Skip to main content
PMC home page
The EMBO Journal logoLink to The EMBO Journal
. 1995 Sep 1;14(17):4357–4364. doi: 10.1002/j.1460-2075.1995.tb00110.x

Identification of two KH domain proteins in the alpha-globin mRNP stability complex.

M Kiledjian 1, X Wang 1, S A Liebhaber 1
PMCID: PMC394520  PMID: 7556077

Abstract

Accumulation of globin mRNAs during erythroid differentiation is dependent on their extraordinary stability. The longevity of human alpha-globin mRNA is associated with a ribonucleoprotein complex (alpha-complex) formed on the 3' untranslated region (3'UTR). One or more of the proteins within this alpha-complex contain strong polycytosine [poly(C)] binding (alpha PCB) activity. In the present report we purify alpha PCB activity from human erythroid K562 cells. Although not able to bind the alpha-globin 3'UTR directly, alpha PCB activity is sufficient to complement alpha-complex formation in a cytosolic extract depleted of poly(C) binding activity. Peptide microsequencing demonstrates that alpha PCB activity contains two structurally related poly(C) binding proteins. These two proteins, alpha-complex protein (alpha CP)-1 and -2, have an overall structural identity of 80% and contain three repeats of the K homology (KH) domain which is found in a subset of RNA binding proteins. Epitope-tagged recombinant alpha CP-1 and alpha CP-2 expressed in cells are each incorporated into the alpha-complex. We conclude that alpha CP-1 and alpha CP-2, members of the KH domain RNA binding protein family, are involved in formation of a sequence-specific alpha-globin mRNP complex associated with alpha-globin mRNA stability. As such this represents the first example of a specific function for this class of proteins and suggests potential roles for other members of this protein family.

Full text

PDF
4357

Images in this article

4358Image
on p.4358
4358Image
on p.4358
4359Image
on p.4359
4360Image
on p.4360
4361Image
on p.4361
4361Image
on p.4361

Selected References

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

  1. Aasheim H. C., Loukianova T., Deggerdal A., Smeland E. B. Tissue specific expression and cDNA structure of a human transcript encoding a nucleic acid binding [oligo(dC)] protein related to the pre-mRNA binding protein K. Nucleic Acids Res. 1994 Mar 25;22(6):959–964. doi: 10.1093/nar/22.6.959. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  3. Ashley C. T., Jr, Wilkinson K. D., Reines D., Warren S. T. FMR1 protein: conserved RNP family domains and selective RNA binding. Science. 1993 Oct 22;262(5133):563–566. doi: 10.1126/science.7692601. [DOI] [PubMed] [Google Scholar]
  4. Atwater J. A., Wisdom R., Verma I. M. Regulated mRNA stability. Annu Rev Genet. 1990;24:519–541. doi: 10.1146/annurev.ge.24.120190.002511. [DOI] [PubMed] [Google Scholar]
  5. Burd C. G., Dreyfuss G. Conserved structures and diversity of functions of RNA-binding proteins. Science. 1994 Jul 29;265(5172):615–621. doi: 10.1126/science.8036511. [DOI] [PubMed] [Google Scholar]
  6. Castiglone Morelli M. A., Stier G., Gibson T., Joseph C., Musco G., Pastore A., Travè G. The KH module has an alpha beta fold. FEBS Lett. 1995 Jan 23;358(2):193–198. doi: 10.1016/0014-5793(94)01422-w. [DOI] [PubMed] [Google Scholar]
  7. Clegg J. B., Weatherall D. J., Milner P. F. Haemoglobin Constant Spring--a chain termination mutant? Nature. 1971 Dec 10;234(5328):337–340. doi: 10.1038/234337a0. [DOI] [PubMed] [Google Scholar]
  8. De Boulle K., Verkerk A. J., Reyniers E., Vits L., Hendrickx J., Van Roy B., Van den Bos F., de Graaff E., Oostra B. A., Willems P. J. A point mutation in the FMR-1 gene associated with fragile X mental retardation. Nat Genet. 1993 Jan;3(1):31–35. doi: 10.1038/ng0193-31. [DOI] [PubMed] [Google Scholar]
  9. Dreyfuss G., Matunis M. J., Piñol-Roma S., Burd C. G. hnRNP proteins and the biogenesis of mRNA. Annu Rev Biochem. 1993;62:289–321. doi: 10.1146/annurev.bi.62.070193.001445. [DOI] [PubMed] [Google Scholar]
  10. Evan G. I., Lewis G. K., Ramsay G., Bishop J. M. Isolation of monoclonal antibodies specific for human c-myc proto-oncogene product. Mol Cell Biol. 1985 Dec;5(12):3610–3616. doi: 10.1128/mcb.5.12.3610. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Furuichi Y., LaFiandra A., Shatkin A. J. 5'-Terminal structure and mRNA stability. Nature. 1977 Mar 17;266(5599):235–239. doi: 10.1038/266235a0. [DOI] [PubMed] [Google Scholar]
  12. Gibson T. J., Thompson J. D., Heringa J. The KH domain occurs in a diverse set of RNA-binding proteins that include the antiterminator NusA and is probably involved in binding to nucleic acid. FEBS Lett. 1993 Jun 21;324(3):361–366. doi: 10.1016/0014-5793(93)80152-k. [DOI] [PubMed] [Google Scholar]
  13. Goller M., Funke B., Gehe-Becker C., Kröger B., Lottspeich F., Horak I. Murine protein which binds preferentially to oligo-C-rich single-stranded nucleic acids. Nucleic Acids Res. 1994 May 25;22(10):1885–1889. doi: 10.1093/nar/22.10.1885. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hahm K., Kim G., Turck C. W., Smale S. T. Isolation of a murine gene encoding a nucleic acid-binding protein with homology to hnRNP K. Nucleic Acids Res. 1993 Aug 11;21(16):3894–3894. doi: 10.1093/nar/21.16.3894. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hunt D. M., Higgs D. R., Winichagoon P., Clegg J. B., Weatherall D. J. Haemoglobin Constant Spring has an unstable alpha chain messenger RNA. Br J Haematol. 1982 Jul;51(3):405–413. doi: 10.1111/j.1365-2141.1982.tb02796.x. [DOI] [PubMed] [Google Scholar]
  16. Jackson R. J. Cytoplasmic regulation of mRNA function: the importance of the 3' untranslated region. Cell. 1993 Jul 16;74(1):9–14. doi: 10.1016/0092-8674(93)90290-7. [DOI] [PubMed] [Google Scholar]
  17. Kenan D. J., Query C. C., Keene J. D. RNA recognition: towards identifying determinants of specificity. Trends Biochem Sci. 1991 Jun;16(6):214–220. doi: 10.1016/0968-0004(91)90088-d. [DOI] [PubMed] [Google Scholar]
  18. Kiledjian M., Dreyfuss G. Primary structure and binding activity of the hnRNP U protein: binding RNA through RGG box. EMBO J. 1992 Jul;11(7):2655–2664. doi: 10.1002/j.1460-2075.1992.tb05331.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Klausner R. D., Rouault T. A., Harford J. B. Regulating the fate of mRNA: the control of cellular iron metabolism. Cell. 1993 Jan 15;72(1):19–28. doi: 10.1016/0092-8674(93)90046-s. [DOI] [PubMed] [Google Scholar]
  20. Lamm G. M., Lamond A. I. Non-snRNP protein splicing factors. Biochim Biophys Acta. 1993 Jun 25;1173(3):247–265. doi: 10.1016/0167-4781(93)90122-t. [DOI] [PubMed] [Google Scholar]
  21. Liebhaber S. A. Alpha thalassemia. Hemoglobin. 1989;13(7-8):685–731. doi: 10.3109/03630268908998845. [DOI] [PubMed] [Google Scholar]
  22. Liebhaber S. A., Kan Y. W. Differentiation of the mRNA transcripts originating from the alpha 1- and alpha 2-globin loci in normals and alpha-thalassemics. J Clin Invest. 1981 Aug;68(2):439–446. doi: 10.1172/JCI110273. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Lodish H. F., Small B. Different lifetimes of reticulocyte messenger RNA. Cell. 1976 Jan;7(1):59–65. doi: 10.1016/0092-8674(76)90255-5. [DOI] [PubMed] [Google Scholar]
  24. Madhani H. D., Guthrie C. Dynamic RNA-RNA interactions in the spliceosome. Annu Rev Genet. 1994;28:1–26. doi: 10.1146/annurev.ge.28.120194.000245. [DOI] [PubMed] [Google Scholar]
  25. Mattaj I. W. RNA recognition: a family matter? Cell. 1993 Jun 4;73(5):837–840. doi: 10.1016/0092-8674(93)90265-r. [DOI] [PubMed] [Google Scholar]
  26. Matunis M. J., Michael W. M., Dreyfuss G. Characterization and primary structure of the poly(C)-binding heterogeneous nuclear ribonucleoprotein complex K protein. Mol Cell Biol. 1992 Jan;12(1):164–171. doi: 10.1128/mcb.12.1.164. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Milner P. F., Clegg J. B., Weatherall D. J. Haemoglobin-H disease due to a unique haemoglobin variant with an elongated alpha-chain. Lancet. 1971 Apr 10;1(7702):729–732. doi: 10.1016/s0140-6736(71)91992-1. [DOI] [PubMed] [Google Scholar]
  28. Nielsen D. A., Shapiro D. J. Insights into hormonal control of messenger RNA stability. Mol Endocrinol. 1990 Jul;4(7):953–957. doi: 10.1210/mend-4-7-953. [DOI] [PubMed] [Google Scholar]
  29. Nigg E. A., Baeuerle P. A., Lührmann R. Nuclear import-export: in search of signals and mechanisms. Cell. 1991 Jul 12;66(1):15–22. doi: 10.1016/0092-8674(91)90135-l. [DOI] [PubMed] [Google Scholar]
  30. Peltz S. W., Brewer G., Bernstein P., Hart P. A., Ross J. Regulation of mRNA turnover in eukaryotic cells. Crit Rev Eukaryot Gene Expr. 1991;1(2):99–126. [PubMed] [Google Scholar]
  31. Peltz S. W., Jacobson A. mRNA stability: in trans-it. Curr Opin Cell Biol. 1992 Dec;4(6):979–983. doi: 10.1016/0955-0674(92)90129-z. [DOI] [PubMed] [Google Scholar]
  32. Pierrat B., Lacroute F., Losson R. The 5' untranslated region of the PPR1 regulatory gene dictates rapid mRNA decay in yeast. Gene. 1993 Sep 6;131(1):43–51. doi: 10.1016/0378-1119(93)90667-r. [DOI] [PubMed] [Google Scholar]
  33. Sachs A. B. Messenger RNA degradation in eukaryotes. Cell. 1993 Aug 13;74(3):413–421. doi: 10.1016/0092-8674(93)80043-e. [DOI] [PubMed] [Google Scholar]
  34. Siomi H., Choi M., Siomi M. C., Nussbaum R. L., Dreyfuss G. Essential role for KH domains in RNA binding: impaired RNA binding by a mutation in the KH domain of FMR1 that causes fragile X syndrome. Cell. 1994 Apr 8;77(1):33–39. doi: 10.1016/0092-8674(94)90232-1. [DOI] [PubMed] [Google Scholar]
  35. Siomi H., Matunis M. J., Michael W. M., Dreyfuss G. The pre-mRNA binding K protein contains a novel evolutionarily conserved motif. Nucleic Acids Res. 1993 Mar 11;21(5):1193–1198. doi: 10.1093/nar/21.5.1193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Siomi H., Siomi M. C., Nussbaum R. L., Dreyfuss G. The protein product of the fragile X gene, FMR1, has characteristics of an RNA-binding protein. Cell. 1993 Jul 30;74(2):291–298. doi: 10.1016/0092-8674(93)90420-u. [DOI] [PubMed] [Google Scholar]
  37. Verkerk A. J., Pieretti M., Sutcliffe J. S., Fu Y. H., Kuhl D. P., Pizzuti A., Reiner O., Richards S., Victoria M. F., Zhang F. P. Identification of a gene (FMR-1) containing a CGG repeat coincident with a breakpoint cluster region exhibiting length variation in fragile X syndrome. Cell. 1991 May 31;65(5):905–914. doi: 10.1016/0092-8674(91)90397-h. [DOI] [PubMed] [Google Scholar]
  38. Volloch V., Housman D. Stability of globin mRNA in terminally differentiating murine erythroleukemia cells. Cell. 1981 Feb;23(2):509–514. doi: 10.1016/0092-8674(81)90146-x. [DOI] [PubMed] [Google Scholar]
  39. Wahle E., Keller W. The biochemistry of 3'-end cleavage and polyadenylation of messenger RNA precursors. Annu Rev Biochem. 1992;61:419–440. doi: 10.1146/annurev.bi.61.070192.002223. [DOI] [PubMed] [Google Scholar]
  40. Wang X., Kiledjian M., Weiss I. M., Liebhaber S. A. Detection and characterization of a 3' untranslated region ribonucleoprotein complex associated with human alpha-globin mRNA stability. Mol Cell Biol. 1995 Mar;15(3):1769–1777. doi: 10.1128/mcb.15.3.1769. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Weiss I. M., Liebhaber S. A. Erythroid cell-specific determinants of alpha-globin mRNA stability. Mol Cell Biol. 1994 Dec;14(12):8123–8132. doi: 10.1128/mcb.14.12.8123. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Weiss I. M., Liebhaber S. A. Erythroid cell-specific mRNA stability elements in the alpha 2-globin 3' nontranslated region. Mol Cell Biol. 1995 May;15(5):2457–2465. doi: 10.1128/mcb.15.5.2457. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Wong G., Müller O., Clark R., Conroy L., Moran M. F., Polakis P., McCormick F. Molecular cloning and nucleic acid binding properties of the GAP-associated tyrosine phosphoprotein p62. Cell. 1992 May 1;69(3):551–558. doi: 10.1016/0092-8674(92)90455-l. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES