Skip to main content
NCBI home page
The EMBO Journal logoLink to The EMBO Journal
. 1989 Dec 1;8(12):3587–3593. doi: 10.1002/j.1460-2075.1989.tb08531.x

Alpha-actinin synthesis can be modulated by antisense probes and is autoregulated in non-muscle cells.

H Schulze 1, A Huckriede 1, A A Noegel 1, M Schleicher 1, B M Jockusch 1
PMCID: PMC402039  PMID: 2479547

Abstract

We used a 279 bp cDNA probe derived from a Dictyostelium alpha-actinin genomic sequence to assay the degree of homology between alpha-actinin from slime molds, mammalian and chicken cells. Recognition of this probe by vertebrate cells was shown in Southern and Northern blots, and by antisense RNA-induced depression of endogenous alpha-actinin synthesis in living cells. Micro-injection of Dictyostelium or chicken gizzard alpha-actinin resulted in incorporation of these proteins in stress fibers, peripheral microfilament belts and adhesion sites. Alpha-actinin-injected cells showed a marked, transient reduction of synthesis of the corresponding endogenous protein. These data emphasize the high degree of conservation of alpha-actinin during evolution and show for the first time autoregulation of synthesis for a microfilament protein.

Full text

PDF
3587

Images in this article

3588Image
on p.3588
3588Image
on p.3588
3588Image
on p.3588
3589Image
on p.3589
3589Image
on p.3589
3590Image
on p.3590
3591Image
on p.3591

Selected References

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

  1. Baron M. D., Davison M. D., Jones P., Critchley D. R. The sequence of chick alpha-actinin reveals homologies to spectrin and calmodulin. J Biol Chem. 1987 Dec 25;262(36):17623–17629. [PubMed] [Google Scholar]
  2. Baron M. D., Davison M. D., Jones P., Patel B., Critchley D. R. Isolation and characterization of a cDNA encoding a chick alpha-actinin. J Biol Chem. 1987 Feb 25;262(6):2558–2561. [PubMed] [Google Scholar]
  3. Ben-Ze'ev A., Farmer S. R., Penman S. Mechanisms of regulating tubulin synthesis in cultured mammalian cells. Cell. 1979 Jun;17(2):319–325. doi: 10.1016/0092-8674(79)90157-0. [DOI] [PubMed] [Google Scholar]
  4. Bishop J. O., Morton J. G., Rosbash M., Richardson M. Three abundance classes in HeLa cell messenger RNA. Nature. 1974 Jul 19;250(463):199–204. doi: 10.1038/250199a0. [DOI] [PubMed] [Google Scholar]
  5. Blikstad I., Eriksson S., Carlsson L. alpha-Actinin promotes polymerization of actin from profilactin. Eur J Biochem. 1980 Aug;109(2):317–323. doi: 10.1111/j.1432-1033.1980.tb04797.x. [DOI] [PubMed] [Google Scholar]
  6. Blin N., Stafford D. W. A general method for isolation of high molecular weight DNA from eukaryotes. Nucleic Acids Res. 1976 Sep;3(9):2303–2308. doi: 10.1093/nar/3.9.2303. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Boschek C. B., Jockusch B. M., Friis R. R., Back R., Grundmann E., Bauer H. Early changes in the distribution and organization of microfilament proteins during cell transformation. Cell. 1981 Apr;24(1):175–184. doi: 10.1016/0092-8674(81)90513-4. [DOI] [PubMed] [Google Scholar]
  8. Brandtzaeg P. Conjugates of immunoglobulin G with different fluorochromes. I. Characterization by anionic-exchange chromatography. Scand J Immunol. 1973;2(3):273–290. doi: 10.1111/j.1365-3083.1973.tb02037.x. [DOI] [PubMed] [Google Scholar]
  9. Bravo R., Celis J. E. Up-dated catalogue of HeLa cell proteins: percentages and characteristics of the major cell polypeptides labeled with a mixture of 16 14C-labeled amino acids. Clin Chem. 1982 Apr;28(4 Pt 2):766–781. [PubMed] [Google Scholar]
  10. Bretscher A., Vandekerckhove J., Weber K. alpha-Actinins from chicken skeletal muscle and smooth muscle show considerable chemical and immunological differences. Eur J Biochem. 1979 Oct;100(1):237–243. doi: 10.1111/j.1432-1033.1979.tb02054.x. [DOI] [PubMed] [Google Scholar]
  11. Burridge K., Feramisco J. R. Non-muscle alpha actinins are calcium-sensitive actin-binding proteins. Nature. 1981 Dec 10;294(5841):565–567. doi: 10.1038/294565a0. [DOI] [PubMed] [Google Scholar]
  12. Byers H. R., Fujiwara K. Stress fibers in cells in situ: immunofluorescence visualization with antiactin, antimyosin, and anti-alpha-actinin. J Cell Biol. 1982 Jun;93(3):804–811. doi: 10.1083/jcb.93.3.804. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Caron J. M., Jones A. L., Kirschner M. W. Autoregulation of tubulin synthesis in hepatocytes and fibroblasts. J Cell Biol. 1985 Nov;101(5 Pt 1):1763–1772. doi: 10.1083/jcb.101.5.1763. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  15. Cleveland D. W., Lopata M. A., Sherline P., Kirschner M. W. Unpolymerized tubulin modulates the level of tubulin mRNAs. Cell. 1981 Aug;25(2):537–546. doi: 10.1016/0092-8674(81)90072-6. [DOI] [PubMed] [Google Scholar]
  16. Cleveland D. W., Pittenger M. F., Feramisco J. R. Elevation of tubulin levels by microinjection suppresses new tubulin synthesis. Nature. 1983 Oct 20;305(5936):738–740. doi: 10.1038/305738a0. [DOI] [PubMed] [Google Scholar]
  17. Condeelis J., Vahey M. A calcium- and pH-regulated protein from Dictyostelium discoideum that cross-links actin filaments. J Cell Biol. 1982 Aug;94(2):466–471. doi: 10.1083/jcb.94.2.466. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Ebashi S., Ebashi F. Alpha-actinin, a new structural protein from striated muscle. I. Preparation and action on actomyosinàtp interaction. J Biochem. 1965 Jul;58(1):7–12. doi: 10.1093/oxfordjournals.jbchem.a128167. [DOI] [PubMed] [Google Scholar]
  19. Feramisco J. R., Blose S. H. Distribution of fluorescently labeled alpha-actinin in living and fixed fibroblasts. J Cell Biol. 1980 Aug;86(2):608–615. doi: 10.1083/jcb.86.2.608. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Feramisco J. R., Burridge K. A rapid purification of alpha-actinin, filamin, and a 130,000-dalton protein from smooth muscle. J Biol Chem. 1980 Feb 10;255(3):1194–1199. [PubMed] [Google Scholar]
