Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1991 Jan 1;112(1):1–11. doi: 10.1083/jcb.112.1.1

Different forms of soluble cytoplasmic mRNA binding proteins and particles in Xenopus laevis oocytes and embryos

PMCID: PMC2288798  PMID: 1670777

Abstract

To gain insight into the mechanisms involved in the formation of maternally stored mRNPs during Xenopus laevis development, we searched for soluble cytoplasmic proteins of the oocyte that are able to selectively bind mRNAs, using as substrate radiolabeled mRNA. In vitro mRNP assembly in solution was followed by UV-cross-linking and RNase digestion, resulting in covalent tagging of polypeptides by nucleotide transfer. Five polypeptides of approximately 54, 56 60, 70, and 100 kD (p54, p56, p60, p70, and p100) have been found to selectively bind mRNA and assemble into mRNPs. These polypeptides, which correspond to previously described native mRNP components, occur in three different particle classes of approximately 4.5S, approximately 6S, and approximately 15S, as also determined by their reactions with antibodies against p54 and p56. Whereas the approximately 4.5S class contains p42, p60, and p70, probably each in the form of individual molecules or small complexes, the approximately 6S particles appears to consist only of p54 and p56, which occur in a near-stoichiometric ratio suggestive of a heterodimer complex. The approximately 15S particles contain, in addition to p54 and p56, p60 and p100 and this is the single occurring form of RNA-binding p100. We have also observed changes in the in vitro mRNA binding properties of these polypeptides during oogenesis and early embryonic development, in relation to their phosphorylation state and to the activity of an approximately 15S particle-associated protein kinase, suggesting that these proteins are involved in the developmental translational regulation of maternal mRNAs.

Full Text

The Full Text of this article is available as a PDF (2.8 MB).

