title: Mammals have two twinfilin isoforms whose subcellular localizations and tissue distributions are differentially regulated.
abstract: Twinfilin is a highly conserved actin monomer-binding protein that regulates cytoskeletal dynamics in organisms from yeast to mammals. In addition to the previously characterized mammalian twinfilin-1, a second protein with approximately 65% sequence identity to twinfilin-1 exists in mouse and humans. However, previous studies failed to identify any actin binding activity in this protein (Rohwer, A., Kittstein, W., Marks, F., and Gschwendt, M. (1999) Eur. J. Biochem. 263, 518-525). Here we show that this protein, which we named twinfilin-2, is indeed an actin monomer-binding protein. Similar to twinfilin-1, mouse twinfilin-2 binds ADP-G-actin with a higher affinity (KD = 0.12 microM) than ATP-G-actin (KD = 1.96 microM) and efficiently inhibits actin filament assembly in vitro. Both mouse twinfilins inhibit the nucleotide exchange on actin monomers and directly interact with capping protein. Furthermore, the actin interactions of mouse twinfilin-1 and twinfilin-2 are inhibited by phosphatidylinositol (4,5)-bisphosphate. Although biochemically very similar, our Northern blots and in situ hybridizations show that these two proteins display distinct expression patterns. Twinfilin-1 is the major isoform in embryos and in most adult mouse non-muscle cell-types, whereas twinfilin-2 is the predominant isoform of adult heart and skeletal muscles. Studies with isoform-specific antibodies demonstrated that although the two proteins show similar localizations in unstimulated cells, they are regulated by different mechanisms. The small GTPases Rac1 and Cdc42 induce the redistribution of twinfilin-1 to membrane ruffles and cell-cell contacts, respectively, but do not affect the localization of twinfilin-2. Taken together, these data show that mammals have two twinfilin isoforms, which are differentially expressed and regulated through distinct cellular signaling pathways.
authors.affiliation: Program in Cellular Biotechnology, Institute of Biotechnology, University of Helsinki, 00014 Helsinki, Finland.
authors.forename: Maria K
authors.initials: MK
authors.lastname: Vartiainen
authors.forename: Elisa M
authors.initials: EM
authors.lastname: Sarkkinen
authors.forename: Tanja
authors.initials: T
authors.lastname: Matilainen
authors.forename: Marjo
authors.initials: M
authors.lastname: Salminen
authors.forename: Pekka
authors.initials: P
authors.lastname: Lappalainen
mesh_terms: D016475:3T3 Cells; D000199:Actins / Q000737:chemistry / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D002352:Carrier Proteins; D002450:Cell Communication; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005786:Gene Expression Regulation; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D017403:In Situ Hybridization; D007700:Kinetics; D051379:Mice; D008840:Microfilament Proteins / Q000737:chemistry*; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases; D012313:RNA / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D015398:Signal Transduction; D013997:Time Factors; D014018:Tissue Distribution; D014407:Tumor Cells, Cultured; D020764:cdc42 GTP-Binding Protein / Q000737:chemistry; D020830:rac1 GTP-Binding Protein / Q000378:metabolism
chemical_list: D000199:Actins; D002352:Carrier Proteins; D018076:DNA, Complementary; D005982:Glutathione Transferase; D008840:Microfilament Proteins; D019269:Phosphatidylinositol 4,5-Diphosphate; D020033:Protein Isoforms; D011505:Protein-Tyrosine Kinases; D012313:RNA; D029701:Saccharomyces cerevisiae Proteins; D020764:cdc42 GTP-Binding Protein; D020830:rac1 GTP-Binding Protein; C485190:Twf1 protein, mouse; C113980:TWF1 protein, S cerevisiae; C485188:TWF1 protein, human; C120744:TWF2 protein, human; C485645:TWF2 protein, mouse