The alpha-helical rod domain of human lamins A and C contains a chromatin binding site (original) (raw)

Abstract

We examined regions of human lamins A and C involved in binding to surfaces of mitotic chromosomes. An Escherichia coli expression system was used to produce full-length lamin A and lamin C, and truncated lamins retaining the central alpha-helical rod domain (residues 34-388) but lacking various amounts of the amino-terminal 'head' and carboxy-terminal 'tail' domains. We found that lamin A, lamin C and lamin fragments lacking the head domain and tail sequences distal to residue 431 efficiently assembled into paracrystals and strongly associated with mitotic chromosomes. Furthermore, the lamin rod domain also associated with chromosomes, although efficient chromosome coating required the pH 5-6 conditions needed to assemble the rod into higher order structures. Biochemical assays showed that chromosomes substantially reduced the critical concentration for assembly of lamin polypeptides into pelletable structures. Association of the lamin rod with chromosomes was abolished by pretrypsinization of chromosomes, and was not seen for vimentin (which possesses a similar rod domain). These data demonstrate that the alpha-helical rod of lamins A and C contains a specific chromosome binding site. Hence, the central rod domain of intermediate filament proteins can be involved in interactions with other cellular structures as well as in filament assembly.

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  1. Aaronson R. P., Blobel G. On the attachment of the nuclear pore complex. J Cell Biol. 1974 Sep;62(3):746–754. doi: 10.1083/jcb.62.3.746. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Aebi U., Cohn J., Buhle L., Gerace L. The nuclear lamina is a meshwork of intermediate-type filaments. Nature. 1986 Oct 9;323(6088):560–564. doi: 10.1038/323560a0. [DOI] [PubMed] [Google Scholar]
  3. Benavente R., Krohne G. Involvement of nuclear lamins in postmitotic reorganization of chromatin as demonstrated by microinjection of lamin antibodies. J Cell Biol. 1986 Nov;103(5):1847–1854. doi: 10.1083/jcb.103.5.1847. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Burnette W. N. "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem. 1981 Apr;112(2):195–203. doi: 10.1016/0003-2697(81)90281-5. [DOI] [PubMed] [Google Scholar]
  5. Dessev G. N. Nuclear envelope structure. Curr Opin Cell Biol. 1992 Jun;4(3):430–435. doi: 10.1016/0955-0674(92)90008-z. [DOI] [PubMed] [Google Scholar]
  6. Fisher D. Z., Chaudhary N., Blobel G. cDNA sequencing of nuclear lamins A and C reveals primary and secondary structural homology to intermediate filament proteins. Proc Natl Acad Sci U S A. 1986 Sep;83(17):6450–6454. doi: 10.1073/pnas.83.17.6450. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Foisner R., Leichtfried F. E., Herrmann H., Small J. V., Lawson D., Wiche G. Cytoskeleton-associated plectin: in situ localization, in vitro reconstitution, and binding to immobilized intermediate filament proteins. J Cell Biol. 1988 Mar;106(3):723–733. doi: 10.1083/jcb.106.3.723. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Franke W. W. Nuclear lamins and cytoplasmic intermediate filament proteins: a growing multigene family. Cell. 1987 Jan 16;48(1):3–4. doi: 10.1016/0092-8674(87)90345-x. [DOI] [PubMed] [Google Scholar]
  9. Franke W. W., Scheer U., Krohne G., Jarasch E. D. The nuclear envelope and the architecture of the nuclear periphery. J Cell Biol. 1981 Dec;91(3 Pt 2):39s–50s. doi: 10.1083/jcb.91.3.39s. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Franke W. W. Structure, biochemistry, and functions of the nuclear envelope. Int Rev Cytol. 1974;Suppl 4:71–236. [PubMed] [Google Scholar]
  11. Georgatos S. D., Blobel G. Two distinct attachment sites for vimentin along the plasma membrane and the nuclear envelope in avian erythrocytes: a basis for a vectorial assembly of intermediate filaments. J Cell Biol. 1987 Jul;105(1):105–115. doi: 10.1083/jcb.105.1.105. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Georgatos S. D., Weaver D. C., Marchesi V. T. Site specificity in vimentin-membrane interactions: intermediate filament subunits associate with the plasma membrane via their head domains. J Cell Biol. 1985 Jun;100(6):1962–1967. doi: 10.1083/jcb.100.6.1962. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Georgatos S. D., Weber K., Geisler N., Blobel G. Binding of two desmin derivatives to the plasma membrane and the nuclear envelope of avian erythrocytes: evidence for a conserved site-specificity in intermediate filament-membrane interactions. Proc Natl Acad Sci U S A. 1987 Oct;84(19):6780–6784. doi: 10.1073/pnas.84.19.6780. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gerace L., Blum A., Blobel G. Immunocytochemical localization of the major polypeptides of the nuclear pore complex-lamina fraction. Interphase and mitotic distribution. J Cell Biol. 1978 Nov;79(2 Pt 1):546–566. doi: 10.1083/jcb.79.2.546. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gerace L., Burke B. Functional organization of the nuclear envelope. Annu Rev Cell Biol. 1988;4:335–374. doi: 10.1146/annurev.cb.04.110188.002003. [DOI] [PubMed] [Google Scholar]
  16. Gieffers C., Krohne G. In vitro reconstitution of recombinant lamin A and a lamin A mutant lacking the carboxy-terminal tail. Eur J Cell Biol. 1991 Aug;55(2):191–199. [PubMed] [Google Scholar]
  17. Glass J. R., Gerace L. Lamins A and C bind and assemble at the surface of mitotic chromosomes. J Cell Biol. 1990 Sep;111(3):1047–1057. doi: 10.1083/jcb.111.3.1047. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Heitlinger E., Peter M., Häner M., Lustig A., Aebi U., Nigg E. A. Expression of chicken lamin B2 in Escherichia coli: characterization of its structure, assembly, and molecular interactions. J Cell Biol. 1991 May;113(3):485–495. doi: 10.1083/jcb.113.3.485. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Heitlinger E., Peter M., Lustig A., Villiger W., Nigg E. A., Aebi U. The role of the head and tail domain in lamin structure and assembly: analysis of bacterially expressed chicken lamin A and truncated B2 lamins. J Struct Biol. 1992 Jan-Feb;108(1):74–89. doi: 10.1016/1047-8477(92)90009-y. [DOI] [PubMed] [Google Scholar]
  20. Höger T. H., Krohne G., Kleinschmidt J. A. Interaction of Xenopus lamins A and LII with chromatin in vitro mediated by a sequence element in the carboxyterminal domain. Exp Cell Res. 1991 Dec;197(2):280–289. doi: 10.1016/0014-4827(91)90434-v. [DOI] [PubMed] [Google Scholar]
  21. Höger T. H., Zatloukal K., Waizenegger I., Krohne G. Characterization of a second highly conserved B-type lamin present in cells previously thought to contain only a single B-type lamin. Chromosoma. 1990 Oct;99(6):379–390. doi: 10.1007/BF01726689. [DOI] [PubMed] [Google Scholar]
  22. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  23. Ludérus M. E., de Graaf A., Mattia E., den Blaauwen J. L., Grande M. A., de Jong L., van Driel R. Binding of matrix attachment regions to lamin B1. Cell. 1992 Sep 18;70(6):949–959. doi: 10.1016/0092-8674(92)90245-8. [DOI] [PubMed] [Google Scholar]
  24. McKeon F. D., Kirschner M. W., Caput D. Homologies in both primary and secondary structure between nuclear envelope and intermediate filament proteins. Nature. 1986 Feb 6;319(6053):463–468. doi: 10.1038/319463a0. [DOI] [PubMed] [Google Scholar]
  25. McLachlan A. D., Stewart M. The 14-fold periodicity in alpha-tropomyosin and the interaction with actin. J Mol Biol. 1976 May 15;103(2):271–298. doi: 10.1016/0022-2836(76)90313-2. [DOI] [PubMed] [Google Scholar]
  26. McLeod M., Stein M., Beach D. The product of the mei3+ gene, expressed under control of the mating-type locus, induces meiosis and sporulation in fission yeast. EMBO J. 1987 Mar;6(3):729–736. doi: 10.1002/j.1460-2075.1987.tb04814.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Meier J., Campbell K. H., Ford C. C., Stick R., Hutchison C. J. The role of lamin LIII in nuclear assembly and DNA replication, in cell-free extracts of Xenopus eggs. J Cell Sci. 1991 Mar;98(Pt 3):271–279. doi: 10.1242/jcs.98.3.271. [DOI] [PubMed] [Google Scholar]
  28. Moir R. D., Donaldson A. D., Stewart M. Expression in Escherichia coli of human lamins A and C: influence of head and tail domains on assembly properties and paracrystal formation. J Cell Sci. 1991 Jun;99(Pt 2):363–372. doi: 10.1242/jcs.99.2.363. [DOI] [PubMed] [Google Scholar]
  29. Nelson W. J., Traub P. Purification of the intermediate filament protein vimentin from Ehrlich ascites tumor cells. J Biol Chem. 1982 May 25;257(10):5536–5543. [PubMed] [Google Scholar]
  30. Newport J. W., Wilson K. L., Dunphy W. G. A lamin-independent pathway for nuclear envelope assembly. J Cell Biol. 1990 Dec;111(6 Pt 1):2247–2259. doi: 10.1083/jcb.111.6.2247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Parry D. A., Steinert P. M. Intermediate filament structure. Curr Opin Cell Biol. 1992 Feb;4(1):94–98. doi: 10.1016/0955-0674(92)90064-j. [DOI] [PubMed] [Google Scholar]
  32. Peter M., Kitten G. T., Lehner C. F., Vorburger K., Bailer S. M., Maridor G., Nigg E. A. Cloning and sequencing of cDNA clones encoding chicken lamins A and B1 and comparison of the primary structures of vertebrate A- and B-type lamins. J Mol Biol. 1989 Aug 5;208(3):393–404. doi: 10.1016/0022-2836(89)90504-4. [DOI] [PubMed] [Google Scholar]
  33. Saiki R. K., Gelfand D. H., Stoffel S., Scharf S. J., Higuchi R., Horn G. T., Mullis K. B., Erlich H. A. Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science. 1988 Jan 29;239(4839):487–491. doi: 10.1126/science.2448875. [DOI] [PubMed] [Google Scholar]
  34. Shoeman R. L., Traub P. The in vitro DNA-binding properties of purified nuclear lamin proteins and vimentin. J Biol Chem. 1990 Jun 5;265(16):9055–9061. [PubMed] [Google Scholar]
  35. Steinert P. M., Roop D. R. Molecular and cellular biology of intermediate filaments. Annu Rev Biochem. 1988;57:593–625. doi: 10.1146/annurev.bi.57.070188.003113. [DOI] [PubMed] [Google Scholar]
  36. Stewart M. Intermediate filaments: structure, assembly and molecular interactions. Curr Opin Cell Biol. 1990 Feb;2(1):91–100. doi: 10.1016/s0955-0674(05)80037-7. [DOI] [PubMed] [Google Scholar]
  37. Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
  38. Tabor S., Richardson C. C. A bacteriophage T7 RNA polymerase/promoter system for controlled exclusive expression of specific genes. Proc Natl Acad Sci U S A. 1985 Feb;82(4):1074–1078. doi: 10.1073/pnas.82.4.1074. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Yuan J., Simos G., Blobel G., Georgatos S. D. Binding of lamin A to polynucleosomes. J Biol Chem. 1991 May 15;266(14):9211–9215. [PubMed] [Google Scholar]