Met, Rac1 and hypoxia-inducible factor-1 alpha (HIF-1α) was evaluated by western blot and quantitative real-time polymerase chain reaction (qRT-PCR). Vascular endothelial growth factor (VEGF) expression was evaluated by enzyme-linked immunosorbent assay (ELISA) and qRT-PCR. We then constructed a recombinant lentiviral vector carrying the decorin gene to transduce ARPE-19 cells. The overexpression of decorin in transduced RPE cells was confirmed by qRT-PCR and western blot. The transduced RPE cells were then cocultured with rhesus macaque choroid-retinal endothelial cells (RF/6A) in a transwell coculture system to observe the effects of decorin overexpression in ARPE-19 cells on the proliferation, migration and tube formation of RF/6A cells. In response to hypoxia, the VEGF concentrations in the culture supernatants increased greatly at 24 and 48 h, and this effect was inhibited significantly and nearly equally in the presence of 50-200 nM decorin. Decorin pretreatment before hypoxia exposure effectively reduced the hypoxia-induced expression of Met, Rac1, HIF-1α and VEGF in ARPE-19 cells. Transfection of small interfering RNA against Met to ARPE-19 cells also resulted in significant downregulation of Rac1, HIF-1α and VEGF under hypoxia, and this effect was similar to that noted with decorin pretreatment alone or with their combination. Results from the coculture system showed that the overexpression of decorin in ARPE-19 cells significantly inhibited the proliferation, migration and tube formation of RF/6A cells. These results indicate that Met pathway activation plays an important role in the upregulation of VEGF in RPE cells under hypoxia. Decorin may interfere with angiogenesis by downregulating hypoxia-induced Met, Rac1, HIF-1α and VEGF expression in RPE cells, which suggests a potential strategy for the inhibition of CNV.Experimental Eye Research 09/2013; · 3.03 Impact Factor