Homeobox protein Hex facilitates serum responsive factor-mediated activation of the SM22alpha gene transcription in embryonic fibroblasts.
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.


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 alpha-actin and SM22alpha but not of calponin and smooth muscle myosin heavy chain. Transient transfection assays showed that Hex activates the transcription from the SM22alpha 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 SM22alpha gene by facilitating the interaction between SRF and its cognate binding site in pluripotent embryonic fibroblasts. This study demonstrates that Hex, a hematopoietically expressed homeobox protein, induces transcription of the SM22alpha gene by facilitating the interaction between SRF and its cognate binding site in embryonic fibroblasts. These findings will provide the clue for understanding the mechanisms by which bone marrow-derived SMC precursor cells undergo differentiation.
Oyama
Yuko
Y
Department of Medicine and Biological Science, University of Gunma, Japan.
Kawai-Kowase
Keiko
K
Sekiguchi
Kenichi
K
Sato
Mahito
M
Sato
Hiroko
H
Yamazaki
Miki
M
Ohyama
Yoshio
Y
Aihara
Yasushi
Y
Iso
Tatsuya
T
Okamaoto
Eichi
E
Nagai
Ryozo
R
Kurabayashi
Masahiko
M
D000199:Actins / Q000096:biosynthesis* / Q000235:genetics; D000818:Animals; D001665:Binding Sites; D002478:Cells, Cultured / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D005347:Fibroblasts / Q000378:metabolism; D005786:Gene Expression Regulation / Q000502:physiology*; D017930:Genes, Reporter; D018398:Homeodomain Proteins / Q000502:physiology*; D051379:Mice; D008840:Microfilament Proteins / Q000096:biosynthesis* / Q000235:genetics; D009124:Muscle Proteins / Q000096:biosynthesis* / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D025941:Protein Interaction Mapping; D011993:Recombinant Fusion Proteins / Q000502:physiology; D027602:Serum Response Element / Q000235:genetics*; D026362:Serum Response Factor / Q000378:metabolism / Q000494:pharmacology*; D014157:Transcription Factors
D000199:Actins; C513080:Hhex protein, mouse; D018398:Homeodomain Proteins; D008840:Microfilament Proteins; D009124:Muscle Proteins; D011993:Recombinant Fusion Proteins; D026362:Serum Response Factor; D014157:Transcription Factors; C051615:transgelin