Skip to main content
PMC home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1992 Jun 2;117(6):1241–1249. doi: 10.1083/jcb.117.6.1241

Localization and specificity of the phospholipid and actin binding sites on the tail of Acanthamoeba myosin IC

PMCID: PMC2289509  PMID: 1607386

Abstract

We used bacterially expressed beta-galactosidase fusion proteins to localize the phospholipid binding domain of Acanthamoeba myosin IC to the region between amino acids 701 and 888 in the NH2-terminal half of the tail. Using a novel immobilized ligand lipid binding assay, we determined that myosin I can bind to several different acidic phospholipids, and that binding requires a minimum of 5 mol% acidic phospholipid in a neutral lipid background. The presence of di- and triglycerides and sterols in the lipid bilayer do not contribute to the affinity of myosin I for membranes. We confirm that the ATP-insensitive actin binding site is contained in the COOH-terminal 30 kD of the tail as previously shown for Acanthamoeba myosin IA. We conclude that the association of the myosin IC tail with acidic phospholipid head groups supplies much of the energy for binding myosin I to biological membranes, but probably not specificity for targeting myosin I isoforms to different cellular locations.

Full Text

The Full Text of this article is available as a PDF (1.5 MB).

Selected References

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

  1. Adams R. J., Pollard T. D. Binding of myosin I to membrane lipids. Nature. 1989 Aug 17;340(6234):565–568. doi: 10.1038/340565a0. [DOI] [PubMed] [Google Scholar]
  2. Baines I. C., Korn E. D. Localization of myosin IC and myosin II in Acanthamoeba castellanii by indirect immunofluorescence and immunogold electron microscopy. J Cell Biol. 1990 Nov;111(5 Pt 1):1895–1904. doi: 10.1083/jcb.111.5.1895. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Brzeska H., Lynch T. J., Korn E. D. Localization of the actin-binding sites of Acanthamoeba myosin IB and effect of limited proteolysis on its actin-activated Mg2+-ATPase activity. J Biol Chem. 1988 Jan 5;263(1):427–435. [PubMed] [Google Scholar]
  4. Chalmers J. J., Kim E., Telford J. N., Wong E. Y., Tacon W. C., Shuler M. L., Wilson D. B. Effects of temperature on Escherichia coli overproducing beta-lactamase or human epidermal growth factor. Appl Environ Microbiol. 1990 Jan;56(1):104–111. doi: 10.1128/aem.56.1.104-111.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Clark R. A., Volpp B. D., Leidal K. G., Nauseef W. M. Two cytosolic components of the human neutrophil respiratory burst oxidase translocate to the plasma membrane during cell activation. J Clin Invest. 1990 Mar;85(3):714–721. doi: 10.1172/JCI114496. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Côté G. P., Albanesi J. P., Ueno T., Hammer J. A., 3rd, Korn E. D. Purification from Dictyostelium discoideum of a low-molecular-weight myosin that resembles myosin I from Acanthamoeba castellanii. J Biol Chem. 1985 Apr 25;260(8):4543–4546. [PubMed] [Google Scholar]
  7. Drenckhahn D., Dermietzel R. Organization of the actin filament cytoskeleton in the intestinal brush border: a quantitative and qualitative immunoelectron microscope study. J Cell Biol. 1988 Sep;107(3):1037–1048. doi: 10.1083/jcb.107.3.1037. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. 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]
  9. Eisenberg E., Kielley W. W. Troponin-tropomyosin complex. Column chromatographic separation and activity of the three, active troponin components with and without tropomyosin present. J Biol Chem. 1974 Aug 10;249(15):4742–4748. [PubMed] [Google Scholar]
  10. Fukui Y., Lynch T. J., Brzeska H., Korn E. D. Myosin I is located at the leading edges of locomoting Dictyostelium amoebae. Nature. 1989 Sep 28;341(6240):328–331. doi: 10.1038/341328a0. [DOI] [PubMed] [Google Scholar]
