RESUMO
The 2 related basic helix loop helix genes, LYL1 and TAL-1 are active in hematopoietic and endothelial lineages. While Tal-1 is essential for both hematopoietic and vascular development, the role of Lyl1 appears to be distinct as deficient mice are viable and display modest hematopoietic defects. Here, we reveal a role for Lyl1 as a major regulator of adult neovascularization. Tumors implanted into Lyl1-deficient mice showed higher proliferation and angiogenesis, as evidenced by enlarged lumens, reduced pericyte coverage and increased permeability, compared with wild type littermates. Of note, Lyl1-deficient tumor vessels exhibited an up-regulation of Tal-1, the VE-Cadherin target gene, as well as Angiopoietin-2, 3 major actors in angiogenesis. Hematopoietic reconstitution experiments demonstrated that this sustained tumor angiogenesis was of endothelial origin. Moreover, the angiogenic phenotype observed in the absence of Lyl1 function was not tumor-restricted as microvessels forming in Matrigel or originating from aortic explants were also more numerous and larger than their wild-type counterparts. Finally, LYL1 depletion in human endothelial cells revealed that LYL1 controls the expression of molecules involved in the stabilization of vascular structures. Together, our data show a role for LYL1 in the postnatal maturation of newly formed blood vessels.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Proteínas de Neoplasias/metabolismo , Neoplasias Experimentais/irrigação sanguínea , Neoplasias Experimentais/metabolismo , Neovascularização Patológica/metabolismo , Angiopoietina-2/biossíntese , Angiopoietina-2/genética , Animais , Antígenos CD/biossíntese , Antígenos CD/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Caderinas/biossíntese , Caderinas/genética , Regulação Neoplásica da Expressão Gênica/genética , Hematopoese/genética , Humanos , Camundongos , Camundongos Mutantes , Proteínas de Neoplasias/genética , Transplante de Neoplasias , Neoplasias Experimentais/genética , Neovascularização Patológica/genética , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteína 1 de Leucemia Linfocítica Aguda de Células T , Regulação para Cima/genéticaRESUMO
The basic helix-loop-helix TAL-1/SCL essential for hematopoietic development is also required during vascular development for embryonic angiogenesis. We reported that TAL-1 acts positively on postnatal angiogenesis by stimulating endothelial morphogenesis. Here, we investigated the functional consequences of TAL-1 silencing in human primary endothelial cells. We found that TAL-1 knockdown caused the inhibition of in vitro tubulomorphogenesis, which was associated with a dramatic reduction in vascular endothelial cadherin (VE-cadherin) at intercellular junctions. Consistently, silencing of TAL-1 as well as of its cofactors E47 and LMO2 down-regulated VE-cadherin at both the mRNA and the protein level. Endogenous VE-cadherin transcription could be activated in nonendothelial HEK-293 cells by the sole concomitant ectopic expression of TAL-1, E47, and LMO2. Transient transfections in human primary endothelial cells derived from umbilical vein (HUVECs) demonstrated that VE-cadherin promoter activity was dependent on the integrity of a specialized E-box associated with a GATA motif and was maximal with the coexpression of the different components of the TAL-1 complex. Finally, chromatin immunoprecipitation assays showed that TAL-1 and its cofactors occupied the VE-cadherin promoter in HUVECs. Together, these data identify VE-cadherin as a bona fide target gene of the TAL-1 complex in the endothelial lineage, providing a first clue to TAL-1 function in angiogenesis.
Assuntos
Antígenos CD/biossíntese , Fatores de Transcrição Hélice-Alça-Hélice Básicos/fisiologia , Caderinas/biossíntese , Proteínas de Ligação a DNA/fisiologia , Células Endoteliais/metabolismo , Metaloproteínas/fisiologia , Proteínas Proto-Oncogênicas/fisiologia , Fatores de Transcrição TCF/fisiologia , Regulação para Cima , Proteínas Adaptadoras de Transdução de Sinal , Sequência de Aminoácidos , Antígenos CD/genética , Sequência de Bases , Caderinas/genética , Células Cultivadas , Colágeno , Combinação de Medicamentos , Células Endoteliais/fisiologia , Endotélio Vascular/citologia , Humanos , Proteínas com Domínio LIM , Laminina , Dados de Sequência Molecular , Neovascularização Fisiológica , Regiões Promotoras Genéticas , Ligação Proteica , Proteoglicanas , Proteína 1 de Leucemia Linfocítica Aguda de Células T , Proteína 1 Semelhante ao Fator 7 de TranscriçãoRESUMO
The two related basic helix-loop-helix, TAL1 and LYL1, and their cofactor LIM-only-2 protein (LMO2) are present in blood and endothelial cells. While their crucial role in early hematopoiesis is well established, their function in endothelial cells and especially in angiogenesis is less understood. Here, we identified ANGIOPOIETIN-2 (ANG-2), which encodes a major regulator of angiogenesis, as a direct transcriptional target of TAL1, LYL1 and LMO2. Knockdown of any of the three transcription factors in human blood and lymphatic endothelial cells caused ANG-2 mRNA and protein down-regulation. Transient transfections showed that the full activity of the ANG-2 promoter required the integrity of a highly conserved Ebox-GATA composite element. Accordingly, chromatin immunoprecipitation assays demonstrated that TAL1, LYL1, LMO2 and GATA2 occupied this region of ANG-2 promoter in human endothelial cells. Furthermore, we showed that LMO2 played a central role in assembling TAL1-E47, LYL1-LYL1 or/and LYL1-TAL1 dimers with GATA2. The resulting complexes were able to activate endogenous ANG-2 expression in endothelial cells as well as in non-endothelial cells. Finally, we showed that ANG-2 gene activation during angiogenesis concurred with the up-regulation of TAL1 and LMO2. Altogether, we identified ANG-2 as a bona fide target gene of LMO2-complexes with TAL1 and/or LYL1, highlighting a new function of the three hematopoietic factors in the endothelial lineage.