Abstract
We have examined the nuclear localization of isoprenylated proteins in CHO-K1 cells labeled with [14C]mevalonate. Nuclear proteins of 68, 70, and 74 kD, posttranslationally modified by an isoprenoid, are also components of a nuclear matrix-intermediate filament preparation from CHO cells. Furthermore, the 68-, 70-, and 74-kD isoprenylated polypeptides are immunoprecipitated from cell extracts with two different anti-lamin antisera. Based on exact two-dimensional comigration with lamin B, both from rat liver lamin and CHO nuclear matrix-intermediate filament preparations, and its immunoprecipitation with anti-lamin antisera, we conclude that the 68-kD isoprenylated protein found in nuclei from [14C]mevalonate-labeled CHO cells is lamin B. The more basic 74-kD isoprenylated nuclear protein is similar in molecular mass and isoelectric pH variants to the lamin A precursor polypeptide reported by others. Starving cells for mevalonate results in a dramatic accumulation of a polypeptide that comigrates on two- dimensional, non-equilibrium pH gradient electrophoresis (NEPHGE) gels with the 74-kD isoprenylated protein. The 70-kD isoprenylated protein, which is resolved on NEPHGE gels as being higher in molecular mass and slightly more basic than lamin B, has not yet been identified.
Full Text
The Full Text of this article is available as a PDF (2.6 MB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Aaronson R. P., Blobel G. Isolation of nuclear pore complexes in association with a lamina. Proc Natl Acad Sci U S A. 1975 Mar;72(3):1007–1011. doi: 10.1073/pnas.72.3.1007. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Alberts A. W., Chen J., Kuron G., Hunt V., Huff J., Hoffman C., Rothrock J., Lopez M., Joshua H., Harris E. Mevinolin: a highly potent competitive inhibitor of hydroxymethylglutaryl-coenzyme A reductase and a cholesterol-lowering agent. Proc Natl Acad Sci U S A. 1980 Jul;77(7):3957–3961. doi: 10.1073/pnas.77.7.3957. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Blobel G., Potter V. R. Nuclei from rat liver: isolation method that combines purity with high yield. Science. 1966 Dec 30;154(3757):1662–1665. doi: 10.1126/science.154.3757.1662. [DOI] [PubMed] [Google Scholar]
- Bruenger E., Rilling H. C. Prenylated proteins from kidney. Biochem Biophys Res Commun. 1986 Aug 29;139(1):209–214. doi: 10.1016/s0006-291x(86)80100-0. [DOI] [PubMed] [Google Scholar]
- 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]
- Cabral F., Gottesman M. M. The determination of similarities in amino acid composition among proteins separated by two-dimensional gel electrophoresis. Anal Biochem. 1978 Dec;91(2):548–556. doi: 10.1016/0003-2697(78)90542-0. [DOI] [PubMed] [Google Scholar]
- Chua N. H., Bennoun P. Thylakoid membrane polypeptides of Chlamydomonas reinhardtii: wild-type and mutant strains deficient in photosystem II reaction center. Proc Natl Acad Sci U S A. 1975 Jun;72(6):2175–2179. doi: 10.1073/pnas.72.6.2175. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cleveland D. W., Fischer S. G., Kirschner M. W., Laemmli U. K. Peptide mapping by limited proteolysis in sodium dodecyl sulfate and analysis by gel electrophoresis. J Biol Chem. 1977 Feb 10;252(3):1102–1106. [PubMed] [Google Scholar]
- DULBECCO R., VOGT M. Plaque formation and isolation of pure lines with poliomyelitis viruses. J Exp Med. 1954 Feb;99(2):167–182. doi: 10.1084/jem.99.2.167. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dagenais A., Bibor-Hardy V., Simard R. Characterization of lamin proteins in BHK cells. Exp Cell Res. 1984 Dec;155(2):435–447. doi: 10.1016/0014-4827(84)90204-0. [DOI] [PubMed] [Google Scholar]
- Dwyer N., Blobel G. A modified procedure for the isolation of a pore complex-lamina fraction from rat liver nuclei. J Cell Biol. 1976 Sep;70(3):581–591. doi: 10.1083/jcb.70.3.581. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Faust J., Krieger M. Expression of specific high capacity mevalonate transport in a Chinese hamster cell variant. J Biol Chem. 1987 Feb 15;262(5):1996–2004. [PubMed] [Google Scholar]
- Gasser S. M., Laemmli U. K. The organisation of chromatin loops: characterization of a scaffold attachment site. EMBO J. 1986 Mar;5(3):511–518. doi: 10.1002/j.1460-2075.1986.tb04240.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gerace L., Blobel G. Nuclear lamina and the structural organization of the nuclear envelope. Cold Spring Harb Symp Quant Biol. 1982;46(Pt 2):967–978. doi: 10.1101/sqb.1982.046.01.090. [DOI] [PubMed] [Google Scholar]
- 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]
- 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]
- 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]
- Habenicht A. J., Glomset J. A., Ross R. Relation of cholesterol and mevalonic acid to the cell cycle in smooth muscle and swiss 3T3 cells stimulated to divide by platelet-derived growth factor. J Biol Chem. 1980 Jun 10;255(11):5134–5140. [PubMed] [Google Scholar]
- Kaufmann S. H., Gibson W., Shaper J. H. Characterization of the major polypeptides of the rat liver nuclear envelope. J Biol Chem. 1983 Feb 25;258(4):2710–2719. [PubMed] [Google Scholar]
- 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]
- 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]
- Krohne G., Franke W. W. Proteins of pore complex--lamina structures from nuclei and nuclear membranes. Methods Enzymol. 1983;96:597–608. doi: 10.1016/s0076-6879(83)96052-4. [DOI] [PubMed] [Google Scholar]
- Krohne G., Franke W. W., Scheer U. The major polypeptides of the nuclear pore complex. Exp Cell Res. 1978 Oct 1;116(1):85–102. doi: 10.1016/0014-4827(78)90067-8. [DOI] [PubMed] [Google Scholar]
- 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]
- 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]
- Laskey R. A., Mills A. D. Quantitative film detection of 3H and 14C in polyacrylamide gels by fluorography. Eur J Biochem. 1975 Aug 15;56(2):335–341. doi: 10.1111/j.1432-1033.1975.tb02238.x. [DOI] [PubMed] [Google Scholar]
- Lebel S., Raymond Y. Lamin B from rat liver nuclei exists both as a lamina protein and as an intrinsic membrane protein. J Biol Chem. 1984 Mar 10;259(5):2693–2696. [PubMed] [Google Scholar]
- Lebel S., Raymond Y. Lamins A, B and C share an epitope with the common domain of intermediate filament proteins. Exp Cell Res. 1987 Apr;169(2):560–565. doi: 10.1016/0014-4827(87)90216-3. [DOI] [PubMed] [Google Scholar]
- 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]
- Lehner C. F., Kurer V., Eppenberger H. M., Nigg E. A. The nuclear lamin protein family in higher vertebrates. Identification of quantitatively minor lamin proteins by monoclonal antibodies. J Biol Chem. 1986 Oct 5;261(28):13293–13301. [PubMed] [Google Scholar]
- Maltese W. A., Sheridan K. M. Isoprenylated proteins in cultured cells: subcellular distribution and changes related to altered morphology and growth arrest induced by mevalonate deprivation. J Cell Physiol. 1987 Dec;133(3):471–481. doi: 10.1002/jcp.1041330307. [DOI] [PubMed] [Google Scholar]
- Maul G. G., Baglia F. A., Newmeyer D. D., Ohlsson-Wilhelm B. M. The major 67 000 molecular weight protein of the clam oocyte nuclear envelope is lamin-like. J Cell Sci. 1984 Apr;67:69–85. doi: 10.1242/jcs.67.1.69. [DOI] [PubMed] [Google Scholar]
- McKeon F. D., Tuffanelli D. L., Fukuyama K., Kirschner M. W. Autoimmune response directed against conserved determinants of nuclear envelope proteins in a patient with linear scleroderma. Proc Natl Acad Sci U S A. 1983 Jul;80(14):4374–4378. doi: 10.1073/pnas.80.14.4374. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Miyakawa T., Tabata M., Tsuchiya E., Fukui S. Biosynthesis and secretion of tremerogen A-10, a polyisoprenyl peptide mating pheromone of Tremella mesenterica. Eur J Biochem. 1985 Mar 15;147(3):489–493. doi: 10.1111/j.0014-2956.1985.00489.x. [DOI] [PubMed] [Google Scholar]
- Newport J. W., Forbes D. J. The nucleus: structure, function, and dynamics. Annu Rev Biochem. 1987;56:535–565. doi: 10.1146/annurev.bi.56.070187.002535. [DOI] [PubMed] [Google Scholar]
- 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]
- Ottaviano Y., Gerace L. Phosphorylation of the nuclear lamins during interphase and mitosis. J Biol Chem. 1985 Jan 10;260(1):624–632. [PubMed] [Google Scholar]
- Peffley D., Sinensky M. Regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase synthesis by a non-sterol mevalonate-derived product in Mev-1 cells. Apparent translational control. J Biol Chem. 1985 Aug 25;260(18):9949–9952. [PubMed] [Google Scholar]
- Quesney-Huneeus V., Wiley M. H., Siperstein M. D. Essential role for mevalonate synthesis in DNA replication. Proc Natl Acad Sci U S A. 1979 Oct;76(10):5056–5060. doi: 10.1073/pnas.76.10.5056. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Scheer U., Kartenbeck J., Trendelenburg M. F., Stadler J., Franke W. W. Experimental disintegration of the nuclear envelope. Evidence for pore-connecting fibrils. J Cell Biol. 1976 Apr;69(1):1–18. doi: 10.1083/jcb.69.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schmidt R. A., Schneider C. J., Glomset J. A. Evidence for post-translational incorporation of a product of mevalonic acid into Swiss 3T3 cell proteins. J Biol Chem. 1984 Aug 25;259(16):10175–10180. [PubMed] [Google Scholar]
- Sefton B. M., Buss J. E. The covalent modification of eukaryotic proteins with lipid. J Cell Biol. 1987 Jun;104(6):1449–1453. doi: 10.1083/jcb.104.6.1449. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Shelton K. R., Egle P. M., Cochran D. L. Nuclear envelope proteins: identification of lamin B subtypes. Biochem Biophys Res Commun. 1981 Dec 15;103(3):975–981. doi: 10.1016/0006-291x(81)90905-0. [DOI] [PubMed] [Google Scholar]
- Shelton K. R., Higgins L. L., Cochran D. L., Ruffolo J. J., Jr, Egle P. M. Nuclear lamins of erythrocyte and liver. J Biol Chem. 1980 Nov 25;255(22):10978–10983. [PubMed] [Google Scholar]
- Sinensky M., Logel J. Defective macromolecule biosynthesis and cell-cycle progression in a mammalian cell starved for mevalonate. Proc Natl Acad Sci U S A. 1985 May;82(10):3257–3261. doi: 10.1073/pnas.82.10.3257. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sinensky M., Torget R., Edwards P. A. Radioimmune precipitation of 3-hydroxy-3-methylglutaryl coenzyme A reductase from Chinese hamster fibroblasts. Effect of 25-hydroxycholesterol. J Biol Chem. 1981 Nov 25;256(22):11774–11779. [PubMed] [Google Scholar]
- Skinner M. K., Griswold M. D. Fluorographic detection of radioactivity in polyacrylamide gels with 2,5-diphenyloxazole in acetic acid and its comparison with existing procedures. Biochem J. 1983 Jan 1;209(1):281–284. doi: 10.1042/bj2090281. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Smith D. E., Gruenbaum Y., Berrios M., Fisher P. A. Biosynthesis and interconversion of Drosophila nuclear lamin isoforms during normal growth and in response to heat shock. J Cell Biol. 1987 Aug;105(2):771–790. doi: 10.1083/jcb.105.2.771. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Suprynowicz F. A., Gerace L. A fractionated cell-free system for analysis of prophase nuclear disassembly. J Cell Biol. 1986 Dec;103(6 Pt 1):2073–2081. doi: 10.1083/jcb.103.6.2073. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Traub P., Perides G., Kühn S., Scherbarth A. Efficient interaction of nonpolar lipids with intermediate filaments of the vimentin type. Eur J Cell Biol. 1987 Feb;43(1):55–64. [PubMed] [Google Scholar]
- Wilcox C. A., Olson E. N. The majority of cellular fatty acid acylated proteins are localized to the cytoplasmic surface of the plasma membrane. Biochemistry. 1987 Feb 24;26(4):1029–1036. doi: 10.1021/bi00378a008. [DOI] [PubMed] [Google Scholar]