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. 1997 Sep;17(9):5473–5484. doi: 10.1128/mcb.17.9.5473

Functional equivalency and diversity of cis-acting elements among yeast replication origins.

S Lin 1, D Kowalski 1
PMCID: PMC232396  PMID: 9271423

Abstract

The DNA replication origins of the yeast Saccharomyces cerevisiae require several short functional elements, most of which are not conserved in sequence. To better characterize ARS305, a replicator from a chromosomal origin, we swapped functional DNA elements of ARS305 with defined elements of ARS1. ARS305 contains elements that are functionally exchangeable with ARS1 A and B1 elements, which are known to bind the origin recognition complex; however, the ARS1 A element differs in that it does not require a 3' box adjacent to the essential autonomously replicating sequence consensus. At the position corresponding to ARS1 B3, ARS305 has a novel element, B4, that can functionally substitute for every type of short element (B1, B2, and B3) in the B domain. Unexpectedly, the replacement of element B4 by ARS1 B3, which binds ABF1p and is known as a replication enhancer, inhibited ARS305 function. ARS305 has no short functional element at or near positions corresponding to the B2 elements in ARS1 and ARS307 but contains an easily unwound region whose functional importance was supported by a broad G+C-rich substitution mutation. Surprisingly, the easily unwound region can functionally substitute for the ARS1 B2 element, even though ARS1 B2 was found to possess a distinct DNA sequence requirement. The functionally conserved B2 element in ARS307 contains a known sequence requirement, and helical stability analysis of linker and minilinker mutations suggested that B2 also contains a DNA unwinding element (DUE). Our findings suggest that yeast replication origins employ a B2 element or a DUE to mediate a common function, DNA unwinding during initiation, although not necessarily through a common mechanism.

