Dimethylarginine dimethylaminohydrolase 2 increases vascular endothelial growth factor expression through Sp1 transcription factor in endothelial cells.
OBJECTIVE
Dimethylarginie dimethylaminohydrolase (DDAH) is a degrading enzyme for asymmetrical dimethylarginine, an endogenous NO synthase inhibitor. The molecular mechanism for DDAH-induced vascular endothelial growth factor (VEGF) expression was examined.


RESULTS
Although the transfection of expression vectors for 2 isoforms of DDAH, DDAH1, or DDAH2 increased DDAH activity in bovine aortic endothelial cells and human umbilical vein endothelial cells, expression and secretion of VEGF were increased only in DDAH2-transfected cells. Knocking down the DDAH2 gene reduced VEGF production, and DDAH2 overexpression enhanced both proliferation and migration of endothelial cells. The VEGF promoter activity was increased by DDAH2 transfection, which was not blocked by an NO synthase (NOS) inhibitor but required the Sp1 sites. DDAH2 overexpression increased nuclear protein levels bound to Sp1 oligonucleotides in endothelial cells. Sp1 small interfering RNA blocked DDAH2-induced upregulation of VEGF. DDAH2 transfection increased nuclear and threonine-phosphorylation levels of Sp1 in a protein kinase A (PKA)-dependent manner. Protein-protein interaction between DDAH2 and PKA was enhanced in DDAH2-transfected cells.


CONCLUSIONS
DDAH2 upregulated the expression of VEGF through Sp1-dependent and NO/NOS system-independent promoter activation. DDAH2-increased Sp1 DNA binding activity was PKA dependent. These mechanisms may provide a novel therapeutic strategy for VEGF-related vasculopathies such as atherosclerosis.
Department of Internal Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, Japan.
Kazuhiro
K
Hasegawa
Shu
S
Wakino
Toru
T
Tanaka
Masumi
M
Kimoto
Satoru
S
Tatematsu
Takeshi
T
Kanda
Kyoko
K
Yoshioka
Koichiro
K
Homma
Naoki
N
Sugano
Masahiko
M
Kurabayashi
Takao
T
Saruta
Koichi
K
Hayashi
D000581:Amidohydrolases / Q000235:genetics / Q000494:pharmacology / Q000502:physiology*; D000818:Animals; D002417:Cattle; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000378:metabolism* / Q000502:physiology; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000494:pharmacology / Q000502:physiology; D051379:Mice; D010766:Phosphorylation / Q000187:drug effects; D011401:Promoter Regions, Genetic / Q000502:physiology; D016329:Sp1 Transcription Factor / Q000378:metabolism / Q000502:physiology*; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*
D007527:Isoenzymes; D016329:Sp1 Transcription Factor; D042461:Vascular Endothelial Growth Factor A; D000581:Amidohydrolases; C059999:dimethylargininase