on July 8, 2004
Published online before print July 8, 2004, doi: 10.1161/01.ATV.0000138404.17519.45
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Submitted on May 10, 2004
Accepted on June 21, 2004
Homeobox Protein Hex Facilitates Serum Responsive Factor-Mediated Activation of the SM22
Gene Transcription in Embryonic Fibroblasts
Yuko Oyama ;
From Department of Medicine and Biological Science (Y.O., K.K.-K., K.S., M.S., H.S., M.Y., Y.O., Y.A., T.I., E.O., M.K.), Graduate School of Medicine, University of Gunma, Gunma; and the Department of Cardiovascular Medicine (R.N.), Graduate School of Medicine, University of Tokyo, Tokyo, Japan.
* To whom correspondence should be addressed. E-mail: mkuraba{at}med.gunma-u.ac.jp.
Objective--Hex (hematopoietically expressed homeobox), a member of homeobox family of transcription factors, has been implicated in the vascular development because of its expression in hemangioblast, a hypothetical stem cell that gives rise to both angioblasts and hematopoietic lineages. In the present study, we examined the role of Hex in the differentiation of vascular smooth muscle cells.
Methods and Results--We constructed adenovirus expressing Hex, to which we refer to as AxCA/Hex, and transduced murine embryonic fibroblasts, 10T1/2 cells. Northern blot analyses showed that Hex increased the mRNA levels of smooth muscle 
-actin and SM22 but not of calponin and smooth muscle myosin heavy chain. Transient transfection assays showed that Hex activates the transcription from the SM22 promoter in a CArG box-dependent manner. Electrophoretic mobility shift assays demonstrate that Hex is not able to bind to CArG box, but binding of serum responsive factor (SRF) to CArG box is enhanced in AxCA/Hex-transduced cells. Recombinant Hex protein produced by in vitro translation system augmented the binding activity of SRF to CArG box. Immunoprecipitation experiments revealed the physical association between Hex and SRF.
Conclusions--Hex induces transcription of the SM22 gene by facilitating the interaction between SRF and its cognate binding site in pluripotent embryonic fibroblasts.
Key words: transcriptional factor • smooth muscle cells • atherosclerosis • homeobox • serum response factor
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