Abstract
Actin-binding proteins filamin A (FLNA) and B (FLNB) are expressed in endothelial cells and play an essential role during vascular development. In order to investigate their role in adult endothelial cell function, we initially confirmed their expression pattern in different adult mouse tissues and cultured cell lines and found that FLNB expression is concentrated mainly in endothelial cells, whereas FLNA is more ubiquitously expressed. Functionally, small interfering RNA knockdown of endogenous FLNB in human umbilical vein endothelial cells inhibited vascular endothelial growth factor (VEGF)-induced in vitro angiogenesis by decreasing endothelial cell migration capacity, whereas FLNA ablation did not alter these parameters. Moreover, FLNB-depleted cells increased their substrate adhesion with more focal adhesions. The molecular mechanism underlying this effect implicates modulation of small GTP-binding protein Rac-1 localization and activity, with altered activation of its downstream effectors p21 protein Cdc42/Rac-activated kinase (PAK)-4/5/6 and its activating guanine nucleotide exchange factor Vav-2. Moreover, our results suggest the existence of a signaling complex, including FLNB, Rac-1, and Vav-2, under basal conditions that would further interact with VEGFR2 and integrin alphavbeta5 after VEGF stimulation. In conclusion, our results reveal a crucial role for FLNB in endothelial cell migration and in the angiogenic process in adult endothelial cells.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Blotting, Northern
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Blotting, Western
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Cell Cycle
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Cell Movement*
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Cells, Cultured
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Chromatin Immunoprecipitation
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Contractile Proteins / antagonists & inhibitors
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Contractile Proteins / genetics
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Contractile Proteins / metabolism*
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Endothelium, Vascular / cytology
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Endothelium, Vascular / metabolism
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Enzyme-Linked Immunosorbent Assay
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Filamins
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Fluorescent Antibody Technique
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Humans
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Immunoenzyme Techniques
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Immunoprecipitation
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Lung / cytology
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Lung / metabolism
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Mice
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Microfilament Proteins / antagonists & inhibitors
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Microfilament Proteins / genetics
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Microfilament Proteins / metabolism*
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Neovascularization, Physiologic
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Protein Binding
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Proto-Oncogene Proteins c-vav / genetics
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Proto-Oncogene Proteins c-vav / metabolism*
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RNA, Messenger / genetics
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RNA, Messenger / metabolism
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RNA, Small Interfering / pharmacology
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Receptors, Vitronectin / genetics
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Receptors, Vitronectin / metabolism
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Reverse Transcriptase Polymerase Chain Reaction
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Umbilical Veins / cytology
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Umbilical Veins / metabolism
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Vascular Endothelial Growth Factor A / genetics
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Vascular Endothelial Growth Factor A / metabolism*
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Vascular Endothelial Growth Factor Receptor-2 / genetics
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Vascular Endothelial Growth Factor Receptor-2 / metabolism
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rac1 GTP-Binding Protein / genetics
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rac1 GTP-Binding Protein / metabolism*
Substances
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Contractile Proteins
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FLNB protein, human
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Filamins
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Microfilament Proteins
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Proto-Oncogene Proteins c-vav
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RAC1 protein, human
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RNA, Messenger
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RNA, Small Interfering
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Receptors, Vitronectin
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VAV2 protein, human
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Vascular Endothelial Growth Factor A
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integrin alphaVbeta5
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Vascular Endothelial Growth Factor Receptor-2
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rac1 GTP-Binding Protein