Skip to main content
PMC home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1993 Dec 2;123(6):1661–1670. doi: 10.1083/jcb.123.6.1661

The role of CaaX-dependent modifications in membrane association of Xenopus nuclear lamin B3 during meiosis and the fate of B3 in transfected mitotic cells

PMCID: PMC2290876  PMID: 8276888

Abstract

Recent evidence shows that the COOH-terminal CaaX motif of lamins is necessary to target newly synthesized proteins to the nuclear envelope membranes. Isoprenylation at the CaaX-cysteine has been taken to explain the different fates of A- and B-type lamins during cell division. A-type lamins, which loose their isoprenylation shortly after incorporation into the lamina structure, become freely soluble upon mitotic nuclear envelope breakdown. Somatic B-type lamins, in contrast, are permanently isoprenylated and, although depolymerized during mitosis, remain associated with remnants of nuclear envelope membranes. However, Xenopus lamin B3, the major B-type lamin of amphibian oocytes and eggs, becomes soluble after nuclear envelope breakdown in meiotic metaphase. Here we show that Xenopus lamin B3 is permanently isoprenylated and carboxyl methylated in oocytes (interphase) and eggs (meiotic metaphase). When transfected into mouse L cells Xenopus lamin B3 is integrated into the host lamina and responds to cell cycle signals in a normal fashion. Notably, the ectopically expressed Xenopus lamin does not form heterooligomers with the endogenous lamins as revealed by a coprecipitation experiment with mitotic lamins. In contrast to the situation in amphibian eggs, a significant portion of lamin B3 remains associated with membranes during mitosis. We conclude from these data that the CaaX motif-mediated modifications, although necessary, are not sufficient for a stable association of lamins with membranes and that additional factors are involved in lamin-membrane binding.

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. Adachi Y., Luke M., Laemmli U. K. Chromosome assembly in vitro: topoisomerase II is required for condensation. Cell. 1991 Jan 11;64(1):137–148. doi: 10.1016/0092-8674(91)90215-k. [DOI] [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. Appelbaum J., Blobel G., Georgatos S. D. In vivo phosphorylation of the lamin B receptor. Binding of lamin B to its nuclear membrane receptor is affected by phosphorylation. J Biol Chem. 1990 Mar 15;265(8):4181–4184. [PubMed] [Google Scholar]
  4. Bailer S. M., Eppenberger H. M., Griffiths G., Nigg E. A. Characterization of A 54-kD protein of the inner nuclear membrane: evidence for cell cycle-dependent interaction with the nuclear lamina. J Cell Biol. 1991 Aug;114(3):389–400. doi: 10.1083/jcb.114.3.389. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Beck L. A., Hosick T. J., Sinensky M. Incorporation of a product of mevalonic acid metabolism into proteins of Chinese hamster ovary cell nuclei. J Cell Biol. 1988 Oct;107(4):1307–1316. doi: 10.1083/jcb.107.4.1307. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Beck L. A., Hosick T. J., Sinensky M. Isoprenylation is required for the processing of the lamin A precursor. J Cell Biol. 1990 May;110(5):1489–1499. doi: 10.1083/jcb.110.5.1489. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Benavente R., Krohne G., Franke W. W. Cell type-specific expression of nuclear lamina proteins during development of Xenopus laevis. Cell. 1985 May;41(1):177–190. doi: 10.1016/0092-8674(85)90072-8. [DOI] [PubMed] [Google Scholar]
  8. 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]
  9. Bossie C. A., Sanders M. M. A cDNA from Drosophila melanogaster encodes a lamin C-like intermediate filament protein. J Cell Sci. 1993 Apr;104(Pt 4):1263–1272. doi: 10.1242/jcs.104.4.1263. [DOI] [PubMed] [Google Scholar]
  10. Burke B., Gerace L. A cell free system to study reassembly of the nuclear envelope at the end of mitosis. Cell. 1986 Feb 28;44(4):639–652. doi: 10.1016/0092-8674(86)90273-4. [DOI] [PubMed] [Google Scholar]
  11. Casey P. J., Solski P. A., Der C. J., Buss J. E. p21ras is modified by a farnesyl isoprenoid. Proc Natl Acad Sci U S A. 1989 Nov;86(21):8323–8327. doi: 10.1073/pnas.86.21.8323. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Chaudhary N., Courvalin J. C. Stepwise reassembly of the nuclear envelope at the end of mitosis. J Cell Biol. 1993 Jul;122(2):295–306. doi: 10.1083/jcb.122.2.295. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Chelsky D., Gutterson N. I., Koshland D. E., Jr A diffusion assay for detection and quantitation of methyl-esterified proteins on polyacrylamide gels. Anal Biochem. 1984 Aug 15;141(1):143–148. doi: 10.1016/0003-2697(84)90437-8. [DOI] [PubMed] [Google Scholar]
  14. Chelsky D., Olson J. F., Koshland D. E., Jr Cell cycle-dependent methyl esterification of lamin B. J Biol Chem. 1987 Mar 25;262(9):4303–4309. [PubMed] [Google Scholar]
  15. Chelsky D., Sobotka C., O'Neill C. L. Lamin B methylation and assembly into the nuclear envelope. J Biol Chem. 1989 May 5;264(13):7637–7643. [PubMed] [Google Scholar]
  16. Clarke S. Protein isoprenylation and methylation at carboxyl-terminal cysteine residues. Annu Rev Biochem. 1992;61:355–386. doi: 10.1146/annurev.bi.61.070192.002035. [DOI] [PubMed] [Google Scholar]
  17. Courvalin J. C., Segil N., Blobel G., Worman H. J. The lamin B receptor of the inner nuclear membrane undergoes mitosis-specific phosphorylation and is a substrate for p34cdc2-type protein kinase. J Biol Chem. 1992 Sep 25;267(27):19035–19038. [PubMed] [Google Scholar]
  18. Cox A. D., Der C. J. Protein prenylation: more than just glue? Curr Opin Cell Biol. 1992 Dec;4(6):1008–1016. doi: 10.1016/0955-0674(92)90133-w. [DOI] [PubMed] [Google Scholar]
  19. Dagenais A., Bibor-Hardy V., Laliberté J. F., Royal A., Simard R. Detection in BHK cells of a precursor form for lamin A. Exp Cell Res. 1985 Dec;161(2):269–276. doi: 10.1016/0014-4827(85)90084-9. [DOI] [PubMed] [Google Scholar]
  20. Dumont J. N. Oogenesis in Xenopus laevis (Daudin). I. Stages of oocyte development in laboratory maintained animals. J Morphol. 1972 Feb;136(2):153–179. doi: 10.1002/jmor.1051360203. [DOI] [PubMed] [Google Scholar]
  21. Döring V., Stick R. Gene structure of nuclear lamin LIII of Xenopus laevis; a model for the evolution of IF proteins from a lamin-like ancestor. EMBO J. 1990 Dec;9(12):4073–4081. doi: 10.1002/j.1460-2075.1990.tb07629.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. 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]
  23. Foisner R., Gerace L. Integral membrane proteins of the nuclear envelope interact with lamins and chromosomes, and binding is modulated by mitotic phosphorylation. Cell. 1993 Jul 2;73(7):1267–1279. doi: 10.1016/0092-8674(93)90355-t. [DOI] [PubMed] [Google Scholar]
  24. Gerace L., Blobel G. The nuclear envelope lamina is reversibly depolymerized during mitosis. Cell. 1980 Jan;19(1):277–287. doi: 10.1016/0092-8674(80)90409-2. [DOI] [PubMed] [Google Scholar]
  25. 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]
  26. Gerace L., Comeau C., Benson M. Organization and modulation of nuclear lamina structure. J Cell Sci Suppl. 1984;1:137–160. doi: 10.1242/jcs.1984.supplement_1.10. [DOI] [PubMed] [Google Scholar]
  27. Hancock J. F., Magee A. I., Childs J. E., Marshall C. J. All ras proteins are polyisoprenylated but only some are palmitoylated. Cell. 1989 Jun 30;57(7):1167–1177. doi: 10.1016/0092-8674(89)90054-8. [DOI] [PubMed] [Google Scholar]
  28. Hancock J. F., Paterson H., Marshall C. J. A polybasic domain or palmitoylation is required in addition to the CAAX motif to localize p21ras to the plasma membrane. Cell. 1990 Oct 5;63(1):133–139. doi: 10.1016/0092-8674(90)90294-o. [DOI] [PubMed] [Google Scholar]
  29. Heald R., McKeon F. Mutations of phosphorylation sites in lamin A that prevent nuclear lamina disassembly in mitosis. Cell. 1990 May 18;61(4):579–589. doi: 10.1016/0092-8674(90)90470-y. [DOI] [PubMed] [Google Scholar]
  30. Holtz D., Tanaka R. A., Hartwig J., McKeon F. The CaaX motif of lamin A functions in conjunction with the nuclear localization signal to target assembly to the nuclear envelope. Cell. 1989 Dec 22;59(6):969–977. doi: 10.1016/0092-8674(89)90753-8. [DOI] [PubMed] [Google Scholar]
  31. Höger T. H., Krohne G., Franke W. W. Amino acid sequence and molecular characterization of murine lamin B as deduced from cDNA clones. Eur J Cell Biol. 1988 Dec;47(2):283–290. [PubMed] [Google Scholar]
  32. 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]
  33. Kita T., Brown M. S., Goldstein J. L. Feedback regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase in livers of mice treated with mevinolin, a competitive inhibitor of the reductase. J Clin Invest. 1980 Nov;66(5):1094–1100. doi: 10.1172/JCI109938. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Kitten G. T., Nigg E. A. The CaaX motif is required for isoprenylation, carboxyl methylation, and nuclear membrane association of lamin B2. J Cell Biol. 1991 Apr;113(1):13–23. doi: 10.1083/jcb.113.1.13. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Krohne G., Dabauvalle M. C., Franke W. W. Cell type-specific differences in protein composition of nuclear pore complex-lamina structures in oocytes and erythrocytes of Xenopus laevis. J Mol Biol. 1981 Sep 5;151(1):121–141. doi: 10.1016/0022-2836(81)90224-2. [DOI] [PubMed] [Google Scholar]
  36. Krohne G., Debus E., Osborn M., Weber K., Franke W. W. A monoclonal antibody against nuclear lamina proteins reveals cell type-specificity in Xenopus laevis. Exp Cell Res. 1984 Jan;150(1):47–59. doi: 10.1016/0014-4827(84)90700-6. [DOI] [PubMed] [Google Scholar]
  37. Krohne G., Waizenegger I., Höger T. H. The conserved carboxy-terminal cysteine of nuclear lamins is essential for lamin association with the nuclear envelope. J Cell Biol. 1989 Nov;109(5):2003–2011. doi: 10.1083/jcb.109.5.2003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. 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]
  39. Laliberté J. F., Dagenais A., Filion M., Bibor-Hardy V., Simard R., Royal A. Identification of distinct messenger RNAs for nuclear lamin C and a putative precursor of nuclear lamin A. J Cell Biol. 1984 Mar;98(3):980–985. doi: 10.1083/jcb.98.3.980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Lehner C. F., Fürstenberger G., Eppenberger H. M., Nigg E. A. Biogenesis of the nuclear lamina: in vivo synthesis and processing of nuclear protein precursors. Proc Natl Acad Sci U S A. 1986 Apr;83(7):2096–2099. doi: 10.1073/pnas.83.7.2096. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Loewinger L., McKeon F. Mutations in the nuclear lamin proteins resulting in their aberrant assembly in the cytoplasm. EMBO J. 1988 Aug;7(8):2301–2309. doi: 10.1002/j.1460-2075.1988.tb03073.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Lutz R. J., Trujillo M. A., Denham K. S., Wenger L., Sinensky M. Nucleoplasmic localization of prelamin A: implications for prenylation-dependent lamin A assembly into the nuclear lamina. Proc Natl Acad Sci U S A. 1992 Apr 1;89(7):3000–3004. doi: 10.1073/pnas.89.7.3000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. 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]
  44. 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]
  45. 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]
  46. Newport J. Nuclear reconstitution in vitro: stages of assembly around protein-free DNA. Cell. 1987 Jan 30;48(2):205–217. doi: 10.1016/0092-8674(87)90424-7. [DOI] [PubMed] [Google Scholar]
  47. Nigg E. A. Nuclear function and organization: the potential of immunochemical approaches. Int Rev Cytol. 1988;110:27–92. doi: 10.1016/s0074-7696(08)61847-1. [DOI] [PubMed] [Google Scholar]
  48. Nigg E. A. The nuclear envelope. Curr Opin Cell Biol. 1989 Jun;1(3):435–440. doi: 10.1016/0955-0674(89)90002-1. [DOI] [PubMed] [Google Scholar]
  49. Padan R., Nainudel-Epszteyn S., Goitein R., Fainsod A., Gruenbaum Y. Isolation and characterization of the Drosophila nuclear envelope otefin cDNA. J Biol Chem. 1990 May 15;265(14):7808–7813. [PubMed] [Google Scholar]
  50. 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]
  51. Peter M., Nakagawa J., Dorée M., Labbé J. C., Nigg E. A. In vitro disassembly of the nuclear lamina and M phase-specific phosphorylation of lamins by cdc2 kinase. Cell. 1990 May 18;61(4):591–602. doi: 10.1016/0092-8674(90)90471-p. [DOI] [PubMed] [Google Scholar]
  52. Reiss Y., Stradley S. J., Gierasch L. M., Brown M. S., Goldstein J. L. Sequence requirement for peptide recognition by rat brain p21ras protein farnesyltransferase. Proc Natl Acad Sci U S A. 1991 Feb 1;88(3):732–736. doi: 10.1073/pnas.88.3.732. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Schafer W. R., Kim R., Sterne R., Thorner J., Kim S. H., Rine J. Genetic and pharmacological suppression of oncogenic mutations in ras genes of yeast and humans. Science. 1989 Jul 28;245(4916):379–385. doi: 10.1126/science.2569235. [DOI] [PubMed] [Google Scholar]
  54. Schafer W. R., Rine J. Protein prenylation: genes, enzymes, targets, and functions. Annu Rev Genet. 1992;26:209–237. doi: 10.1146/annurev.ge.26.120192.001233. [DOI] [PubMed] [Google Scholar]
  55. Senior A., Gerace L. Integral membrane proteins specific to the inner nuclear membrane and associated with the nuclear lamina. J Cell Biol. 1988 Dec;107(6 Pt 1):2029–2036. doi: 10.1083/jcb.107.6.2029. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Sheehan M. A., Mills A. D., Sleeman A. M., Laskey R. A., Blow J. J. Steps in the assembly of replication-competent nuclei in a cell-free system from Xenopus eggs. J Cell Biol. 1988 Jan;106(1):1–12. doi: 10.1083/jcb.106.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Simos G., Georgatos S. D. The inner nuclear membrane protein p58 associates in vivo with a p58 kinase and the nuclear lamins. EMBO J. 1992 Nov;11(11):4027–4036. doi: 10.1002/j.1460-2075.1992.tb05496.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Smith S., Blobel G. The first membrane spanning region of the lamin B receptor is sufficient for sorting to the inner nuclear membrane. J Cell Biol. 1993 Feb;120(3):631–637. doi: 10.1083/jcb.120.3.631. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Stick R., Angres B., Lehner C. F., Nigg E. A. The fates of chicken nuclear lamin proteins during mitosis: evidence for a reversible redistribution of lamin B2 between inner nuclear membrane and elements of the endoplasmic reticulum. J Cell Biol. 1988 Aug;107(2):397–406. doi: 10.1083/jcb.107.2.397. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Stick R., Hausen P. Changes in the nuclear lamina composition during early development of Xenopus laevis. Cell. 1985 May;41(1):191–200. doi: 10.1016/0092-8674(85)90073-x. [DOI] [PubMed] [Google Scholar]
  61. Stick R., Krohne G. Immunological localization of the major architectural protein associated with the nuclear envelope of the Xenopus laevis oocyte. Exp Cell Res. 1982 Apr;138(2):319–313. doi: 10.1016/0014-4827(82)90181-1. [DOI] [PubMed] [Google Scholar]
  62. Stick R. The gene structure of Xenopus nuclear lamin A: a model for the evolution of A-type from B-type lamins by exon shuffling. Chromosoma. 1992 Aug;101(9):566–574. doi: 10.1007/BF00660316. [DOI] [PubMed] [Google Scholar]
  63. Stick R. cDNA cloning of the developmentally regulated lamin LIII of Xenopus laevis. EMBO J. 1988 Oct;7(10):3189–3197. doi: 10.1002/j.1460-2075.1988.tb03186.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  64. Vigers G. P., Lohka M. J. A distinct vesicle population targets membranes and pore complexes to the nuclear envelope in Xenopus eggs. J Cell Biol. 1991 Feb;112(4):545–556. doi: 10.1083/jcb.112.4.545. [DOI] [PMC free article] [PubMed] [Google Scholar]
  65. Vigers G. P., Lohka M. J. Regulation of nuclear envelope precursor functions during cell division. J Cell Sci. 1992 Jun;102(Pt 2):273–284. doi: 10.1242/jcs.102.2.273. [DOI] [PubMed] [Google Scholar]
  66. Vorburger K., Kitten G. T., Nigg E. A. Modification of nuclear lamin proteins by a mevalonic acid derivative occurs in reticulocyte lysates and requires the cysteine residue of the C-terminal CXXM motif. EMBO J. 1989 Dec 20;8(13):4007–4013. doi: 10.1002/j.1460-2075.1989.tb08583.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  67. Vorburger K., Lehner C. F., Kitten G. T., Eppenberger H. M., Nigg E. A. A second higher vertebrate B-type lamin. cDNA sequence determination and in vitro processing of chicken lamin B2. J Mol Biol. 1989 Aug 5;208(3):405–415. doi: 10.1016/0022-2836(89)90505-6. [DOI] [PubMed] [Google Scholar]
  68. Ward G. E., Kirschner M. W. Identification of cell cycle-regulated phosphorylation sites on nuclear lamin C. Cell. 1990 May 18;61(4):561–577. doi: 10.1016/0092-8674(90)90469-u. [DOI] [PubMed] [Google Scholar]
  69. Weber K., Plessmann U., Traub P. Maturation of nuclear lamin A involves a specific carboxy-terminal trimming, which removes the polyisoprenylation site from the precursor; implications for the structure of the nuclear lamina. FEBS Lett. 1989 Nov 6;257(2):411–414. doi: 10.1016/0014-5793(89)81584-4. [DOI] [PubMed] [Google Scholar]
  70. Wilson K. L., Newport J. A trypsin-sensitive receptor on membrane vesicles is required for nuclear envelope formation in vitro. J Cell Biol. 1988 Jul;107(1):57–68. doi: 10.1083/jcb.107.1.57. [DOI] [PMC free article] [PubMed] [Google Scholar]
  71. Wolda S. L., Glomset J. A. Evidence for modification of lamin B by a product of mevalonic acid. J Biol Chem. 1988 May 5;263(13):5997–6000. [PubMed] [Google Scholar]
  72. Worman H. J., Evans C. D., Blobel G. The lamin B receptor of the nuclear envelope inner membrane: a polytopic protein with eight potential transmembrane domains. J Cell Biol. 1990 Oct;111(4):1535–1542. doi: 10.1083/jcb.111.4.1535. [DOI] [PMC free article] [PubMed] [Google Scholar]
  73. Worman H. J., Yuan J., Blobel G., Georgatos S. D. A lamin B receptor in the nuclear envelope. Proc Natl Acad Sci U S A. 1988 Nov;85(22):8531–8534. doi: 10.1073/pnas.85.22.8531. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES