Skip to main content
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1991 Aug;11(8):4135–4146. doi: 10.1128/mcb.11.8.4135

A negative regulator of HO transcription, SIN1 (SPT2), is a nonspecific DNA-binding protein related to HMG1.

W Kruger 1, I Herskowitz 1
PMCID: PMC361230  PMID: 2072912

Abstract

The SIN1 gene was initially identified because mutations in SIN1 bypass the need for SWI1 to activate transcription of the yeast HO gene. We show here that transcription of HO in swi1 sin1 cells efficiently utilizes the normal start site. We have cloned SIN1 and found that it is identical to the previously identified gene SPT2, mutations in which allow transcription from certain mutated regulatory regions. The predicted SIN1/SPT2 protein has a distinctive amino acid composition (45% charged residues, 25% basic and 20% acidic) and has similarity to the mammalian HMG1 protein, a nonhistone component of chromatin. We show that SIN1 is concentrated in the nucleus and binds to DNA with little or no sequence specificity in vitro. It thus exhibits properties of an HMG protein. Addition of random DNA segments to a test promoter alters regulation by SIN1 in a manner similar to addition of a segment from the HO upstream region. Functional analysis of certain SIN1 mutations suggests that SIN1 may be part of a multiprotein complex. On the basis of these results, we propose that SIN1 is a nonhistone component of chromatin which creates the proper context for transcription. Because sin1 mutants exhibit increased loss of chromosome III, SIN1 may also play a role in fidelity of chromosome segregation.

Full text

PDF
4135

Images in this article

Selected References

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

  1. Andrews B. J., Herskowitz I. Identification of a DNA binding factor involved in cell-cycle control of the yeast HO gene. Cell. 1989 Apr 7;57(1):21–29. doi: 10.1016/0092-8674(89)90168-2. [DOI] [PubMed] [Google Scholar]
  2. Breeden L., Nasmyth K. Cell cycle control of the yeast HO gene: cis- and trans-acting regulators. Cell. 1987 Feb 13;48(3):389–397. doi: 10.1016/0092-8674(87)90190-5. [DOI] [PubMed] [Google Scholar]
  3. Butler A. P., Mardian J. K., Olins D. E. Nonhistone chromosomal protein HMG 1 interactions with DNA. Fluorescence and thermal denaturation studies. J Biol Chem. 1985 Sep 5;260(19):10613–10620. [PubMed] [Google Scholar]
  4. Dayhoff M. O., Barker W. C., Hunt L. T. Establishing homologies in protein sequences. Methods Enzymol. 1983;91:524–545. doi: 10.1016/s0076-6879(83)91049-2. [DOI] [PubMed] [Google Scholar]
  5. Desplan C., Theis J., O'Farrell P. H. The Drosophila developmental gene, engrailed, encodes a sequence-specific DNA binding activity. Nature. 1985 Dec 19;318(6047):630–635. doi: 10.1038/318630a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. 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]
  7. 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]
  8. 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]
  9. Fleer R., Nicolet C. M., Pure G. A., Friedberg E. C. RAD4 gene of Saccharomyces cerevisiae: molecular cloning and partial characterization of a gene that is inactivated in Escherichia coli. Mol Cell Biol. 1987 Mar;7(3):1180–1192. doi: 10.1128/mcb.7.3.1180. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Guarente L., Hoar E. Upstream activation sites of the CYC1 gene of Saccharomyces cerevisiae are active when inverted but not when placed downstream of the "TATA box". Proc Natl Acad Sci U S A. 1984 Dec;81(24):7860–7864. doi: 10.1073/pnas.81.24.7860. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Haggren W., Kolodrubetz D. The Saccharomyces cerevisiae ACP2 gene encodes an essential HMG1-like protein. Mol Cell Biol. 1988 Mar;8(3):1282–1289. doi: 10.1128/mcb.8.3.1282. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hall M. N., Hereford L., Herskowitz I. Targeting of E. coli beta-galactosidase to the nucleus in yeast. Cell. 1984 Apr;36(4):1057–1065. doi: 10.1016/0092-8674(84)90055-2. [DOI] [PubMed] [Google Scholar]
  13. Herskowitz I. A regulatory hierarchy for cell specialization in yeast. Nature. 1989 Dec 14;342(6251):749–757. doi: 10.1038/342749a0. [DOI] [PubMed] [Google Scholar]
  14. Hicks J. B., Herskowitz I. Interconversion of Yeast Mating Types I. Direct Observations of the Action of the Homothallism (HO) Gene. Genetics. 1976 Jun;83(2):245–258. doi: 10.1093/genetics/83.2.245. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hirschman J. E., Durbin K. J., Winston F. Genetic evidence for promoter competition in Saccharomyces cerevisiae. Mol Cell Biol. 1988 Nov;8(11):4608–4615. doi: 10.1128/mcb.8.11.4608. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Jantzen H. M., Admon A., Bell S. P., Tjian R. Nucleolar transcription factor hUBF contains a DNA-binding motif with homology to HMG proteins. Nature. 1990 Apr 26;344(6269):830–836. doi: 10.1038/344830a0. [DOI] [PubMed] [Google Scholar]
  17. Jensen R., Sprague G. F., Jr, Herskowitz I. Regulation of yeast mating-type interconversion: feedback control of HO gene expression by the mating-type locus. Proc Natl Acad Sci U S A. 1983 May;80(10):3035–3039. doi: 10.1073/pnas.80.10.3035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Johnson A. D., Herskowitz I. A repressor (MAT alpha 2 Product) and its operator control expression of a set of cell type specific genes in yeast. Cell. 1985 Aug;42(1):237–247. doi: 10.1016/s0092-8674(85)80119-7. [DOI] [PubMed] [Google Scholar]
  19. Kleid D. G., Yansura D., Small B., Dowbenko D., Moore D. M., Grubman M. J., McKercher P. D., Morgan D. O., Robertson B. H., Bachrach H. L. Cloned viral protein vaccine for foot-and-mouth disease: responses in cattle and swine. Science. 1981 Dec 4;214(4525):1125–1129. doi: 10.1126/science.6272395. [DOI] [PubMed] [Google Scholar]
  20. Kolodrubetz D., Burgum A. Duplicated NHP6 genes of Saccharomyces cerevisiae encode proteins homologous to bovine high mobility group protein 1. J Biol Chem. 1990 Feb 25;265(6):3234–3239. [PubMed] [Google Scholar]
  21. Kolodrubetz D., Haggren W., Burgum A. Amino-terminal sequence of a Saccharomyces cerevisiae nuclear protein, NHP6, shows significant identity to bovine HMG1. FEBS Lett. 1988 Sep 26;238(1):175–179. doi: 10.1016/0014-5793(88)80251-5. [DOI] [PubMed] [Google Scholar]
  22. Kostriken R., Heffron F. The product of the HO gene is a nuclease: purification and characterization of the enzyme. Cold Spring Harb Symp Quant Biol. 1984;49:89–96. doi: 10.1101/sqb.1984.049.01.012. [DOI] [PubMed] [Google Scholar]
  23. Kuehl L., Salmond B., Tran L. Concentrations of high-mobility-group proteins in the nucleus and cytoplasm of several rat tissues. J Cell Biol. 1984 Aug;99(2):648–654. doi: 10.1083/jcb.99.2.648. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Lipman D. J., Pearson W. R. Rapid and sensitive protein similarity searches. Science. 1985 Mar 22;227(4693):1435–1441. doi: 10.1126/science.2983426. [DOI] [PubMed] [Google Scholar]
  25. Meeks-Wagner D., Wood J. S., Garvik B., Hartwell L. H. Isolation of two genes that affect mitotic chromosome transmission in S. cerevisiae. Cell. 1986 Jan 17;44(1):53–63. doi: 10.1016/0092-8674(86)90484-8. [DOI] [PubMed] [Google Scholar]
  26. Miller A. M., MacKay V. L., Nasmyth K. A. Identification and comparison of two sequence elements that confer cell-type specific transcription in yeast. Nature. 1985 Apr 18;314(6012):598–603. doi: 10.1038/314598a0. [DOI] [PubMed] [Google Scholar]
  27. Nasmyth K. A repetitive DNA sequence that confers cell-cycle START (CDC28)-dependent transcription of the HO gene in yeast. Cell. 1985 Aug;42(1):225–235. doi: 10.1016/s0092-8674(85)80118-5. [DOI] [PubMed] [Google Scholar]
  28. Nasmyth K. At least 1400 base pairs of 5'-flanking DNA is required for the correct expression of the HO gene in yeast. Cell. 1985 Aug;42(1):213–223. doi: 10.1016/s0092-8674(85)80117-3. [DOI] [PubMed] [Google Scholar]
  29. Nasmyth K. Molecular analysis of a cell lineage. Nature. 1983 Apr 21;302(5910):670–676. doi: 10.1038/302670a0. [DOI] [PubMed] [Google Scholar]
  30. Nonet M. L., Young R. A. Intragenic and extragenic suppressors of mutations in the heptapeptide repeat domain of Saccharomyces cerevisiae RNA polymerase II. Genetics. 1989 Dec;123(4):715–724. doi: 10.1093/genetics/123.4.715. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Nonet M., Sweetser D., Young R. A. Functional redundancy and structural polymorphism in the large subunit of RNA polymerase II. Cell. 1987 Sep 11;50(6):909–915. doi: 10.1016/0092-8674(87)90517-4. [DOI] [PubMed] [Google Scholar]
  32. Orr-Weaver T. L., Szostak J. W., Rothstein R. J. Genetic applications of yeast transformation with linear and gapped plasmids. Methods Enzymol. 1983;101:228–245. doi: 10.1016/0076-6879(83)01017-4. [DOI] [PubMed] [Google Scholar]
  33. Pentecost B. T., Wright J. M., Dixon G. H. Isolation and sequence of cDNA clones coding for a member of the family of high mobility group proteins (HMG-T) in trout and analysis of HMG-T-mRNA's in trout tissues. Nucleic Acids Res. 1985 Jul 11;13(13):4871–4888. doi: 10.1093/nar/13.13.4871. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Peterson C. L., Kruger W., Herskowitz I. A functional interaction between the C-terminal domain of RNA polymerase II and the negative regulator SIN1. Cell. 1991 Mar 22;64(6):1135–1143. doi: 10.1016/0092-8674(91)90268-4. [DOI] [PubMed] [Google Scholar]
  35. Roeder G. S., Beard C., Smith M., Keranen S. Isolation and characterization of the SPT2 gene, a negative regulator of Ty-controlled yeast gene expression. Mol Cell Biol. 1985 Jul;5(7):1543–1553. doi: 10.1128/mcb.5.7.1543. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Roeder G. S., Fink G. R. DNA rearrangements associated with a transposable element in yeast. Cell. 1980 Aug;21(1):239–249. doi: 10.1016/0092-8674(80)90131-2. [DOI] [PubMed] [Google Scholar]
  37. Rotenberg M. O., Woolford J. L., Jr Tripartite upstream promoter element essential for expression of Saccharomyces cerevisiae ribosomal protein genes. Mol Cell Biol. 1986 Feb;6(2):674–687. doi: 10.1128/mcb.6.2.674. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Rotenberg M. O., Woolford J. L., Jr Tripartite upstream promoter element essential for expression of Saccharomyces cerevisiae ribosomal protein genes. Mol Cell Biol. 1986 Feb;6(2):674–687. doi: 10.1128/mcb.6.2.674. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Roth S. Y., Schulman I. G., Cook R. G., Allis C. D. The complete amino acid sequence of an HMG-like protein isolated from the macronucleus of Tetrahymena. Nucleic Acids Res. 1987 Oct 12;15(19):8112–8112. doi: 10.1093/nar/15.19.8112. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Russell D. W., Jensen R., Zoller M. J., Burke J., Errede B., Smith M., Herskowitz I. Structure of the Saccharomyces cerevisiae HO gene and analysis of its upstream regulatory region. Mol Cell Biol. 1986 Dec;6(12):4281–4294. doi: 10.1128/mcb.6.12.4281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Schröter H., Bode J. The binding sites for large and small high-mobility-group (HMG) proteins. Studies on HMG-nucleosome interactions in vitro. Eur J Biochem. 1982 Oct;127(2):429–436. doi: 10.1111/j.1432-1033.1982.tb06890.x. [DOI] [PubMed] [Google Scholar]
  42. Schwartz D. C., Cantor C. R. Separation of yeast chromosome-sized DNAs by pulsed field gradient gel electrophoresis. Cell. 1984 May;37(1):67–75. doi: 10.1016/0092-8674(84)90301-5. [DOI] [PubMed] [Google Scholar]
  43. Silverman S. J., Fink G. R. Effects of Ty insertions on HIS4 transcription in Saccharomyces cerevisiae. Mol Cell Biol. 1984 Jul;4(7):1246–1251. doi: 10.1128/mcb.4.7.1246. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Simchen G., Winston F., Styles C. A., Fink G. R. Ty-mediated gene expression of the LYS2 and HIS4 genes of Saccharomyces cerevisiae is controlled by the same SPT genes. Proc Natl Acad Sci U S A. 1984 Apr;81(8):2431–2434. doi: 10.1073/pnas.81.8.2431. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Smith M. M., Andrésson O. S. DNA sequences of yeast H3 and H4 histone genes from two non-allelic gene sets encode identical H3 and H4 proteins. J Mol Biol. 1983 Sep 25;169(3):663–690. doi: 10.1016/s0022-2836(83)80164-8. [DOI] [PubMed] [Google Scholar]
  46. Snyder M. The SPA2 protein of yeast localizes to sites of cell growth. J Cell Biol. 1989 Apr;108(4):1419–1429. doi: 10.1083/jcb.108.4.1419. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Sprague G. F., Jr, Herskowitz I. Control of yeast cell type by the mating type locus. I. Identification and control of expression of the a-specific gene BAR1. J Mol Biol. 1981 Dec 5;153(2):305–321. doi: 10.1016/0022-2836(81)90280-1. [DOI] [PubMed] [Google Scholar]
  48. Stern M., Jensen R., Herskowitz I. Five SWI genes are required for expression of the HO gene in yeast. J Mol Biol. 1984 Oct 5;178(4):853–868. doi: 10.1016/0022-2836(84)90315-2. [DOI] [PubMed] [Google Scholar]
  49. Sternberg P. W., Stern M. J., Clark I., Herskowitz I. Activation of the yeast HO gene by release from multiple negative controls. Cell. 1987 Feb 27;48(4):567–577. doi: 10.1016/0092-8674(87)90235-2. [DOI] [PubMed] [Google Scholar]
  50. Strathern J., Hicks J., Herskowitz I. Control of cell type in yeast by the mating type locus. The alpha 1-alpha 2 hypothesis. J Mol Biol. 1981 Apr 15;147(3):357–372. doi: 10.1016/0022-2836(81)90488-5. [DOI] [PubMed] [Google Scholar]
  51. Tsuda K., Kikuchi M., Mori K., Waga S., Yoshida M. Primary structure of non-histone protein HMG1 revealed by the nucleotide sequence. Biochemistry. 1988 Aug 9;27(16):6159–6163. doi: 10.1021/bi00416a050. [DOI] [PubMed] [Google Scholar]
  52. Weber S., Isenberg I. High mobility group proteins of Saccharomyces cerevisiae. Biochemistry. 1980 May 13;19(10):2236–2240. doi: 10.1021/bi00551a037. [DOI] [PubMed] [Google Scholar]
  53. 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]

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

RESOURCES