Endothelial cell activation in a VEGF-A gradient: relevance to cell fate decisions.
Distribution of vascular endothelial cell growth factor A (VEGF-A) as a gradient determines microvascular endothelial cell (EC) fate during organogenesis. While much is understood about mechanisms of differential distribution, less is known about how EC perceive and interpret a graded VEGF-A signal to generate positional target gene activation. Using microvascular EC, we analyzed the effect of time and graded VEGF-A input on VEGFR2 autophosphorylation, signal kinase activation and induction of immediate-early genes. The threshold and time to peak activation of VEGFR2 were dependent on signal strength over a 50-fold range in concentration with 3-fold concentration differences readily distinguished. Longer duration of exposure did not compensate for low concentration of VEGF-A, suggesting intensity and duration of signal were not interpreted equivalently. With the same conditions, graded and time-sensitive information was transduced through the PLCgamma/p44/p42MAPK signal pathway but not the parallel AKT pathway. Analysis of MAPK-induced angiogenic immediate-early genes determined that EGR-1, EGR-3, and NR4A1 were dependent on graded input while NR4A2 and DSCR1 were independent with 'switch-like' induction. These data demonstrate rapid, linear integration of VEGF-A levels but independent interpretation of duration of signal and identify potential nodes for segregation of gradient-dependent and -independent responses. These results describe how microvascular EC fate decisions can be determined by comparatively moderate changes in VEGF signal strength, resulting in combinatorial changes in the repertoire of immediate-early genes for transcription effectors.
Division of Neonatology and Pulmonary Biology, Cincinnati Children's Hospital Research Foundation, Department of Pediatrics, University of Cincinnati, Cincinnati, OH 45229, USA. Ann.akeson@cchmc.org
Ann
A
Akeson
Amanda
A
Herman
Diane
D
Wiginton
James
J
Greenberg
D000818:Animals; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D004268:DNA-Binding Proteins; D004305:Dose-Response Relationship, Drug; D051766:Early Growth Response Protein 1 / Q000235:genetics; D051776:Early Growth Response Protein 3 / Q000235:genetics; D004622:Embryo, Mammalian / Q000166:cytology / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D005260:Female; D015870:Gene Expression / Q000187:drug effects / Q000235:genetics; D018507:Gene Expression Regulation, Developmental / Q000502:physiology*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics; D007700:Kinetics; D008168:Lung / Q000166:cytology / Q000378:metabolism; D051379:Mice; D008815:Mice, Inbred Strains; D008822:Mice, Transgenic; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D009124:Muscle Proteins / Q000235:genetics; D057105:Nuclear Receptor Subfamily 4, Group A, Member 1 / Q000235:genetics; D057126:Nuclear Receptor Subfamily 4, Group A, Member 2 / Q000235:genetics; D051966:Phospholipase C gamma / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000494:pharmacology*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism
D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; D047908:Intracellular Signaling Peptides and Proteins; D009124:Muscle Proteins; C539821:NR4A1 protein, human; C539882:NR4A2 protein, human; D057105:Nuclear Receptor Subfamily 4, Group A, Member 1; D057126:Nuclear Receptor Subfamily 4, Group A, Member 2; C108612:RCAN1 protein, human; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; C467480:vascular endothelial growth factor A, mouse; D051776:Early Growth Response Protein 3; D014443:Tyrosine; D040301:Vascular Endothelial Growth Factor Receptor-2; D051057:Proto-Oncogene Proteins c-akt; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D051966:Phospholipase C gamma