title: A LIM domain protein from tobacco involved in actin-bundling and histone gene transcription.
abstract: The two LIM domain-containing proteins from plants (LIMs) typically exhibit a dual cytoplasmic-nuclear distribution, suggesting that, in addition to their previously described roles in actin cytoskeleton organization, they participate in nuclear processes. Using a south-western blot-based screen aimed at identifying factors that bind to plant histone gene promoters, we isolated a positive clone containing the tobacco LIM protein WLIM2 (NtWLIM2) cDNA. Using both green fluorescent protein (GFP) fusion- and immunology-based strategies, we provide clear evidence that NtWLIM2 localizes to the actin cytoskeleton, the nucleus, and the nucleolus. Interestingly, the disruption of the actin cytoskeleton by latrunculin B significantly increases NtWLIM2 nuclear fraction, pinpointing a possible novel cytoskeletal-nuclear crosstalk. Biochemical and electron microscopy experiments reveal the ability of NtWLIM2 to directly bind to actin filaments and to crosslink the latter into thick actin bundles. Electrophoretic mobility shift assays show that NtWLIM2 specifically binds to the conserved octameric cis-elements (Oct) of the Arabidopsis histone H4A748 gene promoter and that this binding largely relies on both LIM domains. Importantly, reporter-based experiments conducted in Arabidopsis and tobacco protoplasts confirm the ability of NtWLIM2 to bind to and activate the H4A748 gene promoter in live cells. Expression studies indicate the constitutive presence of NtWLIM2 mRNA and NtWLIM2 protein during tobacco BY-2 cell proliferation and cell cycle progression, suggesting a role of NtWLIM2 in the activation of basal histone gene expression. Interestingly, both live cell and in vitro data support NtWLIM2 di/oligomerization. We propose that NtWLIM2 functions as an actin-stabilizing protein, which, upon cytoskeleton remodeling, shuttles to the nucleus in order to modify gene expression.
authors.affiliation: Centre de Recherche Public-Santé, 84, Val Fleuri, L-1526 Luxembourg, Luxembourg. daniele.moes@crpsante.lu
authors.forename: Danièle
authors.initials: D
authors.lastname: Moes
authors.forename: Sabrina
authors.initials: S
authors.lastname: Gatti
authors.forename: Céline
authors.initials: C
authors.lastname: Hoffmann
authors.forename: Monika
authors.initials: M
authors.lastname: Dieterle
authors.forename: Flora
authors.initials: F
authors.lastname: Moreau
authors.forename: Katrin
authors.initials: K
authors.lastname: Neumann
authors.forename: Marc
authors.initials: M
authors.lastname: Schumacher
authors.forename: Marc
authors.initials: M
authors.lastname: Diederich
authors.forename: Erwin
authors.initials: E
authors.lastname: Grill
authors.forename: Wen-Hui
authors.initials: WH
authors.lastname: Shen
authors.forename: André
authors.initials: A
authors.lastname: Steinmetz
authors.forename: Clément
authors.initials: C
authors.lastname: Thomas
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D000199:Actins / Q000378:metabolism*; D021581:Active Transport, Cell Nucleus; D002467:Cell Nucleus / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D017930:Genes, Reporter / Q000235:genetics; D006657:Histones / Q000235:genetics*; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D011523:Protoplasts / Q000378:metabolism; D014026:Nicotiana / Q000166:cytology / Q000235:genetics* / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D000199:Actins; D018076:DNA, Complementary; D006657:Histones; D010940:Plant Proteins