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RhoA/ROCK‐dependent moesin phosphorylation regulates AGEe‐induced endothelial response
Author(s) -
Huang Qiaobing,
Wang Jiping,
Liu Hongxia,
Chen Bo,
Li Qiang,
Huang Xuliang
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.855.7
Subject(s) - rhoa , moesin , phosphorylation , microbiology and biotechnology , rho associated protein kinase , chemistry , actin cytoskeleton , signal transduction , cytoskeleton , biology , cell , biochemistry , ezrin
To investigate the involvement of RhoA/ROCK‐dependent moesin phosphorylation in endothelia stimulated by advanced glycation end products (AGEs), human dermal microvascular endothelial cells (HMVECs) were used to observe the monolayer permeability and distribution of F‐actin. Activations of RhoA and ROCK were determined by a luminescence‐based assay and immunoblotting. RhoA N19 or RhoA L63 was transfected to down‐regulate RhoA expression or over‐express RhoA in HMVECs. H‐1152 was employed to block activation of ROCK. Co‐immunoprecipitation was used to confirm the interaction of ROCK and moesin. To identify AGE/ROCK‐induced phosphorylation site in moesin, mutant pcDNA3/HA‐moesinT558A or pcDNA3/HA‐moesinT558D were applied in endothelia. The results showed that AGE‐HSA increased the monolayer permeability and triggered the formation of F‐actin stress fibers. AGE‐HSA enhanced RhoA/ROCK in a time‐ and dose‐dependent manner. RhoA N19 or H‐1152 abolished these AGE‐induced changes, while RhoA L63 reproduced the AGE‐evoked responses. It is confirmed that AGE induced direct interaction of ROCK and moesin. Thr558 was identified as the phosphorylating site for moesin in AGE‐evoked endothelia. These results indicate the effects of RhoA/ROCK pathway and subsequent moesin phosphorylation in AGE‐mediated endothelial dysfunction. (supported by NSFC30771028, 30971201; IRT0730 and G2005CB522601)