  11. Gadasi H., Korn E. D. Evidence for differential intracellular localization of the Acanthamoeba myosin isoenzymes. Nature. 1980 Jul 31;286(5772):452–456. doi: 10.1038/286452a0. [DOI] [PubMed] [Google Scholar]
  12. Garcia A., Coudrier E., Carboni J., Anderson J., Vandekerkhove J., Mooseker M., Louvard D., Arpin M. Partial deduced sequence of the 110-kD-calmodulin complex of the avian intestinal microvillus shows that this mechanoenzyme is a member of the myosin I family. J Cell Biol. 1989 Dec;109(6 Pt 1):2895–2903. doi: 10.1083/jcb.109.6.2895. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hagen S. J., Kiehart D. P., Kaiser D. A., Pollard T. D. Characterization of monoclonal antibodies to Acanthamoeba myosin-I that cross-react with both myosin-II and low molecular mass nuclear proteins. J Cell Biol. 1986 Dec;103(6 Pt 1):2121–2128. doi: 10.1083/jcb.103.6.2121. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hamaguchi M., Hanafusa H. Association of p60src with Triton X-100-resistant cellular structure correlates with morphological transformation. Proc Natl Acad Sci U S A. 1987 Apr;84(8):2312–2316. doi: 10.1073/pnas.84.8.2312. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hammer J. A. Novel myosins. Trends Cell Biol. 1991 Aug;1(2-3):50–56. doi: 10.1016/0962-8924(91)90089-r. [DOI] [PubMed] [Google Scholar]
  16. Hayden S. M., Wolenski J. S., Mooseker M. S. Binding of brush border myosin I to phospholipid vesicles. J Cell Biol. 1990 Aug;111(2):443–451. doi: 10.1083/jcb.111.2.443. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hoshimaru M., Nakanishi S. Identification of a new type of mammalian myosin heavy chain by molecular cloning. Overlap of its mRNA with preprotachykinin B mRNA. J Biol Chem. 1987 Oct 25;262(30):14625–14632. [PubMed] [Google Scholar]
  18. Jung G., Korn E. D., Hammer J. A., 3rd The heavy chain of Acanthamoeba myosin IB is a fusion of myosin-like and non-myosin-like sequences. Proc Natl Acad Sci U S A. 1987 Oct;84(19):6720–6724. doi: 10.1073/pnas.84.19.6720. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Jung G., Saxe C. L., 3rd, Kimmel A. R., Hammer J. A., 3rd Dictyostelium discoideum contains a gene encoding a myosin I heavy chain. Proc Natl Acad Sci U S A. 1989 Aug;86(16):6186–6190. doi: 10.1073/pnas.86.16.6186. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Jung G., Schmidt C. J., Hammer J. A., 3rd Myosin I heavy-chain genes of Acanthamoeba castellanii: cloning of a second gene and evidence for the existence of a third isoform. Gene. 1989 Oct 30;82(2):269–280. doi: 10.1016/0378-1119(89)90052-8. [DOI] [PubMed] [Google Scholar]
  21. Kiehart D. P. Molecular genetic dissection of myosin heavy chain function. Cell. 1990 Feb 9;60(3):347–350. doi: 10.1016/0092-8674(90)90583-z. [DOI] [PubMed] [Google Scholar]
  22. Korn E. D., Hammer J. A., 3rd Myosin I. Curr Opin Cell Biol. 1990 Feb;2(1):57–61. doi: 10.1016/s0955-0674(05)80031-6. [DOI] [PubMed] [Google Scholar]
  23. Korn E. D., Wright P. L. Macromolecular composition of an amoeba plasma membrane. J Biol Chem. 1973 Jan 25;248(2):439–447. [PubMed] [Google Scholar]
  24. Kunkel T. A., Roberts J. D., Zakour R. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol. 1987;154:367–382. doi: 10.1016/0076-6879(87)54085-x. [DOI] [PubMed] [Google Scholar]
  25. Lynch T. J., Albanesi J. P., Korn E. D., Robinson E. A., Bowers B., Fujisaki H. ATPase activities and actin-binding properties of subfragments of Acanthamoeba myosin IA. J Biol Chem. 1986 Dec 25;261(36):17156–17162. [PubMed] [Google Scholar]
  26. MacLean-Fletcher S., Pollard T. D. Mechanism of action of cytochalasin B on actin. Cell. 1980 Jun;20(2):329–341. doi: 10.1016/0092-8674(80)90619-4. [DOI] [PubMed] [Google Scholar]
  27. 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]
  28. Maruta H., Gadasi H., Collins J. H., Korn E. D. Multiple forms of Acanthamoeba myosin I. J Biol Chem. 1979 May 10;254(9):3624–3630. [PubMed] [Google Scholar]
  29. Matsudaira P. T., Burgess D. R. Identification and organization of the components in the isolated microvillus cytoskeleton. J Cell Biol. 1979 Dec;83(3):667–673. doi: 10.1083/jcb.83.3.667. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Mayer L. D., Hope M. J., Cullis P. R. Vesicles of variable sizes produced by a rapid extrusion procedure. Biochim Biophys Acta. 1986 Jun 13;858(1):161–168. doi: 10.1016/0005-2736(86)90302-0. [DOI] [PubMed] [Google Scholar]
  31. Miyata H., Bowers B., Korn E. D. Plasma membrane association of Acanthamoeba myosin I. J Cell Biol. 1989 Oct;109(4 Pt 1):1519–1528. doi: 10.1083/jcb.109.4.1519. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Montell C., Rubin G. M. The Drosophila ninaC locus encodes two photoreceptor cell specific proteins with domains homologous to protein kinases and the myosin heavy chain head. Cell. 1988 Mar 11;52(5):757–772. doi: 10.1016/0092-8674(88)90413-8. [DOI] [PubMed] [Google Scholar]
  33. Pollard T. D., Doberstein S. K., Zot H. G. Myosin-I. Annu Rev Physiol. 1991;53:653–681. doi: 10.1146/annurev.ph.53.030191.003253. [DOI] [PubMed] [Google Scholar]
  34. Pollard T. D., Korn E. D. Acanthamoeba myosin. I. Isolation from Acanthamoeba castellanii of an enzyme similar to muscle myosin. J Biol Chem. 1973 Jul 10;248(13):4682–4690. [PubMed] [Google Scholar]
  35. Porter J. A., Hicks J. L., Williams D. S., Montell C. Differential localizations of and requirements for the two Drosophila ninaC kinase/myosins in photoreceptor cells. J Cell Biol. 1992 Feb;116(3):683–693. doi: 10.1083/jcb.116.3.683. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Rimm D. L., Sinard J. H., Pollard T. D. Location of the head-tail junction of myosin. J Cell Biol. 1989 May;108(5):1783–1789. doi: 10.1083/jcb.108.5.1783. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Rodaway A. R., Sternberg M. J., Bentley D. L. Similarity in membrane proteins. Nature. 1989 Dec 7;342(6250):624–624. doi: 10.1038/342624a0. [DOI] [PubMed] [Google Scholar]
  38. Sinard J. H., Pollard T. D. Microinjection into Acanthamoeba castellanii of monoclonal antibodies to myosin-II slows but does not stop cell locomotion. Cell Motil Cytoskeleton. 1989;12(1):42–52. doi: 10.1002/cm.970120106. [DOI] [PubMed] [Google Scholar]
  39. Stahl M. L., Ferenz C. R., Kelleher K. L., Kriz R. W., Knopf J. L. Sequence similarity of phospholipase C with the non-catalytic region of src. Nature. 1988 Mar 17;332(6161):269–272. doi: 10.1038/332269a0. [DOI] [PubMed] [Google Scholar]
  40. Steers E., Jr, Cuatrecasas P., Pollard H. B. The purification of beta-galactosidase from Escherichia coli by affinity chromatography. J Biol Chem. 1971 Jan 10;246(1):196–200. [PubMed] [Google Scholar]
  41. Stull J. T., Buss J. E. Phosphorylation of cardiac troponin by cyclic adenosine 3':5'-monophosphate-dependent protein kinase. J Biol Chem. 1977 Feb 10;252(3):851–857. [PubMed] [Google Scholar]
  42. Titus M. A., Warrick H. M., Spudich J. A. Multiple actin-based motor genes in Dictyostelium. Cell Regul. 1989 Nov;1(1):55–63. doi: 10.1091/mbc.1.1.55. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Todderud G., Wahl M. I., Rhee S. G., Carpenter G. Stimulation of phospholipase C-gamma 1 membrane association by epidermal growth factor. Science. 1990 Jul 20;249(4966):296–298. doi: 10.1126/science.2374928. [DOI] [PubMed] [Google Scholar]
  44. Ulsamer A. G., Smith F. R., Korn E. D. Lipids of Acanthamoeba castellanii. Composition and effects of phagocytosis on incorporation of radioactive precursors. J Cell Biol. 1969 Oct;43(1):105–114. doi: 10.1083/jcb.43.1.105. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Ulsamer A. G., Wright P. L., Wetzel M. G., Korn E. D. Plasma and phagosome membranes of Acanthamoeba castellanii. J Cell Biol. 1971 Oct;51(1):193–215. doi: 10.1083/jcb.51.1.193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Zot H. G., Doberstein S. K., Pollard T. D. Myosin-I moves actin filaments on a phospholipid substrate: implications for membrane targeting. J Cell Biol. 1992 Jan;116(2):367–376. doi: 10.1083/jcb.116.2.367. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES