Vascular injury induces posttranscriptional regulation of the Id3 gene - Cloning of a novel Id3 isoform expressed during vascular lesion formation inrat and human atherosclerosis

The molecular mechanisms that regulate the proliferation of smooth muscle c ells (SMCs) of the vasculature in response to injury are poorly understood. Members of the inhibitor of DNA binding (Id) class of helix-loop-helix tra nscription factors are known to regulate the growth of a variety of cell ty pes; however, the expression of the various Id genes in SMCs and in vascula r lesions has not been examined. In the present study, the yeast 2-hybrid s ystem was used to clone Id genes from a cultured rat aortic SMC library. By use of ubiquitous E proteins as bait, Id3 and a novel isoform of 1d3 (Id3a ) were cloned. Id3a is the product of alternative splicing of the Id3 gene, resulting in inclusion of a 115-bp "coding intron," which encodes a unique 29-amino acid carboxyl terminus for the Id3a protein. Unlike Id3, Id3a mRN A was not detected in the normal rat carotid artery. However, after balloon injury, Id3a was abundantly expressed throughout the neointimal layer. In addition, mRNA of the human homologue of Id3a (Id3L) was detected in human carotid atherosclerotic plaques. Adenovirus-mediated overexpression of thes e Id3 isoforms in cultured rat aortic SMCs revealed that infection of SMCs with an adenovirus overexpressing Id3a (in contrast to Id3) resulted in a s ignificant decrease in cell number versus AdLacZ-infected cells. DNA fragme ntation analysis suggested that this decrease in SMC viability was due to i ncreased apoptotic activity in cells infected with adenovirus overexpressin g Id3a. These results provide evidence that alternative splicing of the Id3 gene may represent an important mechanism by which neointimal SMC growth i s attenuated during vascular lesion formation.