We have previously shown that maximal expression of the @entity35 @entity50022 ( @entity50022 ) gene in cultured @entity35 aortic smooth muscle cells (SMCs) required the presence of a highly conserved domain ( @entity2036 -1321 and -1095) that contained two positive-acting @entity28898 ( @entity28898 ) binding elements (CArG boxes 1 and 2) and a negative-acting GC-rich element that was recognized by @entity10564 (Madsen, C. S., Hershey, J. C., Hautmann, M. B., White, S. L., and Owens, G. K. (1997) J. Biol. Chem. 272, 6332-6340). In this study, to better understand the functional role of these three cis elements, we created a series of @entity50022 reporter-gene constructs in which each element was mutated either alone or in combination with each other and tested them for activity in transient transfection assays using primary cultured @entity35 aortic SMCs. Results demonstrated that the most proximal @entity28898 binding element (CArG-box1) was active in the absence of CArG-box2, but only upon removal of the GC-rich repressor. In contrast, regardless of sequence context, CArG-box2 was active only when CArG-box1 was present. We further demonstrated using electrophoretic mobility shift assays that @entity10564 binding to the GC-rich repressor element did not prevent @entity28898 binding to the adjacent CArG-box2. Thus, unlike other proteins reported to inhibit @entity28898 activity, the repressor activity associated with the GC-rich element does not appear to function through direct inhibition of @entity28898 binding. As a first step toward understanding the importance of these elements in vivo, we performed in vivo footprinting on the intact @entity35 aorta. We demonstrated that both CArG boxes and the GC-rich element were bound by protein within the animal. Additionally, using the @entity35 carotid injury model we showed that @entity10564 protein was significantly increased in SMCs located within the @entity75 , suggesting that increased expression of this putative repressor factor may contribute to the decreased SM MHC expression within SMCs found in myointimal lesions.

Interaction of CArG elements and a GC-rich repressor element in transcriptional regulation of the XXXX gene in vascular smooth muscle cells.

@entity28898 :: ('501099(Tax:10116)', 'Gene') :: ['serum response factor', 'SRF'] @entity2036 :: ('MESH:D009711', 'Chemical') :: ['nucleotides'] @entity75 :: ('MESH:D001927', 'Disease') :: ['myointimal lesion'] @entity50022 :: ('24582(Tax:10116)', 'Gene') :: ['SM-MHC', 'smooth muscle myosin heavy chain'] @entity35 :: ('10116', 'Species') :: ['rat'] @entity10564 :: ('24790(Tax:10116)', 'Gene') :: ['Sp1']
@entity50022 :: (24582(Tax:10116),Gene) :: ['smooth muscle myosin heavy chain']
