Abstract
We have identified and characterized protein factors from mung bean (Vigna radiata) nuclear extracts that specifically bind the single-stranded G-rich telomeric DNA repeats. Nuclear extracts were prepared from three different types of plant tissue, radicle, hypocotyl, and root, in order to examine changes in the expression patterns of telomere-binding proteins during the development of mung bean. At least three types of specific complexes (A, B, and C) were detected by gel retardation assays with synthetic telomere and nuclear extract from radicle tissue, whereas the two major faster-migrating complexes (A and B) were formed with nuclear extracts from hypocotyl and root tissues. Gel retardation assays also revealed differences in relative amount of each complex forming activity in radicle, hypocotyl, and root nuclear extracts. These data suggest that the expression of telomere-binding proteins is developmentally regulated in plants, and that the factor involved in the formation of complex C may be required during the early stages of development. The binding factors have properties of proteins and are hence designated as mung bean G-rich telomere-binding proteins (MGBP). MGBPs bind DNA substrates with three or more single-stranded TTTAGGG repeats, while none of them show binding affinity to either double-stranded or single-stranded C-rich telomeric DNA. These proteins have a lower affinity to human telomeric sequences than to plant telomeric sequences and do not exhibit a significant binding activity to Tetrahymena telomeric sequence or mutated plant telomeric sequences, indicating that their binding activities are specific to plant telomere. Furthermore, RNase treatment of the nuclear extracts did not affect the complex formation activities. This result indicates that the single-stranded telomere-binding activities may be attributed to a simple protein but not a ribonucleoprotein. The ability of MGBPs to bind specifically the single-stranded TTTAGGG repeats may suggest their in vivo functions in the chromosome ends of plants.
Similar content being viewed by others
References
Blackburn, E.H. 1991. Structure and function of telomeres. Nature 350: 569–573.
Blackburn, E.H. 1992. Telomerase. Annu. Rev. Biochem. 61: 113–129.
Broccoli, D., Smogorzewska, A., Chong, L. and de Lange, T. 1997. Human telomeres contain two distinct Myb-related proteins, TRF1 and TRF2. Nature Genet. 17: 231–235.
Cardenas, M.E., Bianchi, A. and de Lange, T. 1993. A Xenopus egg factor with DNA-binding properties characteristic of terminusspecific telomeric proteins. Genes Dev. 7: 883–894.
Chong, L., Steensel, B.V., Broccoli, D., Erdjument-Bromage, H., Hanish, J., Tempst, P. and de Lange, T. 1995. A human telomeric protein. Science 270: 1663–1667.
Coren, J., Epstein, E. and Vogt, V. 1991. Characterization of a telomere-binding protein from Physarum polycephalum. Mol. Cell. Biol. 11: 2282–2290.
Fajkus, J., Kovarik, A. and Kralovics, R. 1996. Telomerase activity in plant cells. FEBS Lett. 391: 307–309.
Fang, G. and Cech, T.R. 1993. The β subunit of Oxytricha telomerebinding protein promotes G-quartet formation by telomeric DNA. Cell 74: 875–885.
Fitzgerald, M.S., McKnight, T.D. and Shippen, D.E. 1996. Characterization and developmental patterns of telomerase expression in plants. Proc. Natl. Acad. Sci. USA 93: 14422–14427.
Froelich-Ammon, S.J., Dickinson, B.A., Bevilacqua, J.M., Schultz, S.C. and Cech, T.R. 1998. Modulation of telomerase activity by telomere DNA-binding proteins in Oxytricha. Genes Dev. 12: 1504–1514.
Ganal, M.W., Lapitan, N L. and Tanksley, S.D. 1991. Macrostructure of the tomato telomeres. Plant Cell 3: 87–94.
Gottschling, D.E. and Zakian, V.A. 1986. Telomere proteins: specific recognition and protection of the natural termini of Oxytricha macronuclear DNA. Cell 47: 195–205.
Greider, C.W. 1996. Telomere length regulation. Annu. Rev. Biochem. 65: 337–365.
Greider, C.W. 1999. Telomeres do D-loop-T-loop. Cell 97: 419–422.
Griffith, J.D., Comeau, L., Rosenfield, S., Stansel, R.M., Bianchi, A., Moss, H. and de Lange, T. 1999. Mammalian telomeres end in a large duplex loop. Cell 97: 503–514.
Heller, K., Kilian, A., Piatyszek, M.A. and Kleinhofs, A. 1996. Telomerase activity in plant extracts. Mol. Gen. Genet. 252: 342–345.
Hubbard, K., Dhanaraj, S.N., Sethi, K.A., Rhodes, J., Wilusz, J., Small, M.B. and Ozer, H.L. 1995. Alteration of DNA and RNA binding activity of human telomere binding proteins occurs during cellular senescence. Exp. Cell Res. 218: 241–247.
Ishikawa, F., Matunis, M.J., Dreyfuss, G. and Cech, T.R. 1993. Nuclear proteins that bind the pre-mRNA 3' splice site sequence r(UUAG/G) and the human telomeric DNA sequence d(TTAGGG)n. Mol. Cell. Biol. 13: 4301–4310.
Kennedy, B.K., Gotta, M., Sinclair, D.A., Mills, K., McNabb, D.S., Murthy, M., Pak, S.M., Laroche, T., Gasser, S.M. and Guarente, L. 1997. Redistribution of silencing proteins from telomeres to the nucleolus is associated with extension of life span in S. cerevisiae. Cell 89: 381–391.
Kilian, A., Heller, K. and Kleinhofs, A. 1998. Development patterns of telomerase activity in barley and maize. Plant Mol. Biol. 37: 621–628.
Kilian, A. and Kleinhofs, A. 1992. Cloning and mapping of telomere-associated sequences from Hordeum vulgare L. Mol. Gen. Genet. 235: 153–156.
Kilian, A., Stiff, C. and Kleinhofs, A. 1995. Barley telomeres shorten during differentiation but grow in callus culture. Proc. Natl. Acad. Sci. USA 92: 9555–9559.
Kim, J.H., Kim, W.T. and Chung, I.K. 1998. Rice proteins that bind single-stranded G-rich telomere DNA. Plant Mol. Biol. 36: 661–672.
Levis, R.W., Ganesan, R., Houtchens, K., Tolar, L.A. and Sheen, F.M. 1993. Transposons in place of telomeric repeats at a Drosophila telomere. Cell 75: 1083–1093.
Liu, Z. and Gilbert, W. 1994. The yeast KEM1 gene encodes a nuclease specific for G4 tetraplex DNA: implication of in vivo functions for this novel DNA structure. Cell 77: 1083–1092.
Lustig, A.J., Kurtz, S. and Shore, D. 1990. Involvement of the silencer and UAS binding protein RAP1 in regulation of telomere length. Science 250: 549–553.
Makarov, V.L., Hirose, Y. and Langmore, J.P. 1997. Long G tails at both ends of human chromosomes suggest a C strand degradation mechanism for telomere shortening. Cell 88: 657–666.
Nugent, C.I., Hughes, T.R., Lue, N.F. and Lundblad, V. 1996. Cdc13p: a single-strand telomeric DNA-binding protein with a dual role in yeast telomere maintenance. Science 274: 249–252.
Ossipow, V., Laemmli, U.K. and Schibler, U. 1993. A simple method to renature DNA-binding proteins separated by SDSpolyacrylamide gel electrophoresis. Nucl. Acids Res 21: 6040–6041.
Petracek, M.E., Konkel, L.M.C., Kable, M.L. and Berman, J. 1994. A Chlamydomonas protein that binds single-stranded G-rich telomere DNA. EMBO J. 13: 3648–3658.
Price, C.M. 1990. Telomere structure in Euplotes crassus: characterization of DNA-protein interactions and isolation of a telomere-binding protein. Mol. Cell. Biol. 10: 3421–3431.
Price, C.M. and Cech, T.R. 1987. Telomeric DNA-protein interactions of Oxytricha macronuclear DNA. Genes Dev. 1: 783–793.
Price, C.M. and Cech, T.R. 1989. Properties of the telomeric DNA-binding protein from Oxytricha nova. Biochemistry 28: 769–774.
Regad, F., Lebas, M. and Lescure, B. 1994. Interstitial telomeric repeats within the Arabidopsis thaliana genome. J. Mol. Biol. 239: 163–169.
Richards, E.J. and Ausubel, F.M. 1988. Isolation of a higher eukaryotic telomere from Arabidopsis thaliana. Cell 53: 127–136.
Riha, K., Fajkus, J., Siroky, J. and Vyskot, B. 1998. Developmental control of telomere lengths and telomerase activity in plants. Plant Cell 10: 1691–1698.
Schierer, T. and Henderson, E. 1994. A protein from Tetrahymena thermophila that specifically binds parallel-stranded G4-DNA. Biochemistry 33: 2240–2246.
Sheng, H., Hou, Z., Schierer, T., Dobbs, D.L. and Henderson, E. 1995. Identification and characterization of a putative telomere end-binding protein from Tetrahymena thermophila. Mol. Cell. Biol. 15: 1144–1153.
Sussel, L. and Shore, D. 1991. Separation of transcriptional activation and silencing functions of RAP1-encoded repressor/ activator protein 1: isolation of viable mutants affecting both silencing and telomere length. Proc. Natl. Acad. Sci. USA 88: 7749–7753.
van Steensel, B. and de Lange, T. 1997. Control of telomere length by the human telomeric protein TRF1. Nature 385: 740–749.
van Steensel, B., Smogorzewska, A. and de Lange, T. 1998. TRF2 protects human telomeres from end-to-end fusions. Cell 92: 401–413.
Wellinger, R.J., Wolf, A.J. and Zakian, V.A. 1993. Saccharomyces telomeres acquire single-strand TG1-2tails late in S phase. Cell 72: 51–60.
Wright, W.E., Tesmer, V.M., Huffman, K.E., Levene, S.D. and Shay, J.W. 1997. Normal human chromosomes have long G-rich telomeric overhangs at one end. Genes Dev. 11: 2810–2821.
Wu, K.S. and Tanksley, S.D. 1993. Genetic and physical mapping of telomeres and macrosatellites of rice. Plant Mol. Biol. 22: 861–872.
Zakian, V.A. 1995. Telomere: beginning to understand the end. Science 270: 1601–1606.
Zentgraf, U. 1995. Telomere-binding proteins of from Arabidopsis thaliana. Plant Mol. Biol. 27: 467–475.
Zhong, Z., Shiue, L., Kaplan, S. and de Lange, T. 1992. A mammalian factor that binds telomeric TTAGGG repeats in vitro. Mol. Cell. Biol. 12: 4834–4843.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Ho Lee, J., Hyun Kim, J., Taek Kim, W. et al. Characterization and developmental expression of single-stranded telomeric DNA-binding proteins from mung bean (Vigna radiata). Plant Mol Biol 42, 547–557 (2000). https://doi.org/10.1023/A:1006373917321
Issue Date:
DOI: https://doi.org/10.1023/A:1006373917321