Opposite functions of HIF-α isoforms in VEGF induction by TGF-β1 under non-hypoxic conditions.
Transforming growth factor (TGF)-β1 has biphasic functions in prostate tumorigenesis, having a growth-inhibitory effect in the early stages, but in the late stages promoting tumor angiogenesis and metastasis. We demonstrate here that tumor-producing TGF-β1 induces vascular endothelial growth factor (VEGF) in prostate cancer cells, and hypoxia-inducible factor (HIF)-1α and HIF-2α has opposite functions in TGF-β1 regulation of VEGF expression under non-hypoxic conditions. The promoter response of VEGF to TGF-β1 was upregulated by the transfection of HIF-2α or siHIF-1α but downregulated by HIF-1α and siHIF-2α. Both HIF-1α and HIF-2α were induced by TGF-β1 at mRNA and protein levels, however, their nuclear translocation was differentially regulated by TGF-β1, suggesting its association with their opposite effects. VEGF induction by TGF-β1 occurred in a Smad3-dependent manner, and the Smad-binding element 2 (SBE2, -992 to -986) and hypoxia response element (-975 to -968) in the VEGF promoter were required for the promoter response to TGF-β1. Smad3 cooperated with HIF-2α in TGF-β1 activation of VEGF transcription and Smad3 binding to the SBE2 site was greatly impaired by knockdown of HIF-2α expression. Moreover, the VEGF promoter response to TGF-β1 was synergistically elevated by co-transfection of Smad3 and HIF-2α but attenuated by HIF-1α in a dose-dependent manner. Additionally, TGF-β1 was found to increase the stability of VEGF transcript by facilitating the cytoplasmic translocation of a RNA-stabilizing factor HuR. Collectively, our data show that tumor-producing TGF-β1 induces VEGF at the both transcription and post-transcriptional levels through multiple routes including Smad3, HIF-2α and HuR. This study thus suggests that autocrine TGF-β1 production may contribute to tumor angiogenesis via HIF-2α signaling under non-hypoxic conditions, providing a selective growth advantage for prostate tumor cells.
School of Life Sciences and Biotechnology, Korea University, Seoul, Republic of Korea.
K S
KS
Chae
M J
MJ
Kang
J H
JH
Lee
B K
BK
Ryu
M G
MG
Lee
N G
NG
Her
T K
TK
Ha
J
J
Han
Y K
YK
Kim
S G
SG
Chi
D000954:Antigens, Surface / Q000235:genetics / Q000378:metabolism; D051792:Basic Helix-Loop-Helix Proteins / Q000096:biosynthesis* / Q000235:genetics; D015152:Blotting, Northern; D015153:Blotting, Western; D015687:Cell Hypoxia / Q000235:genetics; D045744:Cell Line, Tumor; D047369:Chromatin Immunoprecipitation; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; D024202:Electrophoretic Mobility Shift Assay; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics*; D055785:Gene Knockdown Techniques; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000096:biosynthesis* / Q000235:genetics; D008297:Male; D011401:Promoter Regions, Genetic / Q000235:genetics; D011471:Prostatic Neoplasms / Q000235:genetics / Q000378:metabolism*; D020033:Protein Isoforms / Q000096:biosynthesis / Q000235:genetics; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000235:genetics; D051900:Smad3 Protein / Q000235:genetics / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000235:genetics / Q000378:metabolism*; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism*; D000099162:Endothelial PAS Domain-Containing Protein 1
D000954:Antigens, Surface; D051792:Basic Helix-Loop-Helix Proteins; D051959:ELAV Proteins; D000067780:ELAV-Like Protein 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D020033:Protein Isoforms; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D051900:Smad3 Protein; D053773:Transforming Growth Factor beta1; D042461:Vascular Endothelial Growth Factor A; D000099162:Endothelial PAS Domain-Containing Protein 1; C098770:ELAVL1 protein, human; C497442:HIF1A protein, human; C497790:SMAD3 protein, human