Selected References

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

  1. Anderson D. M., Richter J. D., Chamberlin M. E., Price D. H., Britten R. J., Smith L. D., Davidson E. H. Sequence organization of the poly(A) RNA synthesized and accumulated in lampbrush chromosome stage Xenopus laevis oocytes. J Mol Biol. 1982 Mar 5;155(3):281–309. doi: 10.1016/0022-2836(82)90006-7. [DOI] [PubMed] [Google Scholar]
  2. Ballantine J. E., Woodland H. R., Sturgess E. A. Changes in protein synthesis during the development of Xenopus laevis. J Embryol Exp Morphol. 1979 Jun;51:137–153. [PubMed] [Google Scholar]
  3. Bandziulis R. J., Swanson M. S., Dreyfuss G. RNA-binding proteins as developmental regulators. Genes Dev. 1989 Apr;3(4):431–437. doi: 10.1101/gad.3.4.431. [DOI] [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. Beyer A. L., Christensen M. E., Walker B. W., LeStourgeon W. M. Identification and characterization of the packaging proteins of core 40S hnRNP particles. Cell. 1977 May;11(1):127–138. doi: 10.1016/0092-8674(77)90323-3. [DOI] [PubMed] [Google Scholar]
  7. Blobel G. A protein of molecular weight 78,000 bound to the polyadenylate region of eukaryotic messenger RNAs. Proc Natl Acad Sci U S A. 1973 Mar;70(3):924–928. doi: 10.1073/pnas.70.3.924. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Brawerman G. mRNA decay: finding the right targets. Cell. 1989 Apr 7;57(1):9–10. doi: 10.1016/0092-8674(89)90166-9. [DOI] [PubMed] [Google Scholar]
  9. Crawford D. R., Richter J. D. An RNA-binding protein from Xenopus oocytes is associated with specific message sequences. Development. 1987 Dec;101(4):741–749. doi: 10.1242/dev.101.4.741. [DOI] [PubMed] [Google Scholar]
  10. Cummings A., Sommerville J. Protein kinase activity associated with stored messenger ribonucleoprotein particles of Xenopus oocytes. J Cell Biol. 1988 Jul;107(1):45–56. doi: 10.1083/jcb.107.1.45. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Darnbrough C. H., Ford P. J. Identification in Xenopus laevis of a class of oocyte-specific proteins bound to messenger RNA. Eur J Biochem. 1981 Jan;113(3):415–424. doi: 10.1111/j.1432-1033.1981.tb05081.x. [DOI] [PubMed] [Google Scholar]
  12. Dearsly A. L., Johnson R. M., Barrett P., Sommerville J. Identification of a 60-kDa phosphoprotein that binds stored messenger RNA of Xenopus oocytes. Eur J Biochem. 1985 Jul 1;150(1):95–103. doi: 10.1111/j.1432-1033.1985.tb08993.x. [DOI] [PubMed] [Google Scholar]
  13. Dignam J. D., Lebovitz R. M., Roeder R. G. Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei. Nucleic Acids Res. 1983 Mar 11;11(5):1475–1489. doi: 10.1093/nar/11.5.1475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dingwall C., Dilworth S. M., Black S. J., Kearsey S. E., Cox L. S., Laskey R. A. Nucleoplasmin cDNA sequence reveals polyglutamic acid tracts and a cluster of sequences homologous to putative nuclear localization signals. EMBO J. 1987 Jan;6(1):69–74. doi: 10.1002/j.1460-2075.1987.tb04720.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Dolecki G. J., Smith L. D. Poly(A)+ RNA metabolism during oogenesis in Xenopus laevis. Dev Biol. 1979 Mar;69(1):217–236. doi: 10.1016/0012-1606(79)90287-2. [DOI] [PubMed] [Google Scholar]
  16. Drawbridge J., Grainger J. L., Winkler M. M. Identification and characterization of the poly(A)-binding proteins from the sea urchin: a quantitative analysis. Mol Cell Biol. 1990 Aug;10(8):3994–4006. doi: 10.1128/mcb.10.8.3994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Dreyfuss G. Structure and function of nuclear and cytoplasmic ribonucleoprotein particles. Annu Rev Cell Biol. 1986;2:459–498. doi: 10.1146/annurev.cb.02.110186.002331. [DOI] [PubMed] [Google Scholar]
  18. 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]
  19. Duval C., Bouvet P., Omilli F., Roghi C., Dorel C., LeGuellec R., Paris J., Osborne H. B. Stability of maternal mRNA in Xenopus embryos: role of transcription and translation. Mol Cell Biol. 1990 Aug;10(8):4123–4129. doi: 10.1128/mcb.10.8.4123. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Erikson E., Maller J. L. Purification and characterization of a protein kinase from Xenopus eggs highly specific for ribosomal protein S6. J Biol Chem. 1986 Jan 5;261(1):350–355. [PubMed] [Google Scholar]
  21. Greenberg J. R., Carroll E., 3rd Reconstitution of functional mRNA-protein complexes in a rabbit reticulocyte cell-free translation system. Mol Cell Biol. 1985 Feb;5(2):342–351. doi: 10.1128/mcb.5.2.342. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Greenberg J. R. Proteins crosslinked to messenger RNA by irradiating polyribosomes with ultraviolet light. Nucleic Acids Res. 1980 Dec 11;8(23):5685–5701. doi: 10.1093/nar/8.23.5685. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Greenberg J. R. The polyribosomal mRNA--protein complex is a dynamic structure. Proc Natl Acad Sci U S A. 1981 May;78(5):2923–2926. doi: 10.1073/pnas.78.5.2923. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Hamm J., Kazmaier M., Mattaj I. W. In vitro assembly of U1 snRNPs. EMBO J. 1987 Nov;6(11):3479–3485. doi: 10.1002/j.1460-2075.1987.tb02672.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Hathaway G. M., Traugh J. A. Casein kinases--multipotential protein kinases. Curr Top Cell Regul. 1982;21:101–127. [PubMed] [Google Scholar]
  26. Hunt T. Controlling mRNA lifespan. Nature. 1988 Aug 18;334(6183):567–568. doi: 10.1038/334567a0. [DOI] [PubMed] [Google Scholar]
  27. Hyman L. E., Wormington W. M. Translational inactivation of ribosomal protein mRNAs during Xenopus oocyte maturation. Genes Dev. 1988 May;2(5):598–605. doi: 10.1101/gad.2.5.598. [DOI] [PubMed] [Google Scholar]
  28. Jackson R. J., Standart N. Do the poly(A) tail and 3' untranslated region control mRNA translation? Cell. 1990 Jul 13;62(1):15–24. doi: 10.1016/0092-8674(90)90235-7. [DOI] [PubMed] [Google Scholar]
  29. Kandror K. V., Benumov A. O., Stepanov A. S. Casein kinase II from Rana temporaria oocytes. Intracellular localization and activity during progesterone-induced maturation. Eur J Biochem. 1989 Mar 15;180(2):441–448. doi: 10.1111/j.1432-1033.1989.tb14666.x. [DOI] [PubMed] [Google Scholar]
  30. Kick D., Barrett P., Cummings A., Sommerville J. Phosphorylation of a 60 kDa polypeptide from Xenopus oocytes blocks messenger RNA translation. Nucleic Acids Res. 1987 May 26;15(10):4099–4109. doi: 10.1093/nar/15.10.4099. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Kleinschmidt J. A., Dingwall C., Maier G., Franke W. W. Molecular characterization of a karyophilic, histone-binding protein: cDNA cloning, amino acid sequence and expression of nuclear protein N1/N2 of Xenopus laevis. EMBO J. 1986 Dec 20;5(13):3547–3552. doi: 10.1002/j.1460-2075.1986.tb04681.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Kleinschmidt J. A., Fortkamp E., Krohne G., Zentgraf H., Franke W. W. Co-existence of two different types of soluble histone complexes in nuclei of Xenopus laevis oocytes. J Biol Chem. 1985 Jan 25;260(2):1166–1176. [PubMed] [Google Scholar]
  33. Kloetzel P. M., Johnson R. M., Sommerville J. Interaction of the hnRNA of amphibian oocytes with fibril-forming proteins. Eur J Biochem. 1982 Oct;127(2):301–308. doi: 10.1111/j.1432-1033.1982.tb06870.x. [DOI] [PubMed] [Google Scholar]
  34. Kozak M. A profusion of controls. J Cell Biol. 1988 Jul;107(1):1–7. doi: 10.1083/jcb.107.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. 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]
  36. 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]
  37. Laskey R. A., Honda B. M., Mills A. D., Finch J. T. Nucleosomes are assembled by an acidic protein which binds histones and transfers them to DNA. Nature. 1978 Oct 5;275(5679):416–420. doi: 10.1038/275416a0. [DOI] [PubMed] [Google Scholar]
  38. Lee G., Hynes R., Kirschner M. Temporal and spatial regulation of fibronectin in early Xenopus development. Cell. 1984 Mar;36(3):729–740. doi: 10.1016/0092-8674(84)90353-2. [DOI] [PubMed] [Google Scholar]
  39. Leibold E. A., Munro H. N. Cytoplasmic protein binds in vitro to a highly conserved sequence in the 5' untranslated region of ferritin heavy- and light-subunit mRNAs. Proc Natl Acad Sci U S A. 1988 Apr;85(7):2171–2175. doi: 10.1073/pnas.85.7.2171. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Lindberg U., Sundquist B. Isolation of messenger ribonucleoproteins from mammalian cells. J Mol Biol. 1974 Jun 25;86(2):451–468. doi: 10.1016/0022-2836(74)90030-8. [DOI] [PubMed] [Google Scholar]
  41. Mayrand S., Pederson T. Nuclear ribonucleoprotein particles probed in living cells. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2208–2212. doi: 10.1073/pnas.78.4.2208. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. McGrew L. L., Dworkin-Rastl E., Dworkin M. B., Richter J. D. Poly(A) elongation during Xenopus oocyte maturation is required for translational recruitment and is mediated by a short sequence element. Genes Dev. 1989 Jun;3(6):803–815. doi: 10.1101/gad.3.6.803. [DOI] [PubMed] [Google Scholar]
  43. Melton D. A., Krieg P. A., Rebagliati M. R., Maniatis T., Zinn K., Green M. R. Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter. Nucleic Acids Res. 1984 Sep 25;12(18):7035–7056. doi: 10.1093/nar/12.18.7035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Miyahara K., Shiokawa K., Yamana K. Cellular commitment for post-gastrular increase in alkaline phosphatase activity in Xenopus laevis development. Differentiation. 1982;21(1):45–49. doi: 10.1111/j.1432-0436.1982.tb01194.x. [DOI] [PubMed] [Google Scholar]
  45. Mulner-Lorillon O., Marot J., Cayla X., Pouhle R., Belle R. Purification and characterization of a casein-kinase-II-type enzyme from Xenopus laevis ovary. Biological effects on the meiotic cell division of full-grown oocyte. Eur J Biochem. 1988 Jan 15;171(1-2):107–117. doi: 10.1111/j.1432-1033.1988.tb13765.x. [DOI] [PubMed] [Google Scholar]
  46. O'Farrell P. H. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [PMC free article] [PubMed] [Google Scholar]
  47. 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]
  48. Paris J., Philippe M. Poly(A) metabolism and polysomal recruitment of maternal mRNAs during early Xenopus development. Dev Biol. 1990 Jul;140(1):221–224. doi: 10.1016/0012-1606(90)90070-y. [DOI] [PubMed] [Google Scholar]
  49. Peters J. M., Walsh M. J., Franke W. W. An abundant and ubiquitous homo-oligomeric ring-shaped ATPase particle related to the putative vesicle fusion proteins Sec18p and NSF. EMBO J. 1990 Jun;9(6):1757–1767. doi: 10.1002/j.1460-2075.1990.tb08300.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Richter J. D., Evers D. C. A monoclonal antibody to an oocyte-specific poly(A) RNA-binding protein. J Biol Chem. 1984 Feb 25;259(4):2190–2194. [PubMed] [Google Scholar]
  51. Richter J. D., Smith L. D., Anderson D. M., Davidson E. H. Interspersed poly(A) RNAs of amphibian oocytes are not translatable. J Mol Biol. 1984 Feb 25;173(2):227–241. doi: 10.1016/0022-2836(84)90191-8. [DOI] [PubMed] [Google Scholar]
  52. Richter J. D., Smith L. D. Developmentally regulated RNA binding proteins during oogenesis in Xenopus laevis. J Biol Chem. 1983 Apr 25;258(8):4864–4869. [PubMed] [Google Scholar]
  53. Richter J. D., Smith L. D. Reversible inhibition of translation by Xenopus oocyte-specific proteins. Nature. 1984 May 24;309(5966):378–380. doi: 10.1038/309378a0. [DOI] [PubMed] [Google Scholar]
  54. Ruiz i Altaba A., Perry-O'Keefe H., Melton D. A. Xfin: an embryonic gene encoding a multifingered protein in Xenopus. EMBO J. 1987 Oct;6(10):3065–3070. doi: 10.1002/j.1460-2075.1987.tb02613.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Sachs A. B., Bond M. W., Kornberg R. D. A single gene from yeast for both nuclear and cytoplasmic polyadenylate-binding proteins: domain structure and expression. Cell. 1986 Jun 20;45(6):827–835. doi: 10.1016/0092-8674(86)90557-x. [DOI] [PubMed] [Google Scholar]
  56. Schmidt-Zachmann M. S., Hügle-Dörr B., Franke W. W. A constitutive nucleolar protein identified as a member of the nucleoplasmin family. EMBO J. 1987 Jul;6(7):1881–1890. doi: 10.1002/j.1460-2075.1987.tb02447.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Shiokawa K., Misumi Y., Tashiro K., Nakakura N., Yamana K., Oh-uchida M. Changes in the patterns of RNA synthesis in early embryogenesis of Xenopus laevis. Cell Differ Dev. 1989 Oct;28(1):17–25. doi: 10.1016/0922-3371(89)90019-1. [DOI] [PubMed] [Google Scholar]
  58. Smith S., Stillman B. Purification and characterization of CAF-I, a human cell factor required for chromatin assembly during DNA replication in vitro. Cell. 1989 Jul 14;58(1):15–25. doi: 10.1016/0092-8674(89)90398-x. [DOI] [PubMed] [Google Scholar]
  59. Spirin A. S. "Masked" forms of mRNA. Curr Top Dev Biol. 1966;1:1–38. [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. Swiderski R. E., Richter J. D. Photocrosslinking of proteins to maternal mRNA in Xenopus oocytes. Dev Biol. 1988 Aug;128(2):349–358. doi: 10.1016/0012-1606(88)90297-7. [DOI] [PubMed] [Google Scholar]
  62. Ullrich S. J., Appella E., Mercer W. E. Growth-related expression of a 72,000 molecular weight poly(A)+ mRNA binding protein. Exp Cell Res. 1988 Oct;178(2):273–286. doi: 10.1016/0014-4827(88)90398-9. [DOI] [PubMed] [Google Scholar]
  63. 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]
  64. Woodland H. R. Changes in the polysome content of developing Xenopus laevis embryos. Dev Biol. 1974 Sep;40(1):90–101. doi: 10.1016/0012-1606(74)90111-0. [DOI] [PubMed] [Google Scholar]
  65. Woodland H. R., Flynn J. M., Wyllie A. J. Utilization of stored mRNA in Xenopus embryos and its replacement by newly synthesized transcripts: histone H1 synthesis using interspecies hybrids. Cell. 1979 Sep;18(1):165–171. doi: 10.1016/0092-8674(79)90365-9. [DOI] [PubMed] [Google Scholar]
  66. Zelus B. D., Giebelhaus D. H., Eib D. W., Kenner K. A., Moon R. T. Expression of the poly(A)-binding protein during development of Xenopus laevis. Mol Cell Biol. 1989 Jun;9(6):2756–2760. doi: 10.1128/mcb.9.6.2756. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES