Skip to main content
PMC home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1992 Jun 1;117(5):1023–1029. doi: 10.1083/jcb.117.5.1023

Effects of profilin and profilactin on actin structure and function in living cells

PMCID: PMC2289477  PMID: 1577865

Abstract

Previous studies have yielded conflicting results concerning the physiological role of profilin, a 12-15-kD actin- and phosphoinositide- binding protein, as a regulator of actin polymerization. We have addressed this question by directly microinjecting mammalian profilins, prepared either from an E. coli expression system or from bovine brain, into living normal rat kidney (NRK) cells. The microinjection causes a dose-dependent decrease in F-actin content, as indicated by staining with fluorescent phalloidin, and a dramatic reduction of actin and alpha-actinin along stress fibers. In addition, it has a strong inhibitory effect toward the extension of lamellipodia. However, the injection of profilin causes no detectable perturbation to the cell- substrate focal contact and no apparent depolymerization of filaments in either the nonlamellipodial circumferential band or the contractile ring of dividing cells. Furthermore, cytokinesis of injected cells occurs normally as in control cells. In contrast to pure profilin, high- affinity profilin-actin complexes from brain induce an increase in total cellular F-actin content and an enhanced ruffling activity, suggesting that the complex may dissociate readily in the cell and that there may be multiple states of profilin that differ in their ability to bind or release actin molecules. Our results indicate that profilin and profilactin can function as effective regulators for at least a subset of actin filaments in living cells.

Full Text

The Full Text of this article is available as a PDF (2.1 MB).

Selected References

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

  1. Babcock G., Rubenstein P. A. Synthesis of mammalian profilin in Escherichia coli and its characterization. Cell Motil Cytoskeleton. 1989;14(2):230–236. doi: 10.1002/cm.970140209. [DOI] [PubMed] [Google Scholar]
  2. Cao L. G., Wang Y. L. Mechanism of the formation of contractile ring in dividing cultured animal cells. I. Recruitment of preexisting actin filaments into the cleavage furrow. J Cell Biol. 1990 Apr;110(4):1089–1095. doi: 10.1083/jcb.110.4.1089. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cooper J. A. The role of actin polymerization in cell motility. Annu Rev Physiol. 1991;53:585–605. doi: 10.1146/annurev.ph.53.030191.003101. [DOI] [PubMed] [Google Scholar]
  4. DiNubile M. J., Southwick F. S. Effects of macrophage profilin on actin in the presence and absence of acumentin and gelsolin. J Biol Chem. 1985 Jun 25;260(12):7402–7409. [PubMed] [Google Scholar]
  5. Fishkind D. J., Cao L. G., Wang Y. L. Microinjection of the catalytic fragment of myosin light chain kinase into dividing cells: effects on mitosis and cytokinesis. J Cell Biol. 1991 Sep;114(5):967–975. doi: 10.1083/jcb.114.5.967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Goldschmidt-Clermont P. J., Machesky L. M., Baldassare J. J., Pollard T. D. The actin-binding protein profilin binds to PIP2 and inhibits its hydrolysis by phospholipase C. Science. 1990 Mar 30;247(4950):1575–1578. doi: 10.1126/science.2157283. [DOI] [PubMed] [Google Scholar]
  7. Goldschmidt-Clermont P. J., Machesky L. M., Doberstein S. K., Pollard T. D. Mechanism of the interaction of human platelet profilin with actin. J Cell Biol. 1991 Jun;113(5):1081–1089. doi: 10.1083/jcb.113.5.1081. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Haarer B. K., Brown S. S. Structure and function of profilin. Cell Motil Cytoskeleton. 1990;17(2):71–74. doi: 10.1002/cm.970170202. [DOI] [PubMed] [Google Scholar]
  9. 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]
  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. Kaiser D. A., Goldschmidt-Clermont P. J., Levine B. A., Pollard T. D. Characterization of renatured profilin purified by urea elution from poly-L-proline agarose columns. Cell Motil Cytoskeleton. 1989;14(2):251–262. doi: 10.1002/cm.970140211. [DOI] [PubMed] [Google Scholar]
  12. Kreis T. E., Geiger B., Schlessinger J. Mobility of microinjected rhodamine actin within living chicken gizzard cells determined by fluorescence photobleaching recovery. Cell. 1982 Jul;29(3):835–845. doi: 10.1016/0092-8674(82)90445-7. [DOI] [PubMed] [Google Scholar]
  13. 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]
  14. Lal A. A., Korn E. D. Reinvestigation of the inhibition of actin polymerization by profilin. J Biol Chem. 1985 Aug 25;260(18):10132–10138. [PubMed] [Google Scholar]
  15. 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]
  16. Lazarides E. Two general classes of cytoplasmic actin filaments in tissue culture cells: the role of tropomyosin. J Supramol Struct. 1976;5(4):531(383)–563(415). doi: 10.1002/jss.400050410. [DOI] [PubMed] [Google Scholar]
  17. Lind S. E., Janmey P. A., Chaponnier C., Herbert T. J., Stossel T. P. Reversible binding of actin to gelsolin and profilin in human platelet extracts. J Cell Biol. 1987 Aug;105(2):833–842. doi: 10.1083/jcb.105.2.833. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Machesky L. M., Goldschmidt-Clermont P. J., Pollard T. D. The affinities of human platelet and Acanthamoeba profilin isoforms for polyphosphoinositides account for their relative abilities to inhibit phospholipase C. Cell Regul. 1990 Nov;1(12):937–950. doi: 10.1091/mbc.1.12.937. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Matsudaira P. T., Burgess D. R. SDS microslab linear gradient polyacrylamide gel electrophoresis. Anal Biochem. 1978 Jul 1;87(2):386–396. doi: 10.1016/0003-2697(78)90688-7. [DOI] [PubMed] [Google Scholar]
  20. McKenna N. M., Meigs J. B., Wang Y. L. Exchangeability of alpha-actinin in living cardiac fibroblasts and muscle cells. J Cell Biol. 1985 Dec;101(6):2223–2232. doi: 10.1083/jcb.101.6.2223. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. 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]
  22. Rozycki M., Schutt C. E., Lindberg U. Affinity chromatography-based purification of profilin:actin. Methods Enzymol. 1991;196:100–118. doi: 10.1016/0076-6879(91)96012-g. [DOI] [PubMed] [Google Scholar]
  23. Safer D., Elzinga M., Nachmias V. T. Thymosin beta 4 and Fx, an actin-sequestering peptide, are indistinguishable. J Biol Chem. 1991 Mar 5;266(7):4029–4032. [PubMed] [Google Scholar]
  24. Safer D., Golla R., Nachmias V. T. Isolation of a 5-kilodalton actin-sequestering peptide from human blood platelets. Proc Natl Acad Sci U S A. 1990 Apr;87(7):2536–2540. doi: 10.1073/pnas.87.7.2536. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Sanders M. C., Wang Y. L. Exogenous nucleation sites fail to induce detectable polymerization of actin in living cells. J Cell Biol. 1990 Feb;110(2):359–365. doi: 10.1083/jcb.110.2.359. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Sanger J. M., Dabiri G., Mittal B., Kowalski M. A., Haddad J. G., Sanger J. W. Disruption of microfilament organization in living nonmuscle cells by microinjection of plasma vitamin D-binding protein or DNase I. Proc Natl Acad Sci U S A. 1990 Jul;87(14):5474–5478. doi: 10.1073/pnas.87.14.5474. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Schliwa M., van Blerkom J. Structural interaction of cytoskeletal components. J Cell Biol. 1981 Jul;90(1):222–235. doi: 10.1083/jcb.90.1.222. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Small J. V. Organization of actin in the leading edge of cultured cells: influence of osmium tetroxide and dehydration on the ultrastructure of actin meshworks. J Cell Biol. 1981 Dec;91(3 Pt 1):695–705. doi: 10.1083/jcb.91.3.695. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Southwick F. S., Young C. L. The actin released from profilin--actin complexes is insufficient to account for the increase in F-actin in chemoattractant-stimulated polymorphonuclear leukocytes. J Cell Biol. 1990 Jun;110(6):1965–1973. doi: 10.1083/jcb.110.6.1965. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Spudich J. A., Watt S. The regulation of rabbit skeletal muscle contraction. I. Biochemical studies of the interaction of the tropomyosin-troponin complex with actin and the proteolytic fragments of myosin. J Biol Chem. 1971 Aug 10;246(15):4866–4871. [PubMed] [Google Scholar]
  31. Tilney L. G., Bonder E. M., Coluccio L. M., Mooseker M. S. Actin from Thyone sperm assembles on only one end of an actin filament: a behavior regulated by profilin. J Cell Biol. 1983 Jul;97(1):112–124. doi: 10.1083/jcb.97.1.112. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Vandekerckhove J. S., Kaiser D. A., Pollard T. D. Acanthamoeba actin and profilin can be cross-linked between glutamic acid 364 of actin and lysine 115 of profilin. J Cell Biol. 1989 Aug;109(2):619–626. doi: 10.1083/jcb.109.2.619. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Vojtek A., Haarer B., Field J., Gerst J., Pollard T. D., Brown S., Wigler M. Evidence for a functional link between profilin and CAP in the yeast S. cerevisiae. Cell. 1991 Aug 9;66(3):497–505. doi: 10.1016/0092-8674(81)90013-1. [DOI] [PubMed] [Google Scholar]

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

RESOURCES