Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1991 Nov;11(11):5718–5726. doi: 10.1128/mcb.11.11.5718

CDC68, a yeast gene that affects regulation of cell proliferation and transcription, encodes a protein with a highly acidic carboxyl terminus.

A Rowley 1, R A Singer 1, G C Johnston 1
PMCID: PMC361943  PMID: 1833637

Abstract

The cell cycle of the budding yeast Saccharomyces cerevisiae has been investigated through the study of conditional cdc mutations that specifically affect cell cycle performance. Cells bearing the cdc68-1 mutation (J. A. Prendergast, L. E. Murray, A. Rowley, D. R. Carruthers, R. A. Singer, and G. C. Johnston, Genetics 124:81-90, 1990) are temperature sensitive for the performance of the G1 regulatory event, START. Here we describe the CDC68 gene and present evidence that the CDC68 gene product functions in transcription. CDC68 encodes a 1,035-amino-acid protein with a highly acidic and serine-rich carboxyl terminus. The abundance of transcripts from several unrelated genes is decreased in cdc68-1 mutant cells after transfer to the restrictive temperature, while at least one transcript, from the HSP82 gene, persists in an aberrant fashion. Thus, the cdc68-1 mutation has both positive and negative effects on gene expression. Our findings complement those of Malone et al. (E. A. Malone, C. D. Clark, A. Chiang, and F. Winston, Mol. Cell. Biol. 11:5710-5717, 1991), who have independently identified the CDC68 gene (as SPT16) as a transcriptional suppressor of delta-insertion mutations. Among transcripts that rapidly become depleted in cdc68-1 mutant cells are those of the G1 cyclin genes CLN1, CLN2, and CLN3/WHI1/DAF1, whose activity has been previously shown to be required for the performance of START. The decreased abundance of cyclin transcripts in cdc68-1 mutant cells, coupled with the suppression of cdc68-1-mediated START arrest by the CLN2-1 hyperactive allele of CLN2, shows that the CDC68 gene affects START through cyclin gene expression.

Full text

PDF
5719

Images in this article

5722Image
on p.5722
5722Image
on p.5722
5723Image
on p.5723

Selected References

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

  1. Borkovich K. A., Farrelly F. W., Finkelstein D. B., Taulien J., Lindquist S. hsp82 is an essential protein that is required in higher concentrations for growth of cells at higher temperatures. Mol Cell Biol. 1989 Sep;9(9):3919–3930. doi: 10.1128/mcb.9.9.3919. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Botstein D., Falco S. C., Stewart S. E., Brennan M., Scherer S., Stinchcomb D. T., Struhl K., Davis R. W. Sterile host yeasts (SHY): a eukaryotic system of biological containment for recombinant DNA experiments. Gene. 1979 Dec;8(1):17–24. doi: 10.1016/0378-1119(79)90004-0. [DOI] [PubMed] [Google Scholar]
  3. Carter B. L., Sudbery P. E. Small-sized mutants of Saccharomyces cerevisiae. Genetics. 1980 Nov;96(3):561–566. doi: 10.1093/genetics/96.3.561. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Clark-Adams C. D., Norris D., Osley M. A., Fassler J. S., Winston F. Changes in histone gene dosage alter transcription in yeast. Genes Dev. 1988 Feb;2(2):150–159. doi: 10.1101/gad.2.2.150. [DOI] [PubMed] [Google Scholar]
  5. Clark-Adams C. D., Winston F. The SPT6 gene is essential for growth and is required for delta-mediated transcription in Saccharomyces cerevisiae. Mol Cell Biol. 1987 Feb;7(2):679–686. doi: 10.1128/mcb.7.2.679. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cross F. R. Cell cycle arrest caused by CLN gene deficiency in Saccharomyces cerevisiae resembles START-I arrest and is independent of the mating-pheromone signalling pathway. Mol Cell Biol. 1990 Dec;10(12):6482–6490. doi: 10.1128/mcb.10.12.6482. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cross F. R. DAF1, a mutant gene affecting size control, pheromone arrest, and cell cycle kinetics of Saccharomyces cerevisiae. Mol Cell Biol. 1988 Nov;8(11):4675–4684. doi: 10.1128/mcb.8.11.4675. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Earnshaw W. C. Anionic regions in nuclear proteins. J Cell Biol. 1987 Oct;105(4):1479–1482. doi: 10.1083/jcb.105.4.1479. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Eisenmann D. M., Dollard C., Winston F. SPT15, the gene encoding the yeast TATA binding factor TFIID, is required for normal transcription initiation in vivo. Cell. 1989 Sep 22;58(6):1183–1191. doi: 10.1016/0092-8674(89)90516-3. [DOI] [PubMed] [Google Scholar]
  11. Farrelly F. W., Finkelstein D. B. Complete sequence of the heat shock-inducible HSP90 gene of Saccharomyces cerevisiae. J Biol Chem. 1984 May 10;259(9):5745–5751. [PubMed] [Google Scholar]
  12. Fassler J. S., Winston F. Isolation and analysis of a novel class of suppressor of Ty insertion mutations in Saccharomyces cerevisiae. Genetics. 1988 Feb;118(2):203–212. doi: 10.1093/genetics/118.2.203. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Finkelstein D. B., Strausberg S. Heat shock-regulated production of Escherichia coli beta-galactosidase in Saccharomyces cerevisiae. Mol Cell Biol. 1983 Sep;3(9):1625–1633. doi: 10.1128/mcb.3.9.1625. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Finkelstein D. B., Strausberg S. Identification and expression of a cloned yeast heat shock gene. J Biol Chem. 1983 Feb 10;258(3):1908–1913. [PubMed] [Google Scholar]
  15. Finkelstein D. B., Strausberg S., McAlister L. Alterations of transcription during heat shock of Saccharomyces cerevisiae. J Biol Chem. 1982 Jul 25;257(14):8405–8411. [PubMed] [Google Scholar]
  16. Hadwiger J. A., Wittenberg C., Richardson H. E., de Barros Lopes M., Reed S. I. A family of cyclin homologs that control the G1 phase in yeast. Proc Natl Acad Sci U S A. 1989 Aug;86(16):6255–6259. doi: 10.1073/pnas.86.16.6255. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hartwell L. H. Macromolecule synthesis in temperature-sensitive mutants of yeast. J Bacteriol. 1967 May;93(5):1662–1670. doi: 10.1128/jb.93.5.1662-1670.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hartwell L. H. Saccharomyces cerevisiae cell cycle. Bacteriol Rev. 1974 Jun;38(2):164–198. doi: 10.1128/br.38.2.164-198.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Henikoff S. Unidirectional digestion with exonuclease III in DNA sequence analysis. Methods Enzymol. 1987;155:156–165. doi: 10.1016/0076-6879(87)55014-5. [DOI] [PubMed] [Google Scholar]
  20. Herrick D., Parker R., Jacobson A. Identification and comparison of stable and unstable mRNAs in Saccharomyces cerevisiae. Mol Cell Biol. 1990 May;10(5):2269–2284. doi: 10.1128/mcb.10.5.2269. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hill J. E., Myers A. M., Koerner T. J., Tzagoloff A. Yeast/E. coli shuttle vectors with multiple unique restriction sites. Yeast. 1986 Sep;2(3):163–167. doi: 10.1002/yea.320020304. [DOI] [PubMed] [Google Scholar]
  22. Hinnen A., Hicks J. B., Fink G. R. Transformation of yeast. Proc Natl Acad Sci U S A. 1978 Apr;75(4):1929–1933. doi: 10.1073/pnas.75.4.1929. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hope I. A., Struhl K. Functional dissection of a eukaryotic transcriptional activator protein, GCN4 of yeast. Cell. 1986 Sep 12;46(6):885–894. doi: 10.1016/0092-8674(86)90070-x. [DOI] [PubMed] [Google Scholar]
  24. Jakobsen B. K., Pelham H. R. A conserved heptapeptide restrains the activity of the yeast heat shock transcription factor. EMBO J. 1991 Feb;10(2):369–375. doi: 10.1002/j.1460-2075.1991.tb07958.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Johnston G. C., Pringle J. R., Hartwell L. H. Coordination of growth with cell division in the yeast Saccharomyces cerevisiae. Exp Cell Res. 1977 Mar 1;105(1):79–98. doi: 10.1016/0014-4827(77)90154-9. [DOI] [PubMed] [Google Scholar]
  26. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  27. Lemmon S. K., Pellicena-Palle A., Conley K., Freund C. L. Sequence of the clathrin heavy chain from Saccharomyces cerevisiae and requirement of the COOH terminus for clathrin function. J Cell Biol. 1991 Jan;112(1):65–80. doi: 10.1083/jcb.112.1.65. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Lewin B. Driving the cell cycle: M phase kinase, its partners, and substrates. Cell. 1990 Jun 1;61(5):743–752. doi: 10.1016/0092-8674(90)90181-d. [DOI] [PubMed] [Google Scholar]
  29. Lindquist S., Craig E. A. The heat-shock proteins. Annu Rev Genet. 1988;22:631–677. doi: 10.1146/annurev.ge.22.120188.003215. [DOI] [PubMed] [Google Scholar]
  30. Ma J., Ptashne M. Deletion analysis of GAL4 defines two transcriptional activating segments. Cell. 1987 Mar 13;48(5):847–853. doi: 10.1016/0092-8674(87)90081-x. [DOI] [PubMed] [Google Scholar]
  31. Malone E. A., Clark C. D., Chiang A., Winston F. Mutations in SPT16/CDC68 suppress cis- and trans-acting mutations that affect promoter function in Saccharomyces cerevisiae. Mol Cell Biol. 1991 Nov;11(11):5710–5717. doi: 10.1128/mcb.11.11.5710. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Marchiori F., Meggio F., Marin O., Borin G., Calderan A., Ruzza P., Pinna L. A. Synthetic peptide substrates for casein kinase 2. Assessment of minimum structural requirements for phosphorylation. Biochim Biophys Acta. 1988 Oct 7;971(3):332–338. doi: 10.1016/0167-4889(88)90149-8. [DOI] [PubMed] [Google Scholar]
  33. McAlister L., Finkelstein D. B. Alterations in translatable ribonucleic acid after heat shock of Saccharomyces cerevisiae. J Bacteriol. 1980 Aug;143(2):603–612. doi: 10.1128/jb.143.2.603-612.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. McDaniel D., Caplan A. J., Lee M. S., Adams C. C., Fishel B. R., Gross D. S., Garrard W. T. Basal-level expression of the yeast HSP82 gene requires a heat shock regulatory element. Mol Cell Biol. 1989 Nov;9(11):4789–4798. doi: 10.1128/mcb.9.11.4789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Mendenhall M. D., Jones C. A., Reed S. I. Dual regulation of the yeast CDC28-p40 protein kinase complex: cell cycle, pheromone, and nutrient limitation effects. Cell. 1987 Sep 11;50(6):927–935. doi: 10.1016/0092-8674(87)90519-8. [DOI] [PubMed] [Google Scholar]
  36. Mortimer R. K., Hawthorne D. C. Genetic Mapping in Saccharomyces IV. Mapping of Temperature-Sensitive Genes and Use of Disomic Strains in Localizing Genes. Genetics. 1973 May;74(1):33–54. doi: 10.1093/genetics/74.1.33. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Nash R., Tokiwa G., Anand S., Erickson K., Futcher A. B. The WHI1+ gene of Saccharomyces cerevisiae tethers cell division to cell size and is a cyclin homolog. EMBO J. 1988 Dec 20;7(13):4335–4346. doi: 10.1002/j.1460-2075.1988.tb03332.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Nieto-Sotelo J., Wiederrecht G., Okuda A., Parker C. S. The yeast heat shock transcription factor contains a transcriptional activation domain whose activity is repressed under nonshock conditions. Cell. 1990 Aug 24;62(4):807–817. doi: 10.1016/0092-8674(90)90124-w. [DOI] [PubMed] [Google Scholar]
  39. Nurse P. Universal control mechanism regulating onset of M-phase. Nature. 1990 Apr 5;344(6266):503–508. doi: 10.1038/344503a0. [DOI] [PubMed] [Google Scholar]
  40. Pearson W. R., Lipman D. J. Improved tools for biological sequence comparison. Proc Natl Acad Sci U S A. 1988 Apr;85(8):2444–2448. doi: 10.1073/pnas.85.8.2444. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Penn M. D., Thireos G., Greer H. Temporal analysis of general control of amino acid biosynthesis in Saccharomyces cerevisiae: role of positive regulatory genes in initiation and maintenance of mRNA derepression. Mol Cell Biol. 1984 Mar;4(3):520–528. doi: 10.1128/mcb.4.3.520. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Prendergast J. A., Murray L. E., Rowley A., Carruthers D. R., Singer R. A., Johnston G. C. Size selection identifies new genes that regulate Saccharomyces cerevisiae cell proliferation. Genetics. 1990 Jan;124(1):81–90. doi: 10.1093/genetics/124.1.81. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Reed S. I. G1-specific cyclins: in search of an S-phase-promoting factor. Trends Genet. 1991 Mar;7(3):95–99. doi: 10.1016/0168-9525(91)90279-Y. [DOI] [PubMed] [Google Scholar]
  44. Richardson H. E., Wittenberg C., Cross F., Reed S. I. An essential G1 function for cyclin-like proteins in yeast. Cell. 1989 Dec 22;59(6):1127–1133. doi: 10.1016/0092-8674(89)90768-x. [DOI] [PubMed] [Google Scholar]
  45. Rogers S., Wells R., Rechsteiner M. Amino acid sequences common to rapidly degraded proteins: the PEST hypothesis. Science. 1986 Oct 17;234(4774):364–368. doi: 10.1126/science.2876518. [DOI] [PubMed] [Google Scholar]
  46. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Singer R. A., Johnston G. C. Nalidixic acid causes a transient G1 arrest in the yeast Saccharomyces cerevisiae. Mol Gen Genet. 1979 Oct 2;176(1):37–39. doi: 10.1007/BF00334293. [DOI] [PubMed] [Google Scholar]
  48. Sorger P. K. Yeast heat shock factor contains separable transient and sustained response transcriptional activators. Cell. 1990 Aug 24;62(4):793–805. doi: 10.1016/0092-8674(90)90123-v. [DOI] [PubMed] [Google Scholar]
  49. Souciet J. L., Hubert J. C., Lacroute F. Cloning and restriction mapping of the yeast URA2 gene coding for the carbamyl phosphate synthetase aspartate-transcarbamylase complex. Mol Gen Genet. 1982;186(3):385–390. doi: 10.1007/BF00729458. [DOI] [PubMed] [Google Scholar]
  50. Souciet J. L., Nagy M., Le Gouar M., Lacroute F., Potier S. Organization of the yeast URA2 gene: identification of a defective dihydroorotase-like domain in the multifunctional carbamoylphosphate synthetase-aspartate transcarbamylase complex. Gene. 1989 Jun 30;79(1):59–70. doi: 10.1016/0378-1119(89)90092-9. [DOI] [PubMed] [Google Scholar]
  51. Struhl K., Stinchcomb D. T., Scherer S., Davis R. W. High-frequency transformation of yeast: autonomous replication of hybrid DNA molecules. Proc Natl Acad Sci U S A. 1979 Mar;76(3):1035–1039. doi: 10.1073/pnas.76.3.1035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Sudbery P. E., Goodey A. R., Carter B. L. Genes which control cell proliferation in the yeast Saccharomyces cerevisiae. Nature. 1980 Nov 27;288(5789):401–404. doi: 10.1038/288401a0. [DOI] [PubMed] [Google Scholar]
  53. Swanson M. S., Carlson M., Winston F. SPT6, an essential gene that affects transcription in Saccharomyces cerevisiae, encodes a nuclear protein with an extremely acidic amino terminus. Mol Cell Biol. 1990 Sep;10(9):4935–4941. doi: 10.1128/mcb.10.9.4935. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Swanson M. S., Malone E. A., Winston F. SPT5, an essential gene important for normal transcription in Saccharomyces cerevisiae, encodes an acidic nuclear protein with a carboxy-terminal repeat. Mol Cell Biol. 1991 Jun;11(6):3009–3019. doi: 10.1128/mcb.11.6.3009. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Thomas P. S. Hybridization of denatured RNA transferred or dotted nitrocellulose paper. Methods Enzymol. 1983;100:255–266. doi: 10.1016/0076-6879(83)00060-9. [DOI] [PubMed] [Google Scholar]
  56. Wallace R. B., Miyada C. G. Oligonucleotide probes for the screening of recombinant DNA libraries. Methods Enzymol. 1987;152:432–442. doi: 10.1016/0076-6879(87)52050-x. [DOI] [PubMed] [Google Scholar]
  57. Winston F., Chaleff D. T., Valent B., Fink G. R. Mutations affecting Ty-mediated expression of the HIS4 gene of Saccharomyces cerevisiae. Genetics. 1984 Jun;107(2):179–197. doi: 10.1093/genetics/107.2.179. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Winston F., Dollard C., Malone E. A., Clare J., Kapakos J. G., Farabaugh P., Minehart P. L. Three genes are required for trans-activation of Ty transcription in yeast. Genetics. 1987 Apr;115(4):649–656. doi: 10.1093/genetics/115.4.649. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Wittenberg C., Reed S. I. Control of the yeast cell cycle is associated with assembly/disassembly of the Cdc28 protein kinase complex. Cell. 1988 Sep 23;54(7):1061–1072. doi: 10.1016/0092-8674(88)90121-3. [DOI] [PubMed] [Google Scholar]
  60. Wittenberg C., Sugimoto K., Reed S. I. G1-specific cyclins of S. cerevisiae: cell cycle periodicity, regulation by mating pheromone, and association with the p34CDC28 protein kinase. Cell. 1990 Jul 27;62(2):225–237. doi: 10.1016/0092-8674(90)90361-h. [DOI] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES