Skip to main content
NCBI home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1996 Jun;16(6):2744–2755. doi: 10.1128/mcb.16.6.2744

The SAP, a new family of proteins, associate and function positively with the SIT4 phosphatase.

M M Luke 1, F Della Seta 1, C J Di Como 1, H Sugimoto 1, R Kobayashi 1, K T Arndt 1
PMCID: PMC231265  PMID: 8649382

Abstract

SIT4 is the catalytic subunit of a type 2A-related protein phosphatase in Saccharomyces cerevisiae that is required for G1 cyclin transcription and for bud formation. SIT4 associates with several high-molecular-mass proteins in a cell cycle-dependent fashion. We purified two SIT4-associated proteins, SAP155 and SAP190, and cloned the corresponding genes. By sequence homology, we isolated two additional SAP genes, SAP185 and SAP4. Through such an association is not yet proven for SAP4, each of SAP155, SAP185, and SAP190 physically associates with SIT4 in separate complexes. The SAPs function positively with SIT4, and by several criteria, the loss of all four SAPs is equivalent to the loss of SIT4. The data suggest that the SAPs are not functional in the absence of SIT4 and likewise that SIT4 is not functional in the absence of the SAPs. The SAPs are hyperphoshorylated in cells lacking SIT4, raising the possibility that the SAPs are substrates of SIT4. By sequence similarity, the SAPs fall into two groups, the SAP4/SAP155 group and the SAP185/SAP190 group. Overexpression of a SAP from one group does not suppress the defects due to the loss of the other group. These findings and others indicate that the SAPs have distinct functions.

Full Text

The Full Text of this article is available as a PDF (1.1 MB).

Selected References

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

  1. Arndt K. T., Styles C. A., Fink G. R. A suppressor of a HIS4 transcriptional defect encodes a protein with homology to the catalytic subunit of protein phosphatases. Cell. 1989 Feb 24;56(4):527–537. doi: 10.1016/0092-8674(89)90576-x. [DOI] [PubMed] [Google Scholar]
  2. Bender A., Pringle J. R. Use of a screen for synthetic lethal and multicopy suppressee mutants to identify two new genes involved in morphogenesis in Saccharomyces cerevisiae. Mol Cell Biol. 1991 Mar;11(3):1295–1305. doi: 10.1128/mcb.11.3.1295. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Carlson M., Botstein D. Two differentially regulated mRNAs with different 5' ends encode secreted with intracellular forms of yeast invertase. Cell. 1982 Jan;28(1):145–154. doi: 10.1016/0092-8674(82)90384-1. [DOI] [PubMed] [Google Scholar]
  4. Cohen P. The structure and regulation of protein phosphatases. Annu Rev Biochem. 1989;58:453–508. doi: 10.1146/annurev.bi.58.070189.002321. [DOI] [PubMed] [Google Scholar]
  5. Collins K., Kobayashi R., Greider C. W. Purification of Tetrahymena telomerase and cloning of genes encoding the two protein components of the enzyme. Cell. 1995 Jun 2;81(5):677–686. doi: 10.1016/0092-8674(95)90529-4. [DOI] [PubMed] [Google Scholar]
  6. Cross F. R., Hoek M., McKinney J. D., Tinkelenberg A. H. Role of Swi4 in cell cycle regulation of CLN2 expression. Mol Cell Biol. 1994 Jul;14(7):4779–4787. doi: 10.1128/mcb.14.7.4779. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Di Como C. J., Bose R., Arndt K. T. Overexpression of SIS2, which contains an extremely acidic region, increases the expression of SWI4, CLN1 and CLN2 in sit4 mutants. Genetics. 1995 Jan;139(1):95–107. doi: 10.1093/genetics/139.1.95. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Dirick L., Böhm T., Nasmyth K. Roles and regulation of Cln-Cdc28 kinases at the start of the cell cycle of Saccharomyces cerevisiae. EMBO J. 1995 Oct 2;14(19):4803–4813. doi: 10.1002/j.1460-2075.1995.tb00162.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Doseff A. I., Arndt K. T. LAS1 is an essential nuclear protein involved in cell morphogenesis and cell surface growth. Genetics. 1995 Nov;141(3):857–871. doi: 10.1093/genetics/141.3.857. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Dujon B., Alexandraki D., André B., Ansorge W., Baladron V., Ballesta J. P., Banrevi A., Bolle P. A., Bolotin-Fukuhara M., Bossier P. Complete DNA sequence of yeast chromosome XI. Nature. 1994 Jun 2;369(6479):371–378. doi: 10.1038/369371a0. [DOI] [PubMed] [Google Scholar]
  11. Feng Z. H., Wilson S. E., Peng Z. Y., Schlender K. K., Reimann E. M., Trumbly R. J. The yeast GLC7 gene required for glycogen accumulation encodes a type 1 protein phosphatase. J Biol Chem. 1991 Dec 15;266(35):23796–23801. [PubMed] [Google Scholar]
  12. Fernandez-Sarabia M. J., Sutton A., Zhong T., Arndt K. T. SIT4 protein phosphatase is required for the normal accumulation of SWI4, CLN1, CLN2, and HCS26 RNAs during late G1. Genes Dev. 1992 Dec;6(12A):2417–2428. doi: 10.1101/gad.6.12a.2417. [DOI] [PubMed] [Google Scholar]
  13. Field J., Nikawa J., Broek D., MacDonald B., Rodgers L., Wilson I. A., Lerner R. A., Wigler M. Purification of a RAS-responsive adenylyl cyclase complex from Saccharomyces cerevisiae by use of an epitope addition method. Mol Cell Biol. 1988 May;8(5):2159–2165. doi: 10.1128/mcb.8.5.2159. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Grussenmeyer T., Scheidtmann K. H., Hutchinson M. A., Eckhart W., Walter G. Complexes of polyoma virus medium T antigen and cellular proteins. Proc Natl Acad Sci U S A. 1985 Dec;82(23):7952–7954. doi: 10.1073/pnas.82.23.7952. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Healy A. M., Zolnierowicz S., Stapleton A. E., Goebl M., DePaoli-Roach A. A., Pringle J. R. CDC55, a Saccharomyces cerevisiae gene involved in cellular morphogenesis: identification, characterization, and homology to the B subunit of mammalian type 2A protein phosphatase. Mol Cell Biol. 1991 Nov;11(11):5767–5780. doi: 10.1128/mcb.11.11.5767. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kim Y. J., Francisco L., Chen G. C., Marcotte E., Chan C. S. Control of cellular morphogenesis by the Ip12/Bem2 GTPase-activating protein: possible role of protein phosphorylation. J Cell Biol. 1994 Dec;127(5):1381–1394. doi: 10.1083/jcb.127.5.1381. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kunkel T. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Proc Natl Acad Sci U S A. 1985 Jan;82(2):488–492. doi: 10.1073/pnas.82.2.488. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Lin F. C., Arndt K. T. The role of Saccharomyces cerevisiae type 2A phosphatase in the actin cytoskeleton and in entry into mitosis. EMBO J. 1995 Jun 15;14(12):2745–2759. doi: 10.1002/j.1460-2075.1995.tb07275.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Lowndes N. F., Johnson A. L., Breeden L., Johnston L. H. SWI6 protein is required for transcription of the periodically expressed DNA synthesis genes in budding yeast. Nature. 1992 Jun 11;357(6378):505–508. doi: 10.1038/357505a0. [DOI] [PubMed] [Google Scholar]
  20. Mann D. J., Dombrádi V., Cohen P. T. Drosophila protein phosphatase V functionally complements a SIT4 mutant in Saccharomyces cerevisiae and its amino-terminal region can confer this complementation to a heterologous phosphatase catalytic domain. EMBO J. 1993 Dec;12(12):4833–4842. doi: 10.1002/j.1460-2075.1993.tb06173.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Matsumoto T., Beach D. Interaction of the pim1/spi1 mitotic checkpoint with a protein phosphatase. Mol Biol Cell. 1993 Mar;4(3):337–345. doi: 10.1091/mbc.4.3.337. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Miosga T., Boles E., Schaaff-Gerstenschläger I., Schmitt S., Zimmermann F. K. Sequence and function analysis of a 9.74 kb fragment of Saccharomyces cerevisiae chromosome X including the BCK1 gene. Yeast. 1994 Nov;10(11):1481–1488. doi: 10.1002/yea.320101112. [DOI] [PubMed] [Google Scholar]
  23. Nasmyth K., Dirick L. The role of SWI4 and SWI6 in the activity of G1 cyclins in yeast. Cell. 1991 Sep 6;66(5):995–1013. doi: 10.1016/0092-8674(91)90444-4. [DOI] [PubMed] [Google Scholar]
  24. Ogas J., Andrews B. J., Herskowitz I. Transcriptional activation of CLN1, CLN2, and a putative new G1 cyclin (HCS26) by SWI4, a positive regulator of G1-specific transcription. Cell. 1991 Sep 6;66(5):1015–1026. doi: 10.1016/0092-8674(91)90445-5. [DOI] [PubMed] [Google Scholar]
  25. Ohkura H., Kinoshita N., Miyatani S., Toda T., Yanagida M. The fission yeast dis2+ gene required for chromosome disjoining encodes one of two putative type 1 protein phosphatases. Cell. 1989 Jun 16;57(6):997–1007. doi: 10.1016/0092-8674(89)90338-3. [DOI] [PubMed] [Google Scholar]
  26. Reid B. J., Hartwell L. H. Regulation of mating in the cell cycle of Saccharomyces cerevisiae. J Cell Biol. 1977 Nov;75(2 Pt 1):355–365. doi: 10.1083/jcb.75.2.355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. 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]
  28. Riles L., Dutchik J. E., Baktha A., McCauley B. K., Thayer E. C., Leckie M. P., Braden V. V., Depke J. E., Olson M. V. Physical maps of the six smallest chromosomes of Saccharomyces cerevisiae at a resolution of 2.6 kilobase pairs. Genetics. 1993 May;134(1):81–150. doi: 10.1093/genetics/134.1.81. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Ronne H., Carlberg M., Hu G. Z., Nehlin J. O. Protein phosphatase 2A in Saccharomyces cerevisiae: effects on cell growth and bud morphogenesis. Mol Cell Biol. 1991 Oct;11(10):4876–4884. doi: 10.1128/mcb.11.10.4876. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Rose M. D., Novick P., Thomas J. H., Botstein D., Fink G. R. A Saccharomyces cerevisiae genomic plasmid bank based on a centromere-containing shuttle vector. Gene. 1987;60(2-3):237–243. doi: 10.1016/0378-1119(87)90232-0. [DOI] [PubMed] [Google Scholar]
  31. Sharp P. M., Tuohy T. M., Mosurski K. R. Codon usage in yeast: cluster analysis clearly differentiates highly and lowly expressed genes. Nucleic Acids Res. 1986 Jul 11;14(13):5125–5143. doi: 10.1093/nar/14.13.5125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Shimanuki M., Kinoshita N., Ohkura H., Yoshida T., Toda T., Yanagida M. Isolation and characterization of the fission yeast protein phosphatase gene ppe1+ involved in cell shape control and mitosis. Mol Biol Cell. 1993 Mar;4(3):303–313. doi: 10.1091/mbc.4.3.303. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Sneddon A. A., Cohen P. T., Stark M. J. Saccharomyces cerevisiae protein phosphatase 2A performs an essential cellular function and is encoded by two genes. EMBO J. 1990 Dec;9(13):4339–4346. doi: 10.1002/j.1460-2075.1990.tb07883.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Stuart D., Wittenberg C. Cell cycle-dependent transcription of CLN2 is conferred by multiple distinct cis-acting regulatory elements. Mol Cell Biol. 1994 Jul;14(7):4788–4801. doi: 10.1128/mcb.14.7.4788. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Sutton A., Immanuel D., Arndt K. T. The SIT4 protein phosphatase functions in late G1 for progression into S phase. Mol Cell Biol. 1991 Apr;11(4):2133–2148. doi: 10.1128/mcb.11.4.2133. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Sutton A., Lin F., Arndt K. T. The SIT4 protein phosphatase is required in late G1 for progression into S phase. Cold Spring Harb Symp Quant Biol. 1991;56:75–81. doi: 10.1101/sqb.1991.056.01.011. [DOI] [PubMed] [Google Scholar]
  37. Thomas B. J., Rothstein R. Elevated recombination rates in transcriptionally active DNA. Cell. 1989 Feb 24;56(4):619–630. doi: 10.1016/0092-8674(89)90584-9. [DOI] [PubMed] [Google Scholar]
  38. Tyers M., Tokiwa G., Futcher B. Comparison of the Saccharomyces cerevisiae G1 cyclins: Cln3 may be an upstream activator of Cln1, Cln2 and other cyclins. EMBO J. 1993 May;12(5):1955–1968. doi: 10.1002/j.1460-2075.1993.tb05845.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Tyers M., Tokiwa G., Nash R., Futcher B. The Cln3-Cdc28 kinase complex of S. cerevisiae is regulated by proteolysis and phosphorylation. EMBO J. 1992 May;11(5):1773–1784. doi: 10.1002/j.1460-2075.1992.tb05229.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Wada K., Wada Y., Doi H., Ishibashi F., Gojobori T., Ikemura T. Codon usage tabulated from the GenBank genetic sequence data. Nucleic Acids Res. 1991 Apr 25;19 (Suppl):1981–1986. doi: 10.1093/nar/19.suppl.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. 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]
  42. van Zyl W., Huang W., Sneddon A. A., Stark M., Camier S., Werner M., Marck C., Sentenac A., Broach J. R. Inactivation of the protein phosphatase 2A regulatory subunit A results in morphological and transcriptional defects in Saccharomyces cerevisiae. Mol Cell Biol. 1992 Nov;12(11):4946–4959. doi: 10.1128/mcb.12.11.4946. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES