Reverse transcriptase motifs in the catalytic subunit of telomerase

Science

Volumn 276, Issue 5312, 1997, Pages 561-567

  Lingner, Joachim ^a,d   Hughes, Timothy R ^b   Shevchenko, Andrej ^c   Mann, Matthias ^c   Lundblad, Victoria ^b   Cech, Thomas R ^a  

^a UNIVERSITY OF COLORADO   (United States)

^b BAYLOR COLLEGE OF MEDICINE   (United States)

^c EUROPEAN MOLECULAR BIOLOGY LABORATORY   (Germany)

^d SWISS INST FOR EXP CANCER RESEARCH   (Switzerland)

Author keywords

[No Author keywords available]

Indexed keywords

RIBONUCLEOPROTEIN; RNA DIRECTED DNA POLYMERASE; TELOMERASE;

AMINO ACID SUBSTITUTION; ARTICLE; CHROMOSOME REPLICATION; DNA DETERMINATION; ENZYME PURIFICATION; ENZYME SUBUNIT; GENE DELETION; PRIORITY JOURNAL; PROTEIN FOLDING; SEQUENCE ANALYSIS;

AMINO ACID SEQUENCE; ANIMALS; BINDING SITES; CATALYSIS; CHROMOSOMES; DNA, FUNGAL; DNA-BINDING PROTEINS; EUPLOTES; EVOLUTION, MOLECULAR; FUNGAL PROTEINS; GENES, FUNGAL; GENES, PROTOZOAN; MOLECULAR SEQUENCE DATA; PROTEIN CONFORMATION; RNA; RNA, FUNGAL; RNA, PROTOZOAN; RNA-DIRECTED DNA POLYMERASE; SACCHAROMYCES CEREVISIAE; SACCHAROMYCES CEREVISIAE PROTEINS; SEQUENCE ALIGNMENT; TELOMERASE; TELOMERE; TEMPLATES, GENETIC;

EID: 0030938901     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.276.5312.561     Document Type: Article

References (77)

1. J. D. Watson, Nature 239, 197 (1972); J. Olovnikov, J. Theor. Biol. 41, 181 (1973).
2. J. D. Watson, Nature 239, 197 (1972); J. Olovnikov, J. Theor. Biol. 41, 181 (1973).
3. J. Lingner, J. P. Cooper, T. R. Cech, Science 269, 1533 (1995); for a similar model regarding the macronuclear DNA of hypotrichous cillates, see A. M. Zahler and D. M. Prescott, Nucleic Acids Res. 16, 6953 (1988).
4. J. Lingner, J. P. Cooper, T. R. Cech, Science 269, 1533 (1995); for a similar model regarding the macronuclear DNA of hypotrichous cillates, see A. M. Zahler and D. M. Prescott, Nucleic Acids Res. 16, 6953 (1988).
5. C. W. Greider and E. H. Blackburn, Cell 51, 887 (1987).
6. _, Nature 337, 331 (1989); D. Shippen-Lenz and E. H. Blackburn, Science 247, 546 (1990); D. P. Romero and E. H. Blackburn, Cell 67, 343 (1991).
7. _, Nature 337, 331 (1989); D. Shippen-Lenz and E. H. Blackburn, Science 247, 546 (1990); D. P. Romero and E. H. Blackburn, Cell 67, 343 (1991).
8. _, Nature 337, 331 (1989); D. Shippen-Lenz and E. H. Blackburn, Science 247, 546 (1990); D. P. Romero and E. H. Blackburn, Cell 67, 343 (1991).
9. J. Lingner, L. L. Hendrick, T. R. Cech, Genes Dev. 8, 1984 (1994).
10. M. S. Singer and D. E. Gottschling, Science 266, 404 (1994).
11. M. J. McEachern and E. H. Blackburn, Nature 376, 403 (1995).
12. J. Feng et al., Science 269, 1236 (1995); M. A. Blasco, W. Funk, B. Villeponteau, C. W. Greider, ibid., p. 1267.
13. J. Feng et al., Science 269, 1236 (1995); M. A. Blasco, W. Funk, B. Villeponteau, C. W. Greider, ibid., p. 1267.
14. K. Collins, R. Kobayashi, C. W. Greider, Cell 81, 677 (1995).
15. L. Harrington et al., Science 275, 973 (1997); J. Nakayama, M. Saito, H. Nakamura, A. Matsuura, F. Ishikawa, Cell 88, 875 (1997).
16. L. Harrington et al., Science 275, 973 (1997); J. Nakayama, M. Saito, H. Nakamura, A. Matsuura, F. Ishikawa, Cell 88, 875 (1997).
17. J. Lingner and T. Nakamura, unpublished data. The BLAST search of protein databases was performed at the National Center for Biotechnology Information with GCG (Program Manual for the Wisconsin Package, Version 8.1, Genetics Computer Group, Madison, WI).
18. J. Lingner and T. Nakamura, unpublished data. The BLAST search of protein databases was performed at the National Center for Biotechnology Information with GCG (Program Manual for the Wisconsin Package, Version 8.1, Genetics Computer Group, Madison, WI).
19. J. Lingner and T. R. Cech, Proc. Natl. Acad. Sci. U.S.A. 93, 10712 (1996).
20. D. M. Prescott, Microbiol. Rev. 58, 233 (1994).
21. P. Hammond, T. Lively, T. R. Cech, Mol. Cell. Biol. 17, 296 (1997).
22. 2-terminal peptide t15 (MEVDVDNQADNHGIHSALK) and the sequence LFFAT in the peptide t16 (LFFATMDIEK). There are two discrepancies between the sequence determined by mass spectrometry and that deduced from the DNA sequence. The glutamine in peptide T2 was a lysine, and the order of the first two amino acids in T12 was reversed. In the above sequences, L signifies isoleucine or leucine, two isobaric amino acids that cannot be distinguished by our methods, and the parentheses indicate that the order of the first two amino acids could not be distinguished.
23. D. M. Wilm et al., Nature 379, 466 (1996).
24. M. S. Wilm and M. Mann, Int. J. Mass Spectrom. Ion Proc. 136, 167 (1994); Anal. Chem. 68, 1 (1996).
25. M. S. Wilm and M. Mann, Int. J. Mass Spectrom. Ion Proc. 136, 167 (1994); Anal. Chem. 68, 1 (1996).
26. J. B. Fenn, M. Mann, C. K. Meng, S. F. Wong, C. M. Whitehouse, Science 246, 64 (1989).
27. DNA was isolated from E. aediculatus, treated with mung bean nuclease, and subcloned into the Sma I site of pCR-Script SK (+) (Stratagene). The macronuclear gene encoding p123 was identified by colony hybridization.
28. J. Lingner and T. R. Cech, unpublished data.
29. M. Mann and M. S. Wilm, Anal. Chem. 66, 4390 (1994).
30. P is the probability that the specified segment pair score would be observed in a search of the same size with a random sequence.
31. J.-L. Ferat and F. Michel, Nature 364, 358 (1993).
32. T. S. Lendvay, D. K. Morris, J. Sah, B. Balasubramanian, V. Lundblad, Genetics 144, 1399 (1996).
33. EST1, which has been described [V. Lundblad and J. W. Szostak, Cell 57, 633 (1989)], is not essential for core telomerase activity (33). EST4 is identical to CDC13 [ B. Garvik, M. Carson, L. Hartwell, Mol. Cell. Biol. 15, 6128 (1995)]. Biochemical analysis has shown that Est1 p and Cdc13p are single-stranded telomeric DNA binding proteins that are proposed to facilitate access of telomerase to chromosome ends [ C. I. Nugent, T. R. Hughes, N. F. Lue, V. Lundblad, Science 274, 249 (1996); J. J. Lin and V. A. Zakian, Proc. Natl. Acad. Sci. U.S.A. 93, 13760 (1996); V. Virta-Pearlman, D. K. Morris, V. Lundblad, Genes Dev. 10, 3094 (1996)].
34. EST1, which has been described [V. Lundblad and J. W. Szostak, Cell 57, 633 (1989)], is not essential for core telomerase activity (33). EST4 is identical to CDC13 [ B. Garvik, M. Carson, L. Hartwell, Mol. Cell. Biol. 15, 6128 (1995)]. Biochemical analysis has shown that Est1 p and Cdc13p are single-stranded telomeric DNA binding proteins that are proposed to facilitate access of telomerase to chromosome ends [ C. I. Nugent, T. R. Hughes, N. F. Lue, V. Lundblad, Science 274, 249 (1996); J. J. Lin and V. A. Zakian, Proc. Natl. Acad. Sci. U.S.A. 93, 13760 (1996); V. Virta-Pearlman, D. K. Morris, V. Lundblad, Genes Dev. 10, 3094 (1996)].
35. EST1, which has been described [V. Lundblad and J. W. Szostak, Cell 57, 633 (1989)], is not essential for core telomerase activity (33). EST4 is identical to CDC13 [ B. Garvik, M. Carson, L. Hartwell, Mol. Cell. Biol. 15, 6128 (1995)]. Biochemical analysis has shown that Est1 p and Cdc13p are single-stranded telomeric DNA binding proteins that are proposed to facilitate access of telomerase to chromosome ends [ C. I. Nugent, T. R. Hughes, N. F. Lue, V. Lundblad, Science 274, 249 (1996); J. J. Lin and V. A. Zakian, Proc. Natl. Acad. Sci. U.S.A. 93, 13760 (1996); V. Virta-Pearlman, D. K. Morris, V. Lundblad, Genes Dev. 10, 3094 (1996)].
36. EST1, which has been described [V. Lundblad and J. W. Szostak, Cell 57, 633 (1989)], is not essential for core telomerase activity (33). EST4 is identical to CDC13 [ B. Garvik, M. Carson, L. Hartwell, Mol. Cell. Biol. 15, 6128 (1995)]. Biochemical analysis has shown that Est1 p and Cdc13p are single-stranded telomeric DNA binding proteins that are proposed to facilitate access of telomerase to chromosome ends [ C. I. Nugent, T. R. Hughes, N. F. Lue, V. Lundblad, Science 274, 249 (1996); J. J. Lin and V. A. Zakian, Proc. Natl. Acad. Sci. U.S.A. 93, 13760 (1996); V. Virta-Pearlman, D. K. Morris, V. Lundblad, Genes Dev. 10, 3094 (1996)].
37. EST1, which has been described [V. Lundblad and J. W. Szostak, Cell 57, 633 (1989)], is not essential for core telomerase activity (33). EST4 is identical to CDC13 [ B. Garvik, M. Carson, L. Hartwell, Mol. Cell. Biol. 15, 6128 (1995)]. Biochemical analysis has shown that Est1 p and Cdc13p are single-stranded telomeric DNA binding proteins that are proposed to facilitate access of telomerase to chromosome ends [ C. I. Nugent, T. R. Hughes, N. F. Lue, V. Lundblad, Science 274, 249 (1996); J. J. Lin and V. A. Zakian, Proc. Natl. Acad. Sci. U.S.A. 93, 13760 (1996); V. Virta-Pearlman, D. K. Morris, V. Lundblad, Genes Dev. 10, 3094 (1996)].
38. O. Poch, I. Sauvaget, M. Delarue, N. Tordo, EMBO J. 8, 3867 (1989).
39. Y. Xiong and T. H. Eickbush, ibid. 9, 3353 (1990).
40. V. Lundblad and E. H. Blackburn, Cell 73, 347 (1993).
41. C. Nugent and V. Lundblad, unpublished data.
42. S. G. Sarafianos, V. N. Pandey, N. Kaushik, M. J. Modak, Biochemistry 34, 7207 (1995).
43. 3-tagged gene and was detected on a Northern (RNA) blot. U1 snRNA was not substantially precipitated but was detected in the crude extract.
44. 32P]dTTP (800 Ci/ mmol; 10 μCi/μl), and 10% (v/v) telomerase fraction. After 1 hour at 30°C, the reactions were stopped, digested with proteinase K, ethanol precipitated, and separated as in (5). Fractions were derived from the following strains: Fig. 6, A and B: TVL268 [Mata ura3-52 ade2-101 lys2-801 leu2-Δ1 his3-Δ200/CF (TRP1 SUP11)]; Fig. 6C, lanes 2 to 4: TVL268; Fig. 6C, lanes 6 to 8: TVL268 TLC1-1(HaeIII); Fig. 6D, lanes 1 and 2, and Fig. 6E, lane 1 : AVL78 (Mata leu2 trp1 ura3-52 prb prc pep4-1); Fig. 6D, lanes 3 and 4, and Fig. 6E, lane 2: AVL78 est2-Δ1::UR43; Fig. 6D, lanes 5 and 6, and Fig. 6E, lane 3: AVL78 tlc7-Δ::LEU2; Fig. 6E, lanes 4, 7, 8, and 9: TVL268; Fig. 6E, lanes 5 and 8: TVL268 est2-D530A; and Fig. 6E, lanes 6 and 9: TVL268 est2-D671A. The est2-Δ1 and tlc1-Δ deletion strains were constructed by one-step gene disruption, whereas the est2-D530A, est2-D671A and TLC1-1(HaeIII) mutant alleles were constructed by replacement of the EST2-wt and TLC1-wt wild-type alleles, respectively. In each case, extracts were prepared for analysis after as few generations of growth as possible.
45. M. Cohn and E. H. Blackburn, Science 269, 396 (1995); J. Prescott and E. H. Blackburn, Genes. Dev. 11, 528 (1997).
46. M. Cohn and E. H. Blackburn, Science 269, 396 (1995); J. Prescott and E. H. Blackburn, Genes. Dev. 11, 528 (1997).
47. N. F. Lue and J. C. Wang, J. Biol. Chem. 270, 21453 (1995).
48. L. A. Kohlstaedt, J. Wang, J. M. Friedman, P. A. Rice, T. A. Steitz, Science 256, 1783 (1992); E. Arnold et al., Nature 357, 85 (1992); A. Jacobo-Molina et al., Proc. Natl. Acad. Sci. U.S.A. 90, 6320 (1993).
49. L. A. Kohlstaedt, J. Wang, J. M. Friedman, P. A. Rice, T. A. Steitz, Science 256, 1783 (1992); E. Arnold et al., Nature 357, 85 (1992); A. Jacobo-Molina et al., Proc. Natl. Acad. Sci. U.S.A. 90, 6320 (1993).
50. L. A. Kohlstaedt, J. Wang, J. M. Friedman, P. A. Rice, T. A. Steitz, Science 256, 1783 (1992); E. Arnold et al., Nature 357, 85 (1992); A. Jacobo-Molina et al., Proc. Natl. Acad. Sci. U.S.A. 90, 6320 (1993).
51. D. L. Ollis, P. Brick, T. A. Steitz, Nature 313, 765 (1985). For comparison of different polymerase structures, see C. M. Joyce and T. A. Steitz, Annu. Rev. Biochem. 63, 777 (1994), and R. Sousa, Trends Biochem. Sci. 21, 186 (1996).
52. D. L. Ollis, P. Brick, T. A. Steitz, Nature 313, 765 (1985). For comparison of different polymerase structures, see C. M. Joyce and T. A. Steitz, Annu. Rev. Biochem. 63, 777 (1994), and R. Sousa, Trends Biochem. Sci. 21, 186 (1996).
53. D. L. Ollis, P. Brick, T. A. Steitz, Nature 313, 765 (1985). For comparison of different polymerase structures, see C. M. Joyce and T. A. Steitz, Annu. Rev. Biochem. 63, 777 (1994), and R. Sousa, Trends Biochem. Sci. 21, 186 (1996).
54. J. C. Kennell, H. Wang, A. M. Lambowitz, Mol. Cell. Biol. 14, 3094 (1994); S. Zimmerly et al., Cell 83, 529 (1995).
55. J. C. Kennell, H. Wang, A. M. Lambowitz, Mol. Cell. Biol. 14, 3094 (1994); S. Zimmerly et al., Cell 83, 529 (1995).
56. D. Baltimore, Nature 226, 1209 (1970); H. M. Temin and S. Mizutani, ibid., p. 1211.
57. D. Baltimore, Nature 226, 1209 (1970); H. M. Temin and S. Mizutani, ibid., p. 1211.
58. J. D. Boeke and V. G. Corces, Annu. Rev. Micro. 43, 403 (1989).
59. M. Inouye and S. Inouye, ibid. 45, 163 (1991).
60. H. Wang and A. M. Lambowitz, Cell 75, 1071 (1993).
61. H. M. Temin, Annu. Rev. Genet. 8, 155 (1974).
62. J. E. Darnell and W. F. Doolittle, Proc. Natl. Acad. Sci. U.S.A. 83, 1271 (1986). See also N. Maizels and A. Weiner, in The RNA World, R. F. Gesteland and J. F. Atkins Eds. (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1993), pp. 577-602.
63. J. E. Darnell and W. F. Doolittle, Proc. Natl. Acad. Sci. U.S.A. 83, 1271 (1986). See also N. Maizels and A. Weiner, in The RNA World, R. F. Gesteland and J. F. Atkins Eds. (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1993), pp. 577-602.
64. H. Biessmann et al., Cell 61, 663 (1990); R. W. Levis, R. Ganesan, K. Houtchens, L. A. Tolar, F. Sheen, ibid. 75, 1 (1993).
65. H. Biessmann et al., Cell 61, 663 (1990); R. W. Levis, R. Ganesan, K. Houtchens, L. A. Tolar, F. Sheen, ibid. 75, 1 (1993).
66. M. L. Pardue, O. N. Danilevskaya, K. Lowenhaupt, F. Slot, K. L. Traverse, Trends Genet. 12, 48 (1996).
67. J. Lingner, A. Shevchenko, M. Mann, T. R. Cech, unpublished data.
68. See, for example, A. Shevchenko et al., Proc. Natl. Acad. Sci. U.S.A. 93, 14440 (1996). Reviewed in S. D. Patterson and R. Aebersold, Electrophoresis 16, 791 (1995).
69. See, for example, A. Shevchenko et al., Proc. Natl. Acad. Sci. U.S.A. 93, 14440 (1996). Reviewed in S. D. Patterson and R. Aebersold, Electrophoresis 16, 791 (1995).
70. C. M. Counter et al., EMBO J. 11, 1921 (1992); T. de Lange, Proc. Natl. Acad. Sci. U.S.A. 91, 2882 (1994).
71. C. M. Counter et al., EMBO J. 11, 1921 (1992); T. de Lange, Proc. Natl. Acad. Sci. U.S.A. 91, 2882 (1994).
72. C. Strahl and E. H. Blackburn, Nucleic Acids Res. 22, 893 (1994); Mol. Cell. Biol. 16, 53 (1996).
73. C. Strahl and E. H. Blackburn, Nucleic Acids Res. 22, 893 (1994); Mol. Cell. Biol. 16, 53 (1996).
74. Single-letter abbreviations for the amino acid residues are as follows: A, Ala; C, Cys; D, Asp; E, Glu; F, Phe; G, Gly; H, His; I, Ile; K, Lys; L, Leu; M, Met; N, Asn; P, Pro; Q, Gln; R, Arg; S, Ser; T, Thr; V, Val; W, Trp; and Y, Tyr.
75. D671A mutants) was enhanced in an est2 rad52 strain background (29). The absence of RAD52 gene function has been shown to eliminate a backup pathway for telomere maintenance, thereby enhancing the phenotype of est mutant strains (24, 28).
76. S. Evans and V. Lundblad, unpublished data.
77. We thank C. Grosshans, K. Goodrich, and N. Walcott for technical assistance; T. Nakamura, C. Chapon, A. Zaug, and J. Hansen for many helpful discussions; and D. Underwood and E. Jabri for help with Fig. 7. Supported in part by the Swiss National Science Foundation (J.L.), NIH grant AG11728 and Geron Corporation (V.L.), and a grant from the German Technology Ministry (Bundesministerium für Biologische Forschung) (M.M.).