title: Tropomyosin variants describe distinct functional subcellular domains in differentiated vascular smooth muscle cells.
abstract: Tropomyosin (Tm) is known to be an important gatekeeper of actin function. Tm isoforms are encoded by four genes, and each gene produces several variants by alternative splicing, which have been proposed to play roles in motility, proliferation, and apoptosis. Smooth muscle studies have focused on gizzard smooth muscle, where a heterodimer of Tm from the α-gene (Tmsm-α) and from the β-gene (Tmsm-β) is associated with contractile filaments. In this study we examined Tm in differentiated mammalian vascular smooth muscle (dVSM). Liquid chromatography-tandem mass spectrometry (LC MS/MS) analysis and Western blot screening with variant-specific antibodies revealed that at least five different Tm proteins are expressed in this tissue: Tm6 (Tmsm-α) and Tm2 from the α-gene, Tm1 (Tmsm-β) from the β-gene, Tm5NM1 from the γ-gene, and Tm4 from the δ-gene. Tm6 is by far most abundant in dVSM followed by Tm1, Tm2, Tm5NM1, and Tm4. Coimmunoprecipitation and coimmunofluorescence studies demonstrate that Tm1 and Tm6 coassociate with different actin isoforms and display different intracellular localizations. Using an antibody specific for cytoplasmic γ-actin, we report here the presence of a γ-actin cortical cytoskeleton in dVSM cells. Tm1 colocalizes with cortical cytoplasmic γ-actin and coprecipitates with γ-actin. Tm6, on the other hand, is located on contractile bundles. These data indicate that Tm1 and Tm6 do not form a classical heterodimer in dVSM but rather describe different functional cellular compartments.
authors.affiliation: Health Sciences Dept., Boston University, 635 Commonwealth Ave., Boston, MA 02215, USA.
authors.forename: Cynthia
authors.initials: C
authors.lastname: Gallant
authors.forename: Sarah
authors.initials: S
authors.lastname: Appel
authors.forename: Philip
authors.initials: P
authors.lastname: Graceffa
authors.forename: Paul
authors.initials: P
authors.lastname: Leavis
authors.forename: Jim Jung-Ching
authors.initials: JJ
authors.lastname: Lin
authors.forename: Peter W
authors.initials: PW
authors.lastname: Gunning
authors.forename: Galina
authors.initials: G
authors.lastname: Schevzov
authors.forename: Christine
authors.initials: C
authors.lastname: Chaponnier
authors.forename: Jon
authors.initials: J
authors.lastname: DeGnore
authors.forename: William
authors.initials: W
authors.lastname: Lehman
authors.forename: Kathleen G
authors.initials: KG
authors.lastname: Morgan
mesh_terms: D000199:Actins / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002454:Cell Differentiation / Q000502:physiology*; D002645:Chickens; D005289:Ferrets; D006801:Humans; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000166:cytology*; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000502:physiology*; D011485:Protein Binding; D020033:Protein Isoforms / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D014335:Tropomyosin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D000199:Actins; D020033:Protein Isoforms; D014335:Tropomyosin