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. 1993 Nov 1;123(3):501–512. doi: 10.1083/jcb.123.3.501

Membrane-associated lamins in Xenopus egg extracts: identification of two vesicle populations

PMCID: PMC2200128  PMID: 8227121

Abstract

Nuclear lamin isoforms of vertebrates can be divided into two major classes. The B-type lamins are membrane associated throughout the cell cycle, whereas A-type lamins are recovered from mitotic cell homogenates in membrane-free fractions. A feature of oogenesis in birds and mammals is the nearly exclusive presence of B-type lamins in oocyte nuclear envelopes. In contrast, oocytes and early cleavage embryos of the amphibian Xenopus laevis are believed to contain a single lamin isoform, lamin LIII, which after nuclear envelope breakdown during meiotic maturation is reported to be completely soluble. Consequently, we have reexamined the lamin complement of Xenopus oocyte nuclear envelopes, egg extracts, and early embryos. An mAb (X223) specific for the homologous B-type lamins B2 of mouse and LII of Xenopus somatic cells (Hoger, T., K. Zatloukal, I. Waizenegger, and G. Krohne. 1990. Chromosoma. 99:379-390) recognized a Xenopus oocyte nuclear envelope protein biochemically distinct from lamin LIII and very similar or identical to somatic cell lamin LII. Oocyte lamin LII was detectable in nuclear envelopes of early cleavage embryos. Immunoblotting of fractionated egg extracts revealed that approximately 20-23% of lamin LII and 5-7% of lamin LIII were membrane associated. EM immunolocalization demonstrated that membrane-bound lamins LII and LIII are associated with separate vesicle populations. These findings are relevant to the interpretation of nuclear reconstitution experiments using Xenopus egg extracts.

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Selected References

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

  1. 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]
  2. Beams H. W., Mueller S. Effects of ultracentrifugation on the interphase nucleus of somatic cells with special reference to the nuclear envelope-chromatin relationship. Z Zellforsch Mikrosk Anat. 1970;108(3):297–308. doi: 10.1007/BF00336521. [DOI] [PubMed] [Google Scholar]
  3. 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]
  4. 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]
  5. 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]
  6. 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]
  7. Cordes V., Waizenegger I., Krohne G. Nuclear pore complex glycoprotein p62 of Xenopus laevis and mouse: cDNA cloning and identification of its glycosylated region. Eur J Cell Biol. 1991 Jun;55(1):31–47. [PubMed] [Google Scholar]
  8. Dabauvalle M. C., Loos K., Merkert H., Scheer U. Spontaneous assembly of pore complex-containing membranes ("annulate lamellae") in Xenopus egg extract in the absence of chromatin. J Cell Biol. 1991 Mar;112(6):1073–1082. doi: 10.1083/jcb.112.6.1073. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dabauvalle M. C., Loos K., Scheer U. Identification of a soluble precursor complex essential for nuclear pore assembly in vitro. Chromosoma. 1990 Dec;100(1):56–66. doi: 10.1007/BF00337603. [DOI] [PubMed] [Google Scholar]
  10. Dabauvalle M. C., Scheer U. Assembly of nuclear pore complexes in Xenopus egg extract. Biol Cell. 1991;72(1-2):25–29. doi: 10.1016/0248-4900(91)90074-w. [DOI] [PubMed] [Google Scholar]
  11. 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]
  12. Furukawa K., Hotta Y. cDNA cloning of a germ cell specific lamin B3 from mouse spermatocytes and analysis of its function by ectopic expression in somatic cells. EMBO J. 1993 Jan;12(1):97–106. doi: 10.1002/j.1460-2075.1993.tb05635.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. 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]
  14. 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]
  15. 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]
  16. Hutchison C. J., Cox R., Ford C. C. The control of DNA replication in a cell-free extract that recapitulates a basic cell cycle in vitro. Development. 1988 Jul;103(3):553–566. doi: 10.1242/dev.103.3.553. [DOI] [PubMed] [Google Scholar]
  17. Höger T. H., Grund C., Franke W. W., Krohne G. Immunolocalization of lamins in the thick nuclear lamina of human synovial cells. Eur J Cell Biol. 1991 Feb;54(1):150–156. [PubMed] [Google Scholar]
  18. 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]
  19. Krohne G., Benavente R. The nuclear lamins. A multigene family of proteins in evolution and differentiation. Exp Cell Res. 1986 Jan;162(1):1–10. doi: 10.1016/0014-4827(86)90421-0. [DOI] [PubMed] [Google Scholar]
  20. 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]
  21. Krohne G., Wolin S. L., McKeon F. D., Franke W. W., Kirschner M. W. Nuclear lamin LI of Xenopus laevis: cDNA cloning, amino acid sequence and binding specificity of a member of the lamin B subfamily. EMBO J. 1987 Dec 1;6(12):3801–3808. doi: 10.1002/j.1460-2075.1987.tb02716.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kyhse-Andersen J. Electroblotting of multiple gels: a simple apparatus without buffer tank for rapid transfer of proteins from polyacrylamide to nitrocellulose. J Biochem Biophys Methods. 1984 Dec;10(3-4):203–209. doi: 10.1016/0165-022x(84)90040-x. [DOI] [PubMed] [Google Scholar]
  23. 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]
  24. Laskey R. A., Leno G. H. Assembly of the cell nucleus. Trends Genet. 1990 Dec;6(12):406–410. doi: 10.1016/0168-9525(90)90301-l. [DOI] [PubMed] [Google Scholar]
  25. Lebel S., Lampron C., Royal A., Raymond Y. Lamins A and C appear during retinoic acid-induced differentiation of mouse embryonal carcinoma cells. J Cell Biol. 1987 Sep;105(3):1099–1104. doi: 10.1083/jcb.105.3.1099. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Lebkowski J. S., Laemmli U. K. Non-histone proteins and long-range organization of HeLa interphase DNA. J Mol Biol. 1982 Apr 5;156(2):325–344. doi: 10.1016/0022-2836(82)90332-1. [DOI] [PubMed] [Google Scholar]
  27. Lehner C. F., Stick R., Eppenberger H. M., Nigg E. A. Differential expression of nuclear lamin proteins during chicken development. J Cell Biol. 1987 Jul;105(1):577–587. doi: 10.1083/jcb.105.1.577. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Lourim D., Lin J. J. Expression of nuclear lamin A and muscle-specific proteins in differentiating muscle cells in ovo and in vitro. J Cell Biol. 1989 Aug;109(2):495–504. doi: 10.1083/jcb.109.2.495. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Lourim D., Lin J. J. Expression of wild-type and nuclear localization-deficient human lamin A in chick myogenic cells. J Cell Sci. 1992 Nov;103(Pt 3):863–874. doi: 10.1242/jcs.103.3.863. [DOI] [PubMed] [Google Scholar]
  30. 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]
  31. 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]
  32. 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]
  33. 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]
  34. Newport J., Dunphy W. Characterization of the membrane binding and fusion events during nuclear envelope assembly using purified components. J Cell Biol. 1992 Jan;116(2):295–306. doi: 10.1083/jcb.116.2.295. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. 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]
  36. O'Farrell P. Z., Goodman H. M., O'Farrell P. H. High resolution two-dimensional electrophoresis of basic as well as acidic proteins. Cell. 1977 Dec;12(4):1133–1141. doi: 10.1016/0092-8674(77)90176-3. [DOI] [PubMed] [Google Scholar]
  37. 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]
  38. Röber R. A., Weber K., Osborn M. Differential timing of nuclear lamin A/C expression in the various organs of the mouse embryo and the young animal: a developmental study. Development. 1989 Feb;105(2):365–378. doi: 10.1242/dev.105.2.365. [DOI] [PubMed] [Google Scholar]
  39. Schatten G., Maul G. G., Schatten H., Chaly N., Simerly C., Balczon R., Brown D. L. Nuclear lamins and peripheral nuclear antigens during fertilization and embryogenesis in mice and sea urchins. Proc Natl Acad Sci U S A. 1985 Jul;82(14):4727–4731. doi: 10.1073/pnas.82.14.4727. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Stewart C., Burke B. Teratocarcinoma stem cells and early mouse embryos contain only a single major lamin polypeptide closely resembling lamin B. Cell. 1987 Nov 6;51(3):383–392. doi: 10.1016/0092-8674(87)90634-9. [DOI] [PubMed] [Google Scholar]
  41. 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]
  42. 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]
  43. 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]
  44. 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]
  45. Ulitzur N., Harel A., Feinstein N., Gruenbaum Y. Lamin activity is essential for nuclear envelope assembly in a Drosophila embryo cell-free extract. J Cell Biol. 1992 Oct;119(1):17–25. doi: 10.1083/jcb.119.1.17. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. 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]
  47. 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]
  48. 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]
  49. 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]
  50. 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]
  51. Wolin S. L., Krohne G., Kirschner M. W. A new lamin in Xenopus somatic tissues displays strong homology to human lamin A. EMBO J. 1987 Dec 1;6(12):3809–3818. doi: 10.1002/j.1460-2075.1987.tb02717.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Zewe M., Höger T. H., Fink T., Lichter P., Krohne G., Franke W. W. Gene structure and chromosomal localization of the murine lamin B2 gene. Eur J Cell Biol. 1991 Dec;56(2):342–350. [PubMed] [Google Scholar]

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