Skip to main content
PMC home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1992 Nov 2;119(4):835–842. doi: 10.1083/jcb.119.4.835

Evidence for functional homology in the F-actin binding domains of gelsolin and alpha-actinin: implications for the requirements of severing and capping

PMCID: PMC2289707  PMID: 1331120

Abstract

The F-actin binding domains of gelsolin and alpha-actinin compete for the same site on actin filaments with similar binding affinities. Both contain tandem repeats of approximately 125 amino acids, the first of which is shown to contain the actin-binding site. We have replaced the F-actin binding domain in the NH2-terminal half of gelsolin by that of alpha-actinin. The hybrid severs filaments almost as efficiently as does gelsolin or its NH2-terminal half, but unlike the latter, requires calcium ions. The hybrid binds two actin monomers and caps the barbed ends of filaments in the presence or absence of calcium. The cap produced by the hybrid binds with lower affinity than that of gelsolin and is not stable: It dissociates from filament ends with a half life of approximately 15 min. Although there is no extended sequence homology between these two different F-actin binding domains, our experiments show that they are functionally equivalent and provide new insights into the mechanism of microfilament severing.

Full Text

The Full Text of this article is available as a PDF (1.0 MB).

Selected References

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

  1. Blanchard A., Ohanian V., Critchley D. The structure and function of alpha-actinin. J Muscle Res Cell Motil. 1989 Aug;10(4):280–289. doi: 10.1007/BF01758424. [DOI] [PubMed] [Google Scholar]
  2. Bresnick A. R., Janmey P. A., Condeelis J. Evidence that a 27-residue sequence is the actin-binding site of ABP-120. J Biol Chem. 1991 Jul 15;266(20):12989–12993. [PubMed] [Google Scholar]
  3. Bresnick A. R., Warren V., Condeelis J. Identification of a short sequence essential for actin binding by Dictyostelium ABP-120. J Biol Chem. 1990 Jun 5;265(16):9236–9240. [PubMed] [Google Scholar]
  4. Bryan J. Gelsolin has three actin-binding sites. J Cell Biol. 1988 May;106(5):1553–1562. doi: 10.1083/jcb.106.5.1553. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Byers T. J., Husain-Chishti A., Dubreuil R. R., Branton D., Goldstein L. S. Sequence similarity of the amino-terminal domain of Drosophila beta spectrin to alpha actinin and dystrophin. J Cell Biol. 1989 Oct;109(4 Pt 1):1633–1641. doi: 10.1083/jcb.109.4.1633. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Eichinger L., Noegel A. A., Schleicher M. Domain structure in actin-binding proteins: expression and functional characterization of truncated severin. J Cell Biol. 1991 Feb;112(4):665–676. doi: 10.1083/jcb.112.4.665. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Eichinger L., Schleicher M. Characterization of actin- and lipid-binding domains in severin, a Ca(2+)-dependent F-actin fragmenting protein. Biochemistry. 1992 May 26;31(20):4779–4787. doi: 10.1021/bi00135a006. [DOI] [PubMed] [Google Scholar]
  8. Gill S. C., von Hippel P. H. Calculation of protein extinction coefficients from amino acid sequence data. Anal Biochem. 1989 Nov 1;182(2):319–326. doi: 10.1016/0003-2697(89)90602-7. [DOI] [PubMed] [Google Scholar]
  9. Gorlin J. B., Yamin R., Egan S., Stewart M., Stossel T. P., Kwiatkowski D. J., Hartwig J. H. Human endothelial actin-binding protein (ABP-280, nonmuscle filamin): a molecular leaf spring. J Cell Biol. 1990 Sep;111(3):1089–1105. doi: 10.1083/jcb.111.3.1089. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. 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]
  11. Hemmings L., Kuhlman P. A., Critchley D. R. Analysis of the actin-binding domain of alpha-actinin by mutagenesis and demonstration that dystrophin contains a functionally homologous domain. J Cell Biol. 1992 Mar;116(6):1369–1380. doi: 10.1083/jcb.116.6.1369. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Karinch A. M., Zimmer W. E., Goodman S. R. The identification and sequence of the actin-binding domain of human red blood cell beta-spectrin. J Biol Chem. 1990 Jul 15;265(20):11833–11840. [PubMed] [Google Scholar]
  13. Koenig M., Monaco A. P., Kunkel L. M. The complete sequence of dystrophin predicts a rod-shaped cytoskeletal protein. Cell. 1988 Apr 22;53(2):219–228. doi: 10.1016/0092-8674(88)90383-2. [DOI] [PubMed] [Google Scholar]
  14. Kwiatkowski D. J., Janmey P. A., Yin H. L. Identification of critical functional and regulatory domains in gelsolin. J Cell Biol. 1989 May;108(5):1717–1726. doi: 10.1083/jcb.108.5.1717. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kwiatkowski D. J., Stossel T. P., Orkin S. H., Mole J. E., Colten H. R., Yin H. L. Plasma and cytoplasmic gelsolins are encoded by a single gene and contain a duplicated actin-binding domain. Nature. 1986 Oct 2;323(6087):455–458. doi: 10.1038/323455a0. [DOI] [PubMed] [Google Scholar]
  16. Matsudaira P., Janmey P. Pieces in the actin-severing protein puzzle. Cell. 1988 Jul 15;54(2):139–140. doi: 10.1016/0092-8674(88)90542-9. [DOI] [PubMed] [Google Scholar]
  17. Meyer R. K., Aebi U. Bundling of actin filaments by alpha-actinin depends on its molecular length. J Cell Biol. 1990 Jun;110(6):2013–2024. doi: 10.1083/jcb.110.6.2013. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Murphy D. B., Gray R. O., Grasser W. A., Pollard T. D. Direct demonstration of actin filament annealing in vitro. J Cell Biol. 1988 Jun;106(6):1947–1954. doi: 10.1083/jcb.106.6.1947. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Noegel A. A., Rapp S., Lottspeich F., Schleicher M., Stewart M. The Dictyostelium gelation factor shares a putative actin binding site with alpha-actinins and dystrophin and also has a rod domain containing six 100-residue motifs that appear to have a cross-beta conformation. J Cell Biol. 1989 Aug;109(2):607–618. doi: 10.1083/jcb.109.2.607. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Pollard T. D. Rate constants for the reactions of ATP- and ADP-actin with the ends of actin filaments. J Cell Biol. 1986 Dec;103(6 Pt 2):2747–2754. doi: 10.1083/jcb.103.6.2747. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Pope B., Way M., Weeds A. G. Two of the three actin-binding domains of gelsolin bind to the same subdomain of actin. Implications of capping and severing mechanisms. FEBS Lett. 1991 Mar 11;280(1):70–74. doi: 10.1016/0014-5793(91)80206-i. [DOI] [PubMed] [Google Scholar]
  22. Selve N., Wegner A. Rate constants and equilibrium constants for binding of the gelsolin-actin complex to the barbed ends of actin filaments in the presence and absence of calcium. Eur J Biochem. 1986 Oct 15;160(2):379–387. doi: 10.1111/j.1432-1033.1986.tb09982.x. [DOI] [PubMed] [Google Scholar]
  23. Way M., Gooch J., Pope B., Weeds A. G. Expression of human plasma gelsolin in Escherichia coli and dissection of actin binding sites by segmental deletion mutagenesis. J Cell Biol. 1989 Aug;109(2):593–605. doi: 10.1083/jcb.109.2.593. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Way M., Pope B., Cross R. A., Kendrick-Jones J., Weeds A. G. Expression of the N-terminal domain of dystrophin in E. coli and demonstration of binding to F-actin. FEBS Lett. 1992 Apr 27;301(3):243–245. doi: 10.1016/0014-5793(92)80249-g. [DOI] [PubMed] [Google Scholar]
  25. Way M., Pope B., Gooch J., Hawkins M., Weeds A. G. Identification of a region in segment 1 of gelsolin critical for actin binding. EMBO J. 1990 Dec;9(12):4103–4109. doi: 10.1002/j.1460-2075.1990.tb07632.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Way M., Pope B., Weeds A. G. Are the conserved sequences in segment 1 of gelsolin important for binding actin? J Cell Biol. 1992 Mar;116(5):1135–1143. doi: 10.1083/jcb.116.5.1135. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Way M., Pope B., Weeds A. Molecular biology of actin binding proteins: evidence for a common structural domain in the F-actin binding sites of gelsolin and alpha-actinin. J Cell Sci Suppl. 1991;14:91–94. doi: 10.1242/jcs.1991.supplement_14.19. [DOI] [PubMed] [Google Scholar]
  28. Way M., Weeds A. Nucleotide sequence of pig plasma gelsolin. Comparison of protein sequence with human gelsolin and other actin-severing proteins shows strong homologies and evidence for large internal repeats. J Mol Biol. 1988 Oct 20;203(4):1127–1133. doi: 10.1016/0022-2836(88)90132-5. [DOI] [PubMed] [Google Scholar]
  29. Weber A., Pring M., Lin S. L., Bryan J. Role of the N- and C-terminal actin-binding domains of gelsolin in barbed filament end capping. Biochemistry. 1991 Sep 24;30(38):9327–9334. doi: 10.1021/bi00102a027. [DOI] [PubMed] [Google Scholar]
  30. Weeds A. G., Gooch J., Hawkins M., Pope B., Way M. Role of actin-binding proteins in cytoskeletal dynamics. Biochem Soc Trans. 1991 Nov;19(4):1016–1020. doi: 10.1042/bst0191016. [DOI] [PubMed] [Google Scholar]
  31. Yin H. L. Gelsolin: calcium- and polyphosphoinositide-regulated actin-modulating protein. Bioessays. 1987 Oct;7(4):176–179. doi: 10.1002/bies.950070409. [DOI] [PubMed] [Google Scholar]
  32. Yin H. L., Iida K., Janmey P. A. Identification of a polyphosphoinositide-modulated domain in gelsolin which binds to the sides of actin filaments. J Cell Biol. 1988 Mar;106(3):805–812. doi: 10.1083/jcb.106.3.805. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Yu F. X., Zhou D. M., Yin H. L. Chimeric and truncated gCap39 elucidate the requirements for actin filament severing and end capping by the gelsolin family of proteins. J Biol Chem. 1991 Oct 15;266(29):19269–19275. [PubMed] [Google Scholar]
  34. de Arruda M. V., Watson S., Lin C. S., Leavitt J., Matsudaira P. Fimbrin is a homologue of the cytoplasmic phosphoprotein plastin and has domains homologous with calmodulin and actin gelation proteins. J Cell Biol. 1990 Sep;111(3):1069–1079. doi: 10.1083/jcb.111.3.1069. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES