Skip to main content
NCBI home page
The EMBO Journal logoLink to The EMBO Journal
. 1991 Apr;10(4):757–766. doi: 10.1002/j.1460-2075.1991.tb08007.x

Mbh 1: a novel gelsolin/severin-related protein which binds actin in vitro and exhibits nuclear localization in vivo.

G C Prendergast 1, E B Ziff 1
PMCID: PMC452713  PMID: 1849072

Abstract

We describe the characterization of a novel cDNA, mbh1 (myc basic motif homolog-1), which was found during a search for candidate factors which might interact with the c-Myc oncoprotein. Embedded within the amino acid sequence encoded by mbh1 is a region distantly related to the basic/helix-loop-helix (B/HLH) DNA-binding motif and a potential nuclear localization signal. Mbh1 encodes a polypeptide structurally similar to the actin-severing proteins gelsolin and severin. Translation of mbh1 RNA in rabbit reticulocyte extracts produces an approximately 45 kd protein capable of binding actin-coupled agarose beads in vitro in a Ca2(+)-dependent manner. Antiserum raised to a trpE/mbh1 bacterial fusion protein recognizes an approximately 45 kb protein in murine 3T3 fibroblasts, suggesting that the cDNA encodes the complete Mbh1 protein. Examination of Mbh1 localization in 3T3 fibroblasts by indirect immunofluorescence reveals a larger cell population showing diffuse staining, and a smaller population exhibiting a distinct nuclear stain. Western analysis corroborates this intracellular localization and indicates that total cellular levels and localization of Mbh1 are not affected by the cell growth state. The data suggest that Mbh1 may play a role in regulating cytoplasmic and/or nuclear architecture through potential interactions with actin.

Full text

PDF
757

Images in this article

758Image
on p.758
760Image
on p.760
761Image
on p.761
762Image
on p.762
763Image
on p.763
764Image
on p.764

Selected References

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

  1. Ampe C., Vandekerckhove J. The F-actin capping proteins of Physarum polycephalum: cap42(a) is very similar, if not identical, to fragmin and is structurally and functionally very homologous to gelsolin; cap42(b) is Physarum actin. EMBO J. 1987 Dec 20;6(13):4149–4157. doi: 10.1002/j.1460-2075.1987.tb02761.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. André E., Lottspeich F., Schleicher M., Noegel A. Severin, gelsolin, and villin share a homologous sequence in regions presumed to contain F-actin severing domains. J Biol Chem. 1988 Jan 15;263(2):722–727. [PubMed] [Google Scholar]
  3. Ankenbauer T., Kleinschmidt J. A., Walsh M. J., Weiner O. H., Franke W. W. Identification of a widespread nuclear actin binding protein. Nature. 1989 Dec 14;342(6251):822–825. doi: 10.1038/342822a0. [DOI] [PubMed] [Google Scholar]
  4. Battey J., Moulding C., Taub R., Murphy W., Stewart T., Potter H., Lenoir G., Leder P. The human c-myc oncogene: structural consequences of translocation into the IgH locus in Burkitt lymphoma. Cell. 1983 Oct;34(3):779–787. doi: 10.1016/0092-8674(83)90534-2. [DOI] [PubMed] [Google Scholar]
  5. Bazari W. L., Matsudaira P., Wallek M., Smeal T., Jakes R., Ahmed Y. Villin sequence and peptide map identify six homologous domains. Proc Natl Acad Sci U S A. 1988 Jul;85(14):4986–4990. doi: 10.1073/pnas.85.14.4986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Beckmann H., Su L. K., Kadesch T. TFE3: a helix-loop-helix protein that activates transcription through the immunoglobulin enhancer muE3 motif. Genes Dev. 1990 Feb;4(2):167–179. doi: 10.1101/gad.4.2.167. [DOI] [PubMed] [Google Scholar]
  7. Benezra R., Davis R. L., Lockshon D., Turner D. L., Weintraub H. The protein Id: a negative regulator of helix-loop-helix DNA binding proteins. Cell. 1990 Apr 6;61(1):49–59. doi: 10.1016/0092-8674(90)90214-y. [DOI] [PubMed] [Google Scholar]
  8. Blackwell T. K., Kretzner L., Blackwood E. M., Eisenman R. N., Weintraub H. Sequence-specific DNA binding by the c-Myc protein. Science. 1990 Nov 23;250(4984):1149–1151. doi: 10.1126/science.2251503. [DOI] [PubMed] [Google Scholar]
  9. Cai M., Davis R. W. Yeast centromere binding protein CBF1, of the helix-loop-helix protein family, is required for chromosome stability and methionine prototrophy. Cell. 1990 May 4;61(3):437–446. doi: 10.1016/0092-8674(90)90525-j. [DOI] [PubMed] [Google Scholar]
  10. Carron C. P., Hwo S. Y., Dingus J., Benson D. M., Meza I., Bryan J. A re-evaluation of cytoplasmic gelsolin localization. J Cell Biol. 1986 Jan;102(1):237–245. doi: 10.1083/jcb.102.1.237. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Church G. M., Gilbert W. Genomic sequencing. Proc Natl Acad Sci U S A. 1984 Apr;81(7):1991–1995. doi: 10.1073/pnas.81.7.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Clark T. G., Merriam R. W. Diffusible and bound actin nuclei of Xenopus laevis oocytes. Cell. 1977 Dec;12(4):883–891. doi: 10.1016/0092-8674(77)90152-0. [DOI] [PubMed] [Google Scholar]
  13. Cooper J. A., Loftus D. J., Frieden C., Bryan J., Elson E. L. Localization and mobility of gelsolin in cells. J Cell Biol. 1988 Apr;106(4):1229–1240. doi: 10.1083/jcb.106.4.1229. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dang C. V., Lee W. M. Nuclear and nucleolar targeting sequences of c-erb-A, c-myb, N-myc, p53, HSP70, and HIV tat proteins. J Biol Chem. 1989 Oct 25;264(30):18019–18023. [PubMed] [Google Scholar]
  15. Davis R. L., Cheng P. F., Lassar A. B., Weintraub H. The MyoD DNA binding domain contains a recognition code for muscle-specific gene activation. Cell. 1990 Mar 9;60(5):733–746. doi: 10.1016/0092-8674(90)90088-v. [DOI] [PubMed] [Google Scholar]
  16. Davis R. L., Weintraub H., Lassar A. B. Expression of a single transfected cDNA converts fibroblasts to myoblasts. Cell. 1987 Dec 24;51(6):987–1000. doi: 10.1016/0092-8674(87)90585-x. [DOI] [PubMed] [Google Scholar]
  17. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Dieffenbach C. W., SenGupta D. N., Krause D., Sawzak D., Silverman R. H. Cloning of murine gelsolin and its regulation during differentiation of embryonal carcinoma cells. J Biol Chem. 1989 Aug 5;264(22):13281–13288. [PubMed] [Google Scholar]
  19. Egly J. M., Miyamoto N. G., Moncollin V., Chambon P. Is actin a transcription initiation factor for RNA polymerase B? EMBO J. 1984 Oct;3(10):2363–2371. doi: 10.1002/j.1460-2075.1984.tb02141.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Frohman M. A., Dush M. K., Martin G. R. Rapid production of full-length cDNAs from rare transcripts: amplification using a single gene-specific oligonucleotide primer. Proc Natl Acad Sci U S A. 1988 Dec;85(23):8998–9002. doi: 10.1073/pnas.85.23.8998. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Gizang-Ginsberg E., Ziff E. B. Nerve growth factor regulates tyrosine hydroxylase gene transcription through a nucleoprotein complex that contains c-Fos. Genes Dev. 1990 Apr;4(4):477–491. doi: 10.1101/gad.4.4.477. [DOI] [PubMed] [Google Scholar]
  22. Gorham J. D., Baker H., Kegler D., Ziff E. B. The expression of the neuronal intermediate filament protein peripherin in the rat embryo. Brain Res Dev Brain Res. 1990 Dec 15;57(2):235–248. doi: 10.1016/0165-3806(90)90049-5. [DOI] [PubMed] [Google Scholar]
  23. Jackson D. A., Cook P. R. Replication occurs at a nucleoskeleton. EMBO J. 1986 Jun;5(6):1403–1410. doi: 10.1002/j.1460-2075.1986.tb04374.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Jackson D. A., Cook P. R. Transcription occurs at a nucleoskeleton. EMBO J. 1985 Apr;4(4):919–925. doi: 10.1002/j.1460-2075.1985.tb03719.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Jockusch B. M., Becker M., Hindennach I., Jockusch H. Slime mould actin: homology to vertebrate actin and presence in the nucleus. Exp Cell Res. 1974 Dec;89(2):241–246. doi: 10.1016/0014-4827(74)90787-3. [DOI] [PubMed] [Google Scholar]
  26. Karlin S., Altschul S. F. Methods for assessing the statistical significance of molecular sequence features by using general scoring schemes. Proc Natl Acad Sci U S A. 1990 Mar;87(6):2264–2268. doi: 10.1073/pnas.87.6.2264. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Kouzarides T., Ziff E. The role of the leucine zipper in the fos-jun interaction. Nature. 1988 Dec 15;336(6200):646–651. doi: 10.1038/336646a0. [DOI] [PubMed] [Google Scholar]
  28. 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]
  29. 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]
  30. Landschulz W. H., Johnson P. F., McKnight S. L. The leucine zipper: a hypothetical structure common to a new class of DNA binding proteins. Science. 1988 Jun 24;240(4860):1759–1764. doi: 10.1126/science.3289117. [DOI] [PubMed] [Google Scholar]
  31. Lawrence J. B., Singer R. H., Marselle L. M. Highly localized tracks of specific transcripts within interphase nuclei visualized by in situ hybridization. Cell. 1989 May 5;57(3):493–502. doi: 10.1016/0092-8674(89)90924-0. [DOI] [PubMed] [Google Scholar]
  32. Magri E., Zaccarini M., Grazi E. The interaction of histone and protamine with actin. Possible involvement in the formation of the mitotic spindle. Biochem Biophys Res Commun. 1978 Jun 29;82(4):1207–1210. doi: 10.1016/0006-291x(78)90315-7. [DOI] [PubMed] [Google Scholar]
  33. 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]
  34. 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]
  35. Murre C., McCaw P. S., Baltimore D. A new DNA binding and dimerization motif in immunoglobulin enhancer binding, daughterless, MyoD, and myc proteins. Cell. 1989 Mar 10;56(5):777–783. doi: 10.1016/0092-8674(89)90682-x. [DOI] [PubMed] [Google Scholar]
  36. Murre C., McCaw P. S., Vaessin H., Caudy M., Jan L. Y., Jan Y. N., Cabrera C. V., Buskin J. N., Hauschka S. D., Lassar A. B. Interactions between heterologous helix-loop-helix proteins generate complexes that bind specifically to a common DNA sequence. Cell. 1989 Aug 11;58(3):537–544. doi: 10.1016/0092-8674(89)90434-0. [DOI] [PubMed] [Google Scholar]
  37. 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]
  38. Prendergast G. C., Cole M. D. Posttranscriptional regulation of cellular gene expression by the c-myc oncogene. Mol Cell Biol. 1989 Jan;9(1):124–134. doi: 10.1128/mcb.9.1.124. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Prendergast G. C., Ziff E. B. DNA-binding motif. Nature. 1989 Oct 5;341(6241):392–392. doi: 10.1038/341392a0. [DOI] [PubMed] [Google Scholar]
  40. Prendergast G. C., Ziff E. B. Methylation-sensitive sequence-specific DNA binding by the c-Myc basic region. Science. 1991 Jan 11;251(4990):186–189. doi: 10.1126/science.1987636. [DOI] [PubMed] [Google Scholar]
  41. Rajchel A., Chan Y. L., Wool I. G. The primary structure of rat ribosomal protein L32. Nucleic Acids Res. 1988 Mar 25;16(5):2347–2347. doi: 10.1093/nar/16.5.2347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Richardson J. S., Richardson D. C. Amino acid preferences for specific locations at the ends of alpha helices. Science. 1988 Jun 17;240(4859):1648–1652. doi: 10.1126/science.3381086. [DOI] [PubMed] [Google Scholar]
  43. Rimm D. L., Pollard T. D. Purification and characterization of an Acanthamoeba nuclear actin-binding protein. J Cell Biol. 1989 Aug;109(2):585–591. doi: 10.1083/jcb.109.2.585. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Scheer U., Hinssen H., Franke W. W., Jockusch B. M. Microinjection of actin-binding proteins and actin antibodies demonstrates involvement of nuclear actin in transcription of lampbrush chromosomes. Cell. 1984 Nov;39(1):111–122. doi: 10.1016/0092-8674(84)90196-x. [DOI] [PubMed] [Google Scholar]
  45. Schröder H. C., Zahn R. K., Müller W. E. Role of actin and tubulin in the regulation of poly(A) polymerase-endoribonuclease IV complex from calf thymus. J Biol Chem. 1982 Mar 10;257(5):2305–2309. [PubMed] [Google Scholar]
  46. Schuler G. D., Altschul S. F., Lipman D. J. A workbench for multiple alignment construction and analysis. Proteins. 1991;9(3):180–190. doi: 10.1002/prot.340090304. [DOI] [PubMed] [Google Scholar]
  47. Spector D. L. Higher order nuclear organization: three-dimensional distribution of small nuclear ribonucleoprotein particles. Proc Natl Acad Sci U S A. 1990 Jan;87(1):147–151. doi: 10.1073/pnas.87.1.147. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Spindler K. R., Rosser D. S., Berk A. J. Analysis of adenovirus transforming proteins from early regions 1A and 1B with antisera to inducible fusion antigens produced in Escherichia coli. J Virol. 1984 Jan;49(1):132–141. doi: 10.1128/jvi.49.1.132-141.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. 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]
  50. Villares R., Cabrera C. V. The achaete-scute gene complex of D. melanogaster: conserved domains in a subset of genes required for neurogenesis and their homology to myc. Cell. 1987 Jul 31;50(3):415–424. doi: 10.1016/0092-8674(87)90495-8. [DOI] [PubMed] [Google Scholar]
  51. 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]
  52. 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]
  53. Yin H. L., Janmey P. A., Schleicher M. Severin is a gelsolin prototype. FEBS Lett. 1990 May 7;264(1):78–80. doi: 10.1016/0014-5793(90)80769-f. [DOI] [PubMed] [Google Scholar]
  54. Yin H. L., Stossel T. P. Control of cytoplasmic actin gel-sol transformation by gelsolin, a calcium-dependent regulatory protein. Nature. 1979 Oct 18;281(5732):583–586. doi: 10.1038/281583a0. [DOI] [PubMed] [Google Scholar]

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

RESOURCES