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Selected References

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  1. Bell S. P., Stillman B. ATP-dependent recognition of eukaryotic origins of DNA replication by a multiprotein complex. Nature. 1992 May 14;357(6374):128–134. doi: 10.1038/357128a0. [DOI] [PubMed] [Google Scholar]
  2. Borowiec J. A., Hurwitz J. Localized melting and structural changes in the SV40 origin of replication induced by T-antigen. EMBO J. 1988 Oct;7(10):3149–3158. doi: 10.1002/j.1460-2075.1988.tb03182.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bramhill D., Kornberg A. Duplex opening by dnaA protein at novel sequences in initiation of replication at the origin of the E. coli chromosome. Cell. 1988 Mar 11;52(5):743–755. doi: 10.1016/0092-8674(88)90412-6. [DOI] [PubMed] [Google Scholar]
  4. Breslauer K. J., Frank R., Blöcker H., Marky L. A. Predicting DNA duplex stability from the base sequence. Proc Natl Acad Sci U S A. 1986 Jun;83(11):3746–3750. doi: 10.1073/pnas.83.11.3746. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Brewer B. J., Diller J. D., Friedman K. L., Kolor K. M., Raghuraman M. K., Fangman W. L. The topography of chromosome replication in yeast. Cold Spring Harb Symp Quant Biol. 1993;58:425–434. doi: 10.1101/sqb.1993.058.01.049. [DOI] [PubMed] [Google Scholar]
  6. Brewer B. J., Fangman W. L. The localization of replication origins on ARS plasmids in S. cerevisiae. Cell. 1987 Nov 6;51(3):463–471. doi: 10.1016/0092-8674(87)90642-8. [DOI] [PubMed] [Google Scholar]
  7. Buchman A. R., Kornberg R. D. A yeast ARS-binding protein activates transcription synergistically in combination with other weak activating factors. Mol Cell Biol. 1990 Mar;10(3):887–897. doi: 10.1128/mcb.10.3.887. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Celniker S. E., Sweder K., Srienc F., Bailey J. E., Campbell J. L. Deletion mutations affecting autonomously replicating sequence ARS1 of Saccharomyces cerevisiae. Mol Cell Biol. 1984 Nov;4(11):2455–2466. doi: 10.1128/mcb.4.11.2455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cocker J. H., Piatti S., Santocanale C., Nasmyth K., Diffley J. F. An essential role for the Cdc6 protein in forming the pre-replicative complexes of budding yeast. Nature. 1996 Jan 11;379(6561):180–182. doi: 10.1038/379180a0. [DOI] [PubMed] [Google Scholar]
  10. DePamphilis M. L. Eukaryotic DNA replication: anatomy of an origin. Annu Rev Biochem. 1993;62:29–63. doi: 10.1146/annurev.bi.62.070193.000333. [DOI] [PubMed] [Google Scholar]
  11. Deshpande A. M., Newlon C. S. The ARS consensus sequence is required for chromosomal origin function in Saccharomyces cerevisiae. Mol Cell Biol. 1992 Oct;12(10):4305–4313. doi: 10.1128/mcb.12.10.4305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Diffley J. F., Cocker J. H., Dowell S. J., Rowley A. Two steps in the assembly of complexes at yeast replication origins in vivo. Cell. 1994 Jul 29;78(2):303–316. doi: 10.1016/0092-8674(94)90299-2. [DOI] [PubMed] [Google Scholar]
  13. Dowell S. J., Romanowski P., Diffley J. F. Interaction of Dbf4, the Cdc7 protein kinase regulatory subunit, with yeast replication origins in vivo. Science. 1994 Aug 26;265(5176):1243–1246. doi: 10.1126/science.8066465. [DOI] [PubMed] [Google Scholar]
  14. Dubey D. D., Davis L. R., Greenfeder S. A., Ong L. Y., Zhu J. G., Broach J. R., Newlon C. S., Huberman J. A. Evidence suggesting that the ARS elements associated with silencers of the yeast mating-type locus HML do not function as chromosomal DNA replication origins. Mol Cell Biol. 1991 Oct;11(10):5346–5355. doi: 10.1128/mcb.11.10.5346. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Fangman W. L., Brewer B. J. A question of time: replication origins of eukaryotic chromosomes. Cell. 1992 Oct 30;71(3):363–366. doi: 10.1016/0092-8674(92)90505-7. [DOI] [PubMed] [Google Scholar]
  16. Fox C. A., Loo S., Dillin A., Rine J. The origin recognition complex has essential functions in transcriptional silencing and chromosomal replication. Genes Dev. 1995 Apr 15;9(8):911–924. doi: 10.1101/gad.9.8.911. [DOI] [PubMed] [Google Scholar]
  17. Friedman K. L., Diller J. D., Ferguson B. M., Nyland S. V., Brewer B. J., Fangman W. L. Multiple determinants controlling activation of yeast replication origins late in S phase. Genes Dev. 1996 Jul 1;10(13):1595–1607. doi: 10.1101/gad.10.13.1595. [DOI] [PubMed] [Google Scholar]
  18. Gille H., Messer W. Localized DNA melting and structural pertubations in the origin of replication, oriC, of Escherichia coli in vitro and in vivo. EMBO J. 1991 Jun;10(6):1579–1584. doi: 10.1002/j.1460-2075.1991.tb07678.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Greenfeder S. A., Newlon C. S. A replication map of a 61-kb circular derivative of Saccharomyces cerevisiae chromosome III. Mol Biol Cell. 1992 Sep;3(9):999–1013. doi: 10.1091/mbc.3.9.999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hay R. T., DePamphilis M. L. Initiation of SV40 DNA replication in vivo: location and structure of 5' ends of DNA synthesized in the ori region. Cell. 1982 Apr;28(4):767–779. doi: 10.1016/0092-8674(82)90056-3. [DOI] [PubMed] [Google Scholar]
  21. Holmes D. S., Quigley M. A rapid boiling method for the preparation of bacterial plasmids. Anal Biochem. 1981 Jun;114(1):193–197. doi: 10.1016/0003-2697(81)90473-5. [DOI] [PubMed] [Google Scholar]
  22. Hsiao C. L., Carbon J. High-frequency transformation of yeast by plasmids containing the cloned yeast ARG4 gene. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3829–3833. doi: 10.1073/pnas.76.8.3829. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Huang R. Y., Kowalski D. A DNA unwinding element and an ARS consensus comprise a replication origin within a yeast chromosome. EMBO J. 1993 Dec;12(12):4521–4531. doi: 10.1002/j.1460-2075.1993.tb06141.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Huang R. Y., Kowalski D. Multiple DNA elements in ARS305 determine replication origin activity in a yeast chromosome. Nucleic Acids Res. 1996 Mar 1;24(5):816–823. doi: 10.1093/nar/24.5.816. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Huberman J. A., Spotila L. D., Nawotka K. A., el-Assouli S. M., Davis L. R. The in vivo replication origin of the yeast 2 microns plasmid. Cell. 1987 Nov 6;51(3):473–481. doi: 10.1016/0092-8674(87)90643-x. [DOI] [PubMed] [Google Scholar]
  26. Huberman J. A., Zhu J. G., Davis L. R., Newlon C. S. Close association of a DNA replication origin and an ARS element on chromosome III of the yeast, Saccharomyces cerevisiae. Nucleic Acids Res. 1988 Jul 25;16(14A):6373–6384. doi: 10.1093/nar/16.14.6373. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Ito H., Fukuda Y., Murata K., Kimura A. Transformation of intact yeast cells treated with alkali cations. J Bacteriol. 1983 Jan;153(1):163–168. doi: 10.1128/jb.153.1.163-168.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Kearsey S. Structural requirements for the function of a yeast chromosomal replicator. Cell. 1984 May;37(1):299–307. doi: 10.1016/0092-8674(84)90326-x. [DOI] [PubMed] [Google Scholar]
  29. Kowalski D., Eddy M. J. The DNA unwinding element: a novel, cis-acting component that facilitates opening of the Escherichia coli replication origin. EMBO J. 1989 Dec 20;8(13):4335–4344. doi: 10.1002/j.1460-2075.1989.tb08620.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Liang C., Weinreich M., Stillman B. ORC and Cdc6p interact and determine the frequency of initiation of DNA replication in the genome. Cell. 1995 Jun 2;81(5):667–676. doi: 10.1016/0092-8674(95)90528-6. [DOI] [PubMed] [Google Scholar]
  31. Lin S., Kowalski D. DNA helical instability facilitates initiation at the SV40 replication origin. J Mol Biol. 1994 Jan 14;235(2):496–507. doi: 10.1006/jmbi.1994.1009. [DOI] [PubMed] [Google Scholar]
  32. Loo S., Laurenson P., Foss M., Dillin A., Rine J. Roles of ABF1, NPL3, and YCL54 in silencing in Saccharomyces cerevisiae. Genetics. 1995 Nov;141(3):889–902. doi: 10.1093/genetics/141.3.889. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Marahrens Y., Stillman B. A yeast chromosomal origin of DNA replication defined by multiple functional elements. Science. 1992 Feb 14;255(5046):817–823. doi: 10.1126/science.1536007. [DOI] [PubMed] [Google Scholar]
  34. Marahrens Y., Stillman B. Replicator dominance in a eukaryotic chromosome. EMBO J. 1994 Jul 15;13(14):3395–3400. doi: 10.1002/j.1460-2075.1994.tb06642.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Matsumoto K., Ishimi Y. Single-stranded-DNA-binding protein-dependent DNA unwinding of the yeast ARS1 region. Mol Cell Biol. 1994 Jul;14(7):4624–4632. doi: 10.1128/mcb.14.7.4624. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Miller C. A., Kowalski D. cis-acting components in the replication origin from ribosomal DNA of Saccharomyces cerevisiae. Mol Cell Biol. 1993 Sep;13(9):5360–5369. doi: 10.1128/mcb.13.9.5360. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Muzi-Falconi M., Brown G. W., Kelly T. J. Controlling initiation during the cell cycle. DNA replication. Curr Biol. 1996 Mar 1;6(3):229–233. doi: 10.1016/s0960-9822(02)00464-5. [DOI] [PubMed] [Google Scholar]
  38. Natale D. A., Schubert A. E., Kowalski D. DNA helical stability accounts for mutational defects in a yeast replication origin. Proc Natl Acad Sci U S A. 1992 Apr 1;89(7):2654–2658. doi: 10.1073/pnas.89.7.2654. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Natale D. A., Umek R. M., Kowalski D. Ease of DNA unwinding is a conserved property of yeast replication origins. Nucleic Acids Res. 1993 Feb 11;21(3):555–560. doi: 10.1093/nar/21.3.555. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Newlon C. S., Lipchitz L. R., Collins I., Deshpande A., Devenish R. J., Green R. P., Klein H. L., Palzkill T. G., Ren R. B., Synn S. Analysis of a circular derivative of Saccharomyces cerevisiae chromosome III: a physical map and identification and location of ARS elements. Genetics. 1991 Oct;129(2):343–357. doi: 10.1093/genetics/129.2.343. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Newlon C. S., Theis J. F. The structure and function of yeast ARS elements. Curr Opin Genet Dev. 1993 Oct;3(5):752–758. doi: 10.1016/s0959-437x(05)80094-2. [DOI] [PubMed] [Google Scholar]
  42. Palzkill T. G., Newlon C. S. A yeast replication origin consists of multiple copies of a small conserved sequence. Cell. 1988 May 6;53(3):441–450. doi: 10.1016/0092-8674(88)90164-x. [DOI] [PubMed] [Google Scholar]
  43. Parsons R., Anderson M. E., Tegtmeyer P. Three domains in the simian virus 40 core origin orchestrate the binding, melting, and DNA helicase activities of T antigen. J Virol. 1990 Feb;64(2):509–518. doi: 10.1128/jvi.64.2.509-518.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Perrin S., Gilliland G. Site-specific mutagenesis using asymmetric polymerase chain reaction and a single mutant primer. Nucleic Acids Res. 1990 Dec 25;18(24):7433–7438. doi: 10.1093/nar/18.24.7433. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Piatti S., Böhm T., Cocker J. H., Diffley J. F., Nasmyth K. Activation of S-phase-promoting CDKs in late G1 defines a "point of no return" after which Cdc6 synthesis cannot promote DNA replication in yeast. Genes Dev. 1996 Jun 15;10(12):1516–1531. doi: 10.1101/gad.10.12.1516. [DOI] [PubMed] [Google Scholar]
  46. Rao H., Marahrens Y., Stillman B. Functional conservation of multiple elements in yeast chromosomal replicators. Mol Cell Biol. 1994 Nov;14(11):7643–7651. doi: 10.1128/mcb.14.11.7643. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Rao H., Stillman B. The origin recognition complex interacts with a bipartite DNA binding site within yeast replicators. Proc Natl Acad Sci U S A. 1995 Mar 14;92(6):2224–2228. doi: 10.1073/pnas.92.6.2224. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Rivier D. H., Rine J. An origin of DNA replication and a transcription silencer require a common element. Science. 1992 May 1;256(5057):659–663. doi: 10.1126/science.1585179. [DOI] [PubMed] [Google Scholar]
  49. Rowley A., Cocker J. H., Harwood J., Diffley J. F. Initiation complex assembly at budding yeast replication origins begins with the recognition of a bipartite sequence by limiting amounts of the initiator, ORC. EMBO J. 1995 Jun 1;14(11):2631–2641. doi: 10.1002/j.1460-2075.1995.tb07261.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Rowley A., Dowell S. J., Diffley J. F. Recent developments in the initiation of chromosomal DNA replication: a complex picture emerges. Biochim Biophys Acta. 1994 Apr 6;1217(3):239–256. doi: 10.1016/0167-4781(94)90283-6. [DOI] [PubMed] [Google Scholar]
  51. Shirahige K., Iwasaki T., Rashid M. B., Ogasawara N., Yoshikawa H. Location and characterization of autonomously replicating sequences from chromosome VI of Saccharomyces cerevisiae. Mol Cell Biol. 1993 Aug;13(8):5043–5056. doi: 10.1128/mcb.13.8.5043. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Stillman B. Cell cycle control of DNA replication. Science. 1996 Dec 6;274(5293):1659–1664. doi: 10.1126/science.274.5293.1659. [DOI] [PubMed] [Google Scholar]
  53. Stinchcomb D. T., Struhl K., Davis R. W. Isolation and characterisation of a yeast chromosomal replicator. Nature. 1979 Nov 1;282(5734):39–43. doi: 10.1038/282039a0. [DOI] [PubMed] [Google Scholar]
  54. Theis J. F., Newlon C. S. Domain B of ARS307 contains two functional elements and contributes to chromosomal replication origin function. Mol Cell Biol. 1994 Nov;14(11):7652–7659. doi: 10.1128/mcb.14.11.7652. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Umek R. M., Kowalski D. The ease of DNA unwinding as a determinant of initiation at yeast replication origins. Cell. 1988 Feb 26;52(4):559–567. doi: 10.1016/0092-8674(88)90469-2. [DOI] [PubMed] [Google Scholar]
  56. Van Houten J. V., Newlon C. S. Mutational analysis of the consensus sequence of a replication origin from yeast chromosome III. Mol Cell Biol. 1990 Aug;10(8):3917–3925. doi: 10.1128/mcb.10.8.3917. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Walker S. S., Francesconi S. C., Eisenberg S. A DNA replication enhancer in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1990 Jun;87(12):4665–4669. doi: 10.1073/pnas.87.12.4665. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Walker S. S., Malik A. K., Eisenberg S. Analysis of the interactions of functional domains of a nuclear origin of replication from Saccharomyces cerevisiae. Nucleic Acids Res. 1991 Nov 25;19(22):6255–6262. doi: 10.1093/nar/19.22.6255. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Wold M. S. Replication protein A: a heterotrimeric, single-stranded DNA-binding protein required for eukaryotic DNA metabolism. Annu Rev Biochem. 1997;66:61–92. doi: 10.1146/annurev.biochem.66.1.61. [DOI] [PubMed] [Google Scholar]
  60. Yung B. Y., Kornberg A. The dnaA initiator protein binds separate domains in the replication origin of Escherichia coli. J Biol Chem. 1989 Apr 15;264(11):6146–6150. [PubMed] [Google Scholar]

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