Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1993 Jan 2;120(2):421–435. doi: 10.1083/jcb.120.2.421

Cofilin is an essential component of the yeast cortical cytoskeleton

PMCID: PMC2119511  PMID: 8421056

Abstract

We have biochemically identified the Saccharomyces cerevisiae homologue of the mammalian actin binding protein cofilin. Cofilin and related proteins isolated from diverse organisms are low molecular weight proteins (15-20 kD) that possess several activities in vitro. All bind to monomeric actin and sever filaments, and some can stably associate with filaments. In this study, we demonstrate using viscosity, sedimentation, and actin assembly rate assays that yeast cofilin (16 kD) possesses all of these properties. Cloning and sequencing of the S. cerevisiae cofilin gene (COF1) revealed that yeast cofilin is 41% identical in amino acid sequence to mammalian cofilin and, surprisingly, has homology to a protein outside the family of cofilin- like proteins. The NH2-terminal 16kD of Abp1p, a 65-kD yeast protein identified by its ability to bind to actin filaments, is 23% identical to yeast cofilin. Immunofluorescence experiments showed that, like Abp1p, cofilin is associated with the membrane actin cytoskeleton. A complete disruption of the COF1 gene was created in diploid cells. Sporulation and tetrad analysis revealed that yeast cofilin has an essential function in vivo. Although Abp1p shares sequence similarity with cofilin and has the same distribution as cofilin in the cell, multiple copies of the ABP1 gene cannot compensate for the loss of cofilin. Thus, cofilin and Abp1p are structurally related but functionally distinct components of the yeast membrane cytoskeleton.

Full Text

The Full Text of this article is available as a PDF (3.0 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Adams A. E., Botstein D., Drubin D. G. Requirement of yeast fimbrin for actin organization and morphogenesis in vivo. Nature. 1991 Dec 5;354(6352):404–408. doi: 10.1038/354404a0. [DOI] [PubMed] [Google Scholar]
  2. Adams A. E., Pringle J. R. Relationship of actin and tubulin distribution to bud growth in wild-type and morphogenetic-mutant Saccharomyces cerevisiae. J Cell Biol. 1984 Mar;98(3):934–945. doi: 10.1083/jcb.98.3.934. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Amatruda J. F., Cannon J. F., Tatchell K., Hug C., Cooper J. A. Disruption of the actin cytoskeleton in yeast capping protein mutants. Nature. 1990 Mar 22;344(6264):352–354. doi: 10.1038/344352a0. [DOI] [PubMed] [Google Scholar]
  4. Amatruda J. F., Cooper J. A. Purification, characterization, and immunofluorescence localization of Saccharomyces cerevisiae capping protein. J Cell Biol. 1992 Jun;117(5):1067–1076. doi: 10.1083/jcb.117.5.1067. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Attri A. K., Lewis M. S., Korn E. D. The formation of actin oligomers studied by analytical ultracentrifugation. J Biol Chem. 1991 Apr 15;266(11):6815–6824. [PubMed] [Google Scholar]
  6. Bamburg J. R., Harris H. E., Weeds A. G. Partial purification and characterization of an actin depolymerizing factor from brain. FEBS Lett. 1980 Nov 17;121(1):178–182. doi: 10.1016/0014-5793(80)81292-0. [DOI] [PubMed] [Google Scholar]
  7. Barnes G., Drubin D. G., Stearns T. The cytoskeleton of Saccharomyces cerevisiae. Curr Opin Cell Biol. 1990 Feb;2(1):109–115. doi: 10.1016/s0955-0674(05)80040-7. [DOI] [PubMed] [Google Scholar]
  8. Bernstein B. W., Bamburg J. R. Tropomyosin binding to F-actin protects the F-actin from disassembly by brain actin-depolymerizing factor (ADF). Cell Motil. 1982;2(1):1–8. doi: 10.1002/cm.970020102. [DOI] [PubMed] [Google Scholar]
  9. Chenevert J., Corrado K., Bender A., Pringle J., Herskowitz I. A yeast gene (BEM1) necessary for cell polarization whose product contains two SH3 domains. Nature. 1992 Mar 5;356(6364):77–79. doi: 10.1038/356077a0. [DOI] [PubMed] [Google Scholar]
  10. Cicchetti P., Mayer B. J., Thiel G., Baltimore D. Identification of a protein that binds to the SH3 region of Abl and is similar to Bcr and GAP-rho. Science. 1992 Aug 7;257(5071):803–806. doi: 10.1126/science.1379745. [DOI] [PubMed] [Google Scholar]
  11. Cooper J. A., Blum J. D., Williams R. C., Jr, Pollard T. D. Purification and characterization of actophorin, a new 15,000-dalton actin-binding protein from Acanthamoeba castellanii. J Biol Chem. 1986 Jan 5;261(1):477–485. [PubMed] [Google Scholar]
  12. Cooper J. A., Walker S. B., Pollard T. D. Pyrene actin: documentation of the validity of a sensitive assay for actin polymerization. J Muscle Res Cell Motil. 1983 Apr;4(2):253–262. doi: 10.1007/BF00712034. [DOI] [PubMed] [Google Scholar]
  13. Dingwall C., Laskey R. A. Protein import into the cell nucleus. Annu Rev Cell Biol. 1986;2:367–390. doi: 10.1146/annurev.cb.02.110186.002055. [DOI] [PubMed] [Google Scholar]
  14. Drubin D. G., Miller K. G., Botstein D. Yeast actin-binding proteins: evidence for a role in morphogenesis. J Cell Biol. 1988 Dec;107(6 Pt 2):2551–2561. doi: 10.1083/jcb.107.6.2551. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Drubin D. G., Mulholland J., Zhu Z. M., Botstein D. Homology of a yeast actin-binding protein to signal transduction proteins and myosin-I. Nature. 1990 Jan 18;343(6255):288–290. doi: 10.1038/343288a0. [DOI] [PubMed] [Google Scholar]
  16. Gehrung S., Snyder M. The SPA2 gene of Saccharomyces cerevisiae is important for pheromone-induced morphogenesis and efficient mating. J Cell Biol. 1990 Oct;111(4):1451–1464. doi: 10.1083/jcb.111.4.1451. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Gerring S. L., Connelly C., Hieter P. Positional mapping of genes by chromosome blotting and chromosome fragmentation. Methods Enzymol. 1991;194:57–77. doi: 10.1016/0076-6879(91)94007-y. [DOI] [PubMed] [Google Scholar]
  18. Gimeno C. J., Ljungdahl P. O., Styles C. A., Fink G. R. Unipolar cell divisions in the yeast S. cerevisiae lead to filamentous growth: regulation by starvation and RAS. Cell. 1992 Mar 20;68(6):1077–1090. doi: 10.1016/0092-8674(92)90079-r. [DOI] [PubMed] [Google Scholar]
  19. Haarer B. K., Lillie S. H., Adams A. E., Magdolen V., Bandlow W., Brown S. S. Purification of profilin from Saccharomyces cerevisiae and analysis of profilin-deficient cells. J Cell Biol. 1990 Jan;110(1):105–114. doi: 10.1083/jcb.110.1.105. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hartwig J. H., Kwiatkowski D. J. Actin-binding proteins. Curr Opin Cell Biol. 1991 Feb;3(1):87–97. doi: 10.1016/0955-0674(91)90170-4. [DOI] [PubMed] [Google Scholar]
  21. Iida K., Matsumoto S., Yahara I. The KKRKK sequence is involved in heat shock-induced nuclear translocation of the 18-kDa actin-binding protein, cofilin. Cell Struct Funct. 1992 Feb;17(1):39–46. doi: 10.1247/csf.17.39. [DOI] [PubMed] [Google Scholar]
  22. Iida K., Yahara I. Reversible induction of actin rods in mouse C3H-2K cells by incubation in salt buffers and by treatment with non-ionic detergents. Exp Cell Res. 1986 Jun;164(2):492–506. doi: 10.1016/0014-4827(86)90047-9. [DOI] [PubMed] [Google Scholar]
  23. Isenberg G., Aebi U., Pollard T. D. An actin-binding protein from Acanthamoeba regulates actin filament polymerization and interactions. Nature. 1980 Dec 4;288(5790):455–459. doi: 10.1038/288455a0. [DOI] [PubMed] [Google Scholar]
  24. Ito H., Fukuda Y., Murata K., Kimura A. Transformation of intact yeast cells treated with alkali cations. J Bacteriol. 1983 Jan;153(1):163–168. doi: 10.1128/jb.153.1.163-168.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Jackson C. L., Hartwell L. H. Courtship in S. cerevisiae: both cell types choose mating partners by responding to the strongest pheromone signal. Cell. 1990 Nov 30;63(5):1039–1051. doi: 10.1016/0092-8674(90)90507-b. [DOI] [PubMed] [Google Scholar]
  26. Janmey P. A., Stossel T. P. Modulation of gelsolin function by phosphatidylinositol 4,5-bisphosphate. Nature. 1987 Jan 22;325(6102):362–364. doi: 10.1038/325362a0. [DOI] [PubMed] [Google Scholar]
  27. Johnston G. C., Prendergast J. A., Singer R. A. The Saccharomyces cerevisiae MYO2 gene encodes an essential myosin for vectorial transport of vesicles. J Cell Biol. 1991 May;113(3):539–551. doi: 10.1083/jcb.113.3.539. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Koch C. A., Anderson D., Moran M. F., Ellis C., Pawson T. SH2 and SH3 domains: elements that control interactions of cytoplasmic signaling proteins. Science. 1991 May 3;252(5006):668–674. doi: 10.1126/science.1708916. [DOI] [PubMed] [Google Scholar]
  29. Koffer A., Edgar A. J., Bamburg J. R. Identification of two species of actin depolymerizing factor in cultures of BHK cells. J Muscle Res Cell Motil. 1988 Aug;9(4):320–328. doi: 10.1007/BF01773875. [DOI] [PubMed] [Google Scholar]
  30. Kouyama T., Mihashi K. Fluorimetry study of N-(1-pyrenyl)iodoacetamide-labelled F-actin. Local structural change of actin protomer both on polymerization and on binding of heavy meromyosin. Eur J Biochem. 1981;114(1):33–38. [PubMed] [Google Scholar]
  31. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  32. Lassing I., Lindberg U. Specific interaction between phosphatidylinositol 4,5-bisphosphate and profilactin. Nature. 1985 Apr 4;314(6010):472–474. doi: 10.1038/314472a0. [DOI] [PubMed] [Google Scholar]
  33. Liu H. P., Bretscher A. Disruption of the single tropomyosin gene in yeast results in the disappearance of actin cables from the cytoskeleton. Cell. 1989 Apr 21;57(2):233–242. doi: 10.1016/0092-8674(89)90961-6. [DOI] [PubMed] [Google Scholar]
  34. Liu H. P., Bretscher A. Purification of tropomyosin from Saccharomyces cerevisiae and identification of related proteins in Schizosaccharomyces and Physarum. Proc Natl Acad Sci U S A. 1989 Jan;86(1):90–93. doi: 10.1073/pnas.86.1.90. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Mabuchi I. An actin-depolymerizing protein (depactin) from starfish oocytes: properties and interaction with actin. J Cell Biol. 1983 Nov;97(5 Pt 1):1612–1621. doi: 10.1083/jcb.97.5.1612. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. MacLean-Fletcher S. D., Pollard T. D. Viscometric analysis of the gelation of Acanthamoeba extracts and purification of two gelation factors. J Cell Biol. 1980 May;85(2):414–428. doi: 10.1083/jcb.85.2.414. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Maciver S. K., Zot H. G., Pollard T. D. Characterization of actin filament severing by actophorin from Acanthamoeba castellanii. J Cell Biol. 1991 Dec;115(6):1611–1620. doi: 10.1083/jcb.115.6.1611. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Matsuzaki F., Matsumoto S., Yahara I., Yonezawa N., Nishida E., Sakai H. Cloning and characterization of porcine brain cofilin cDNA. Cofilin contains the nuclear transport signal sequence. J Biol Chem. 1988 Aug 15;263(23):11564–11568. [PubMed] [Google Scholar]
  39. Moriyama K., Matsumoto S., Nishida E., Sakai H., Yahara I. Nucleotide sequence of mouse cofilin cDNA. Nucleic Acids Res. 1990 May 25;18(10):3053–3053. doi: 10.1093/nar/18.10.3053. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Moriyama K., Nishida E., Yonezawa N., Sakai H., Matsumoto S., Iida K., Yahara I. Destrin, a mammalian actin-depolymerizing protein, is closely related to cofilin. Cloning and expression of porcine brain destrin cDNA. J Biol Chem. 1990 Apr 5;265(10):5768–5773. [PubMed] [Google Scholar]
  41. Moriyama K., Yonezawa N., Sakai H., Yahara I., Nishida E. Mutational analysis of an actin-binding site of cofilin and characterization of chimeric proteins between cofilin and destrin. J Biol Chem. 1992 Apr 15;267(11):7240–7244. [PubMed] [Google Scholar]
  42. Mortimer R. K., Schild D., Contopoulou C. R., Kans J. A. Genetic map of Saccharomyces cerevisiae, edition 10. Yeast. 1989 Sep-Oct;5(5):321–403. doi: 10.1002/yea.320050503. [DOI] [PubMed] [Google Scholar]
  43. Nishida E., Iida K., Yonezawa N., Koyasu S., Yahara I., Sakai H. Cofilin is a component of intranuclear and cytoplasmic actin rods induced in cultured cells. Proc Natl Acad Sci U S A. 1987 Aug;84(15):5262–5266. doi: 10.1073/pnas.84.15.5262. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Nishida E., Maekawa S., Muneyuki E., Sakai H. Action of a 19K protein from porcine brain on actin polymerization: a new functional class of actin-binding proteins. J Biochem. 1984 Feb;95(2):387–398. doi: 10.1093/oxfordjournals.jbchem.a134619. [DOI] [PubMed] [Google Scholar]
  45. Nishida E., Maekawa S., Sakai H. Cofilin, a protein in porcine brain that binds to actin filaments and inhibits their interactions with myosin and tropomyosin. Biochemistry. 1984 Oct 23;23(22):5307–5313. doi: 10.1021/bi00317a032. [DOI] [PubMed] [Google Scholar]
  46. Nishida E., Muneyuki E., Maekawa S., Ohta Y., Sakai H. An actin-depolymerizing protein (destrin) from porcine kidney. Its action on F-actin containing or lacking tropomyosin. Biochemistry. 1985 Nov 5;24(23):6624–6630. doi: 10.1021/bi00344a049. [DOI] [PubMed] [Google Scholar]
  47. Novick P., Botstein D. Phenotypic analysis of temperature-sensitive yeast actin mutants. Cell. 1985 Feb;40(2):405–416. doi: 10.1016/0092-8674(85)90154-0. [DOI] [PubMed] [Google Scholar]
  48. Ohta Y., Nishida E., Sakai H., Miyamoto E. Dephosphorylation of cofilin accompanies heat shock-induced nuclear accumulation of cofilin. J Biol Chem. 1989 Sep 25;264(27):16143–16148. [PubMed] [Google Scholar]
  49. Olson M. V., Dutchik J. E., Graham M. Y., Brodeur G. M., Helms C., Frank M., MacCollin M., Scheinman R., Frank T. Random-clone strategy for genomic restriction mapping in yeast. Proc Natl Acad Sci U S A. 1986 Oct;83(20):7826–7830. doi: 10.1073/pnas.83.20.7826. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Pardee J. D., Spudich J. A. Purification of muscle actin. Methods Enzymol. 1982;85(Pt B):164–181. doi: 10.1016/0076-6879(82)85020-9. [DOI] [PubMed] [Google Scholar]
  51. Pfeffer S. R., Drubin D. G., Kelly R. B. Identification of three coated vesicle components as alpha- and beta-tubulin linked to a phosphorylated 50,000-dalton polypeptide. J Cell Biol. 1983 Jul;97(1):40–47. doi: 10.1083/jcb.97.1.40. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Pollard T. D. A falling ball apparatus to measure filament cross-linking. Methods Cell Biol. 1982;24:301–311. doi: 10.1016/s0091-679x(08)60663-9. [DOI] [PubMed] [Google Scholar]
  53. Pollard T. D., Cooper J. A. Actin and actin-binding proteins. A critical evaluation of mechanisms and functions. Annu Rev Biochem. 1986;55:987–1035. doi: 10.1146/annurev.bi.55.070186.005011. [DOI] [PubMed] [Google Scholar]
  54. Pringle J. R., Adams A. E., Drubin D. G., Haarer B. K. Immunofluorescence methods for yeast. Methods Enzymol. 1991;194:565–602. doi: 10.1016/0076-6879(91)94043-c. [DOI] [PubMed] [Google Scholar]
  55. Rose M. D., Novick P., Thomas J. H., Botstein D., Fink G. R. A Saccharomyces cerevisiae genomic plasmid bank based on a centromere-containing shuttle vector. Gene. 1987;60(2-3):237–243. doi: 10.1016/0378-1119(87)90232-0. [DOI] [PubMed] [Google Scholar]
  56. Schiestl R. H., Gietz R. D. High efficiency transformation of intact yeast cells using single stranded nucleic acids as a carrier. Curr Genet. 1989 Dec;16(5-6):339–346. doi: 10.1007/BF00340712. [DOI] [PubMed] [Google Scholar]
  57. Sikorski R. S., Hieter P. A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. Genetics. 1989 May;122(1):19–27. doi: 10.1093/genetics/122.1.19. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Snyder M. The SPA2 protein of yeast localizes to sites of cell growth. J Cell Biol. 1989 Apr;108(4):1419–1429. doi: 10.1083/jcb.108.4.1419. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Stossel T. P., Chaponnier C., Ezzell R. M., Hartwig J. H., Janmey P. A., Kwiatkowski D. J., Lind S. E., Smith D. B., Southwick F. S., Yin H. L. Nonmuscle actin-binding proteins. Annu Rev Cell Biol. 1985;1:353–402. doi: 10.1146/annurev.cb.01.110185.002033. [DOI] [PubMed] [Google Scholar]
  60. Sutoh K., Mabuchi I. End-label fingerprintings show that an N-terminal segment of depactin participates in interaction with actin. Biochemistry. 1989 Jan 10;28(1):102–106. doi: 10.1021/bi00427a015. [DOI] [PubMed] [Google Scholar]
  61. Takagi T., Konishi K., Mabuchi I. Amino acid sequence of starfish oocyte depactin. J Biol Chem. 1988 Mar 5;263(7):3097–3102. [PubMed] [Google Scholar]
  62. Vandekerckhove J. Actin-binding proteins. Curr Opin Cell Biol. 1990 Feb;2(1):41–50. doi: 10.1016/s0955-0674(05)80029-8. [DOI] [PubMed] [Google Scholar]
  63. Vandekerckhove J., Vancompernolle K. Structural relationships of actin-binding proteins. Curr Opin Cell Biol. 1992 Feb;4(1):36–42. doi: 10.1016/0955-0674(92)90056-i. [DOI] [PubMed] [Google Scholar]
  64. Woolford J. L., Jr Nuclear pre-mRNA splicing in yeast. Yeast. 1989 Nov-Dec;5(6):439–457. doi: 10.1002/yea.320050604. [DOI] [PubMed] [Google Scholar]
  65. Yonezawa N., Homma Y., Yahara I., Sakai H., Nishida E. A short sequence responsible for both phosphoinositide binding and actin binding activities of cofilin. J Biol Chem. 1991 Sep 15;266(26):17218–17221. [PubMed] [Google Scholar]
  66. Yonezawa N., Nishida E., Iida K., Kumagai H., Yahara I., Sakai H. Inhibition of actin polymerization by a synthetic dodecapeptide patterned on the sequence around the actin-binding site of cofilin. J Biol Chem. 1991 Jun 5;266(16):10485–10489. [PubMed] [Google Scholar]
  67. Yonezawa N., Nishida E., Iida K., Yahara I., Sakai H. Inhibition of the interactions of cofilin, destrin, and deoxyribonuclease I with actin by phosphoinositides. J Biol Chem. 1990 May 25;265(15):8382–8386. [PubMed] [Google Scholar]
  68. Yonezawa N., Nishida E., Koyasu S., Maekawa S., Ohta Y., Yahara I., Sakai H. Distribution among tissues and intracellular localization of cofilin, a 21kDa actin-binding protein. Cell Struct Funct. 1987 Oct;12(5):443–452. doi: 10.1247/csf.12.443. [DOI] [PubMed] [Google Scholar]
  69. Yonezawa N., Nishida E., Ohba M., Seki M., Kumagai H., Sakai H. An actin-interacting heptapeptide in the cofilin sequence. Eur J Biochem. 1989 Jul 15;183(1):235–238. doi: 10.1111/j.1432-1033.1989.tb14918.x. [DOI] [PubMed] [Google Scholar]
  70. Yonezawa N., Nishida E., Sakai H. pH control of actin polymerization by cofilin. J Biol Chem. 1985 Nov 25;260(27):14410–14412. [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES