Identification and characterization of novel smoothelin isoforms in vascular smooth muscle

Smoothelin is a cytoskeletal protein specifically expressed in differentiat ed smooth muscle cells and has been shown to colocalize with smooth muscle alpha actin, In addition to the small smoothelin isoform of 59 kD, we recen tly identified a large smoothelin isoform of 117 kD, The aim of this study was to identify and characterize novel smoothelin isoforms, The genomic str ucture and sequence of the smoothelin gene were determined by genomic PCR, RT-PCR and DNA sequencing. Comparison of the cDNA and genomic sequences sho ws that the small smoothelin isoform is generated by transcription initiati on 10 kb downstream of the start site of the large isoform. In addition to the known smoothelin cDNA(cl isoform) we identified two novel cDNA variants (c2 and c3 isoform) that are generated by alternative splicing within a re gion, which shows similarity to the spectrin family of F-actin cross-linkin g proteins. Visceral organs express the c1 form, while the c2 form prevails in well-vascularized tissue as analyzed by RT-PCR. We then generated speci fic antibodies against the major smoothelin isoforms and could show by West ern blotting and immunohistochemistry that the large isoform is specificall y expressed. in vascular smooth muscle cells, while the small isoform is ab undant in visceral smooth muscle. These results strongly suggest that the s moothelin gene contains a vascular and a visceral smooth muscle promoter. T he cell-type-specific expression of smoothelin isoforms that are associated with actin filaments may play a role in the modulation of the contractile properties of different smooth muscle cell types. Copyright (C) 2001 S. Kar ger AG, Basel.