  21. Feramisco J. R. Microinjection of fluorescently labeled alpha-actinin into living fibroblasts. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3967–3971. doi: 10.1073/pnas.76.8.3967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Fujiwara K., Porter M. E., Pollard T. D. Alpha-actinin localization in the cleavage furrow during cytokinesis. J Cell Biol. 1978 Oct;79(1):268–275. doi: 10.1083/jcb.79.1.268. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Füchtbauer A., Jockusch B. M., Maruta H., Kilimann M. W., Isenberg G. Disruption of microfilament organization after injection of F-actin capping proteins into living tissue culture cells. 1983 Jul 28-Aug 3Nature. 304(5924):361–364. doi: 10.1038/304361a0. [DOI] [PubMed] [Google Scholar]
  24. Gache Y., Landon F., Olomucki A. Polymorphism of alpha-actinin from human blood platelets. Homodimeric and heterodimeric forms. Eur J Biochem. 1984 May 15;141(1):57–61. doi: 10.1111/j.1432-1033.1984.tb08156.x. [DOI] [PubMed] [Google Scholar]
  25. Geiger B., Singer S. J. The participation of alpha-actinin in the capping of cell membrane components. Cell. 1979 Jan;16(1):213–222. doi: 10.1016/0092-8674(79)90202-2. [DOI] [PubMed] [Google Scholar]
  26. Gordon W. E., 3rd Immunofluorescent and ultrastructural studies of "sarcomeric" units in stress fibers of cultured non-muscle cells. Exp Cell Res. 1978 Dec;117(2):253–260. doi: 10.1016/0014-4827(78)90138-6. [DOI] [PubMed] [Google Scholar]
  27. Graessmann M., Graessman A. "Early" simian-virus-40-specific RNA contains information for tumor antigen formation and chromatin replication. Proc Natl Acad Sci U S A. 1976 Feb;73(2):366–370. doi: 10.1073/pnas.73.2.366. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Hoessli D., Rungger-Brändle E., Jockusch B. M., Gabbiani G. Lymphocyte alpha-actinin. Relationship to cell membrane and co-capping with surface receptors. J Cell Biol. 1980 Feb;84(2):305–314. doi: 10.1083/jcb.84.2.305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Jockusch B. M., Isenberg G. Interaction of alpha-actinin and vinculin with actin: opposite effects on filament network formation. Proc Natl Acad Sci U S A. 1981 May;78(5):3005–3009. doi: 10.1073/pnas.78.5.3005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Jockusch H., Jockusch B. M. Structural organization of the Z-line protein, alpha-actinin, in developing skeletal muscle cells. Dev Biol. 1980 Mar;75(1):231–238. doi: 10.1016/0012-1606(80)90158-x. [DOI] [PubMed] [Google Scholar]
  31. Krawczyk Z., Wu C. Isolation of RNA for dot hybridization by heparin-DNase I treatment of whole cell lysate. Anal Biochem. 1987 Aug 15;165(1):20–27. doi: 10.1016/0003-2697(87)90195-3. [DOI] [PubMed] [Google Scholar]
  32. Kreis T. E., Birchmeier W. Stress fiber sarcomeres of fibroblasts are contractile. Cell. 1980 Nov;22(2 Pt 2):555–561. doi: 10.1016/0092-8674(80)90365-7. [DOI] [PubMed] [Google Scholar]
  33. Landon F., Gache Y., Touitou H., Olomucki A. Properties of two isoforms of human blood platelet alpha-actinin. Eur J Biochem. 1985 Dec 2;153(2):231–237. doi: 10.1111/j.1432-1033.1985.tb09291.x. [DOI] [PubMed] [Google Scholar]
  34. Landon F., Olomucki A. Isolation and physico-chemical properties of blood platelet alpha-actinin. Biochim Biophys Acta. 1983 Jan 12;742(1):129–134. doi: 10.1016/0167-4838(83)90368-0. [DOI] [PubMed] [Google Scholar]
  35. Lane B. P., Elias J., Drummond E. Immunoelectronmicroscopic localization of alpha-actinin in skeletal muscle cells. J Histochem Cytochem. 1977 Jan;25(1):69–72. doi: 10.1177/25.1.833423. [DOI] [PubMed] [Google Scholar]
  36. Langanger G., de Mey J., Moeremans M., Daneels G., de Brabander M., Small J. V. Ultrastructural localization of alpha-actinin and filamin in cultured cells with the immunogold staining (IGS) method. J Cell Biol. 1984 Oct;99(4 Pt 1):1324–1334. doi: 10.1083/jcb.99.4.1324. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Lazarides E., Burridge K. Alpha-actinin: immunofluorescent localization of a muscle structural protein in nonmuscle cells. Cell. 1975 Nov;6(3):289–298. doi: 10.1016/0092-8674(75)90180-4. [DOI] [PubMed] [Google Scholar]
  38. Lemanski L. F., Tu Z. H. Immunofluorescent studies for myosin, actin, tropomyosin and alpha-actinin in cultured cardiomyopathic hamster heart cells. Dev Biol. 1983 Jun;97(2):338–348. doi: 10.1016/0012-1606(83)90091-x. [DOI] [PubMed] [Google Scholar]
  39. Mariash C. N., Seelig S., Oppenheimer J. H. A rapid, inexpensive, quantitative technique for the analysis of two-dimensional electrophoretograms. Anal Biochem. 1982 Apr;121(2):388–394. doi: 10.1016/0003-2697(82)90498-5. [DOI] [PubMed] [Google Scholar]
  40. Masaki T., Endo M., Ebashi S. Localization of 6S component of a alpha-actinin at Z-band. J Biochem. 1967 Nov;62(5):630–632. doi: 10.1093/oxfordjournals.jbchem.a128717. [DOI] [PubMed] [Google Scholar]
  41. 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]
  42. Messing J., Vieira J. A new pair of M13 vectors for selecting either DNA strand of double-digest restriction fragments. Gene. 1982 Oct;19(3):269–276. doi: 10.1016/0378-1119(82)90016-6. [DOI] [PubMed] [Google Scholar]
  43. Noegel A., Witke W., Schleicher M. Calcium-sensitive non-muscle alpha-actinin contains EF-hand structures and highly conserved regions. FEBS Lett. 1987 Sep 14;221(2):391–396. doi: 10.1016/0014-5793(87)80962-6. [DOI] [PubMed] [Google Scholar]
  44. O'Farrell P. H. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [PMC free article] [PubMed] [Google Scholar]
  45. Pachter J. S., Yen T. J., Cleveland D. W. Autoregulation of tubulin expression is achieved through specific degradation of polysomal tubulin mRNAs. Cell. 1987 Oct 23;51(2):283–292. doi: 10.1016/0092-8674(87)90155-3. [DOI] [PubMed] [Google Scholar]
  46. Pittenger M. F., Cleveland D. W. Retention of autoregulatory control of tubulin synthesis in cytoplasts: demonstration of a cytoplasmic mechanism that regulates the level of tubulin expression. J Cell Biol. 1985 Nov;101(5 Pt 1):1941–1952. doi: 10.1083/jcb.101.5.1941. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Podlubnaya Z. A., Tskhovrebova L. A., Zaalishtsbvili M. M., Stefanenko G. A. Electron microscopic study of alpha-actinin. J Mol Biol. 1975 Feb 25;92(2):357–359. doi: 10.1016/0022-2836(75)90234-x. [DOI] [PubMed] [Google Scholar]
  48. Sanger J. M., Mittal B., Pochapin M. B., Sanger J. W. Stress fiber and cleavage furrow formation in living cells microinjected with fluorescently labeled alpha-actinin. Cell Motil Cytoskeleton. 1987;7(3):209–220. doi: 10.1002/cm.970070304. [DOI] [PubMed] [Google Scholar]
  49. Sanger J. W., Mittal B., Sanger J. M. Analysis of myofibrillar structure and assembly using fluorescently labeled contractile proteins. J Cell Biol. 1984 Mar;98(3):825–833. doi: 10.1083/jcb.98.3.825. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Sanger J. W., Mittal B., Sanger J. M. Formation of myofibrils in spreading chick cardiac myocytes. Cell Motil. 1984;4(6):405–416. doi: 10.1002/cm.970040602. [DOI] [PubMed] [Google Scholar]
  51. Sanger J. W., Mittal B., Sanger J. M. Interaction of fluorescently-labeled contractile proteins with the cytoskeleton in cell models. J Cell Biol. 1984 Sep;99(3):918–928. doi: 10.1083/jcb.99.3.918. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Sanger J. W., Sanger J. M., Jockusch B. M. Differential response of three types of actin filament bundles to depletion of cellular ATP levels. Eur J Cell Biol. 1983 Sep;31(2):197–204. [PubMed] [Google Scholar]
  53. Schachat F. H., Canine A. C., Briggs M. M., Reedy M. C. The presence of two skeletal muscle alpha-actinins correlates with troponin-tropomyosin expression and Z-line width. J Cell Biol. 1985 Sep;101(3):1001–1008. doi: 10.1083/jcb.101.3.1001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Schleicher M., Noegel A., Schwarz T., Wallraff E., Brink M., Faix J., Gerisch G., Isenberg G. A Dictyostelium mutant with severe defects in alpha-actinin: its characterization using cDNA probes and monoclonal antibodies. J Cell Sci. 1988 May;90(Pt 1):59–71. doi: 10.1242/jcs.90.1.59. [DOI] [PubMed] [Google Scholar]
  55. Small J. V., Rinnerthaler G., Hinssen H. Organization of actin meshworks in cultured cells: the leading edge. Cold Spring Harb Symp Quant Biol. 1982;46(Pt 2):599–611. doi: 10.1101/sqb.1982.046.01.056. [DOI] [PubMed] [Google Scholar]
  56. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  57. Stickel S. K., Wang Y. L. Alpha-actinin-containing aggregates in transformed cells are highly dynamic structures. J Cell Biol. 1987 Jun;104(6):1521–1526. doi: 10.1083/jcb.104.6.1521. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Stromer M. H., Goll D. E. Studies on purified -actinin. II. Electron microscopic studies on the competitive binding of -actinin and tropomyosin to Z-line extracted myofibrils. J Mol Biol. 1972 Jun 28;67(3):489–494. doi: 10.1016/0022-2836(72)90465-2. [DOI] [PubMed] [Google Scholar]
  59. Witke W., Nellen W., Noegel A. Homologous recombination in the Dictyostelium alpha-actinin gene leads to an altered mRNA and lack of the protein. EMBO J. 1987 Dec 20;6(13):4143–4148. doi: 10.1002/j.1460-2075.1987.tb02760.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Witke W., Schleicher M., Lottspeich F., Noegel A. Studies on the transcription, translation, and structure of alpha-actinin in Dictyostelium discoideum. J Cell Biol. 1986 Sep;103(3):969–975. doi: 10.1083/jcb.103.3.969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  61. Yen T. J., Gay D. A., Pachter J. S., Cleveland D. W. Autoregulated changes in stability of polyribosome-bound beta-tubulin mRNAs are specified by the first 13 translated nucleotides. Mol Cell Biol. 1988 Mar;8(3):1224–1235. doi: 10.1128/mcb.8.3.1224. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES