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Angiostatin K1‐3 induces E‐selectin via AP1 and Ets1: a mediator for anti‐angiogenic action of K1‐3
Author(s) -
CHEN Y.H.,
HUANG Y.H.,
WU H.L.,
WU M.P.,
CHANG W.T.,
KUO Y.Z.,
LU K.C.,
WU L.W.
Publication year - 2008
Publication title -
journal of thrombosis and haemostasis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.947
H-Index - 178
eISSN - 1538-7836
pISSN - 1538-7933
DOI - 10.1111/j.1538-7836.2008.03139.x
Subject(s) - ap 1 transcription factor , chromatin immunoprecipitation , angiostatin , microbiology and biotechnology , gene knockdown , transactivation , immunoprecipitation , blot , chemistry , biology , promoter , angiogenesis , gene expression , cancer research , biochemistry , gene
Summary. Background : Angiostatin, a circulating angiogenic inhibitor, is an internal fragment of plasminogen and consists of several isoforms, K1‐3 included. We previously showed that K1‐3 was the most potent angiostatin to induce E‐selectin mRNA expression. The purpose of this study was to identify the mechanism responsible for K1‐3‐induced E‐selectin expression and investigate the role of E‐selectin in the anti‐angiogenic action of K1‐3. Methods and results : Quantitative real time RT‐PCR and Western blotting analyses confirmed a time‐dependent increase of E‐selectin mRNA and protein induced by K1‐3. Subcellular fractionation and immunofluorescence microscopy showed the co‐localization of K1‐3‐induced E‐selectin with caveolin 1 (Cav1) in lipid rafts in which E‐selectin may behave as a signaling receptor. Promoter‐driven reporter assays and site‐directed mutagenesis showed that K1‐3 induced E‐selectin expression via promoter activation and AP1 and Ets‐1 binding sites in the proximal E‐selectin promoter were required for E‐selectin induction. The in vivo binding of both protein complexes to the proximal promoter was confirmed by chromatin immunoprecipitation (ChIP). Although K1‐3 induced the activation of ERK1/2 and JNK, only repression of JNK activation attenuated the induction of E‐selectin by K1‐3. A modulatory role of E‐selectin in the anti‐angiogenic action of K1‐3 was manifested by both overexpression and knockdown of E‐selectin followed by cell proliferation assay. Conclusions : We show that K1‐3 induced E‐selectin expression via AP1 and Ets‐1 binding to the proximal E‐selectin promoter (−356/+1), which was positively mediated by JNK activation. Our findings also demonstrate E‐selectin as a novel target for the anti‐angiogenic therapy.