Skip to main content
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1996 Dec 1;24(23):4599–4607. doi: 10.1093/nar/24.23.4599

Comparative amino acid sequence analysis of the C6 zinc cluster family of transcriptional regulators.

P Schjerling 1, S Holmberg 1
PMCID: PMC146297  PMID: 8967907

Abstract

The C6 zinc cluster family of fungal regulatory proteins shares as DNA-binding motif the C6 zinc cluster, also known as the Zn(II)2Cys6 binuclear cluster. This family includes transcriptional activators like Gal4p, Leu3p, Hap1p, Put3p and Cha4p from Saccharomyces cerevisiae, qutA and amdR from Aspergillus, nit4 from Neurospora and Ntf1 from Schizosaccharomyces pombe. Seventy-nine proteins were retrieved from databases by homology to the C6 zinc cluster. All were fungal and 56 were found in the entire genome sequence of S.cerevisiae. Sequence analysis suggests that 60 of the 79 proteins possess one or more coiled-coil dimerization regions succeeding the C6 zinc cluster. Previous comparisons of Gal4p and seven other C6 zinc cluster proteins identified an additional region with weak homology. This region, designated the middle homology region (MHR), was shown to be present in 50 of the 79 proteins. Although reported mutation and deletion analyses suggest a role of MHR in regulation of protein activity, no function has yet been assigned specifically to this region. We find that the family of MHR sequences is confined to C6 zinc cluster proteins and hypothesize that one MHR function is to assist the C6 zinc cluster in DNA target discrimination.

Full Text

The Full Text of this article is available as a PDF (1.7 MB).

Selected References

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

  1. Anderson S. F., Steber C. M., Esposito R. E., Coleman J. E. UME6, a negative regulator of meiosis in Saccharomyces cerevisiae, contains a C-terminal Zn2Cys6 binuclear cluster that binds the URS1 DNA sequence in a zinc-dependent manner. Protein Sci. 1995 Sep;4(9):1832–1843. doi: 10.1002/pro.5560040918. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Baum J. A., Geever R., Giles N. H. Expression of qa-1F activator protein: identification of upstream binding sites in the qa gene cluster and localization of the DNA-binding domain. Mol Cell Biol. 1987 Mar;7(3):1256–1266. doi: 10.1128/mcb.7.3.1256. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Berger B., Wilson D. B., Wolf E., Tonchev T., Milla M., Kim P. S. Predicting coiled coils by use of pairwise residue correlations. Proc Natl Acad Sci U S A. 1995 Aug 29;92(18):8259–8263. doi: 10.1073/pnas.92.18.8259. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Carey M., Kakidani H., Leatherwood J., Mostashari F., Ptashne M. An amino-terminal fragment of GAL4 binds DNA as a dimer. J Mol Biol. 1989 Oct 5;209(3):423–432. doi: 10.1016/0022-2836(89)90007-7. [DOI] [PubMed] [Google Scholar]
  5. Chasman D. I., Kornberg R. D. GAL4 protein: purification, association with GAL80 protein, and conserved domain structure. Mol Cell Biol. 1990 Jun;10(6):2916–2923. doi: 10.1128/mcb.10.6.2916. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Coornaert D., Vissers S., André B. The pleiotropic UGA35(DURL) regulatory gene of Saccharomyces cerevisiae: cloning, sequence and identity with the DAL81 gene. Gene. 1991 Jan 15;97(2):163–171. doi: 10.1016/0378-1119(91)90048-g. [DOI] [PubMed] [Google Scholar]
  7. De Rijcke M., Seneca S., Punyammalee B., Glansdorff N., Crabeel M. Characterization of the DNA target site for the yeast ARGR regulatory complex, a sequence able to mediate repression or induction by arginine. Mol Cell Biol. 1992 Jan;12(1):68–81. doi: 10.1128/mcb.12.1.68. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Delahodde A., Delaveau T., Jacq C. Positive autoregulation of the yeast transcription factor Pdr3p, which is involved in control of drug resistance. Mol Cell Biol. 1995 Aug;15(8):4043–4051. doi: 10.1128/mcb.15.8.4043. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Delaveau T., Delahodde A., Carvajal E., Subik J., Jacq C. PDR3, a new yeast regulatory gene, is homologous to PDR1 and controls the multidrug resistance phenomenon. Mol Gen Genet. 1994 Sep 1;244(5):501–511. doi: 10.1007/BF00583901. [DOI] [PubMed] [Google Scholar]
  10. 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]
  11. Feng B., Marzluf G. A. The regulatory protein NIT4 that mediates nitrate induction in Neurospora crassa contains a complex tripartite activation domain with a novel leucine-rich, acidic motif. Curr Genet. 1996 May;29(6):537–548. doi: 10.1007/BF02426958. [DOI] [PubMed] [Google Scholar]
  12. Friden P., Reynolds C., Schimmel P. A large internal deletion converts yeast LEU3 to a constitutive transcriptional activator. Mol Cell Biol. 1989 Sep;9(9):4056–4060. doi: 10.1128/mcb.9.9.4056. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Friden P., Schimmel P. LEU3 of Saccharomyces cerevisiae activates multiple genes for branched-chain amino acid biosynthesis by binding to a common decanucleotide core sequence. Mol Cell Biol. 1988 Jul;8(7):2690–2697. doi: 10.1128/mcb.8.7.2690. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Giles N. H., Geever R. F., Asch D. K., Avalos J., Case M. E. The Wilhelmine E. Key 1989 invitational lecture. Organization and regulation of the qa (quinic acid) genes in Neurospora crassa and other fungi. J Hered. 1991 Jan-Feb;82(1):1–7. doi: 10.1093/jhered/82.1.1. [DOI] [PubMed] [Google Scholar]
  15. Giniger E., Ptashne M. Transcription in yeast activated by a putative amphipathic alpha helix linked to a DNA binding unit. Nature. 1987 Dec 17;330(6149):670–672. doi: 10.1038/330670a0. [DOI] [PubMed] [Google Scholar]
  16. Giniger E., Varnum S. M., Ptashne M. Specific DNA binding of GAL4, a positive regulatory protein of yeast. Cell. 1985 Apr;40(4):767–774. doi: 10.1016/0092-8674(85)90336-8. [DOI] [PubMed] [Google Scholar]
  17. Gray W. M., Fassler J. S. Isolation and analysis of the yeast TEA1 gene, which encodes a zinc cluster Ty enhancer-binding protein. Mol Cell Biol. 1996 Jan;16(1):347–358. doi: 10.1128/mcb.16.1.347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Gribskov M., Lüthy R., Eisenberg D. Profile analysis. Methods Enzymol. 1990;183:146–159. doi: 10.1016/0076-6879(90)83011-w. [DOI] [PubMed] [Google Scholar]
  19. Gribskov M., McLachlan A. D., Eisenberg D. Profile analysis: detection of distantly related proteins. Proc Natl Acad Sci U S A. 1987 Jul;84(13):4355–4358. doi: 10.1073/pnas.84.13.4355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Halvorsen Y. D., Nandabalan K., Dickson R. C. Identification of base and backbone contacts used for DNA sequence recognition and high-affinity binding by LAC9, a transcription activator containing a C6 zinc finger. Mol Cell Biol. 1991 Apr;11(4):1777–1784. doi: 10.1128/mcb.11.4.1777. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Henikoff S., Henikoff J. G. Amino acid substitution matrices from protein blocks. Proc Natl Acad Sci U S A. 1992 Nov 15;89(22):10915–10919. doi: 10.1073/pnas.89.22.10915. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Holmberg S., Schjerling P. Cha4p of Saccharomyces cerevisiae activates transcription via serine/threonine response elements. Genetics. 1996 Oct;144(2):467–478. doi: 10.1093/genetics/144.2.467. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Johnston M. A model fungal gene regulatory mechanism: the GAL genes of Saccharomyces cerevisiae. Microbiol Rev. 1987 Dec;51(4):458–476. doi: 10.1128/mr.51.4.458-476.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Johnston S. A., Salmeron J. M., Jr, Dincher S. S. Interaction of positive and negative regulatory proteins in the galactose regulon of yeast. Cell. 1987 Jul 3;50(1):143–146. doi: 10.1016/0092-8674(87)90671-4. [DOI] [PubMed] [Google Scholar]
  25. Katzmann D. J., Burnett P. E., Golin J., Mahé Y., Moye-Rowley W. S. Transcriptional control of the yeast PDR5 gene by the PDR3 gene product. Mol Cell Biol. 1994 Jul;14(7):4653–4661. doi: 10.1128/mcb.14.7.4653. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Keegan L., Gill G., Ptashne M. Separation of DNA binding from the transcription-activating function of a eukaryotic regulatory protein. Science. 1986 Feb 14;231(4739):699–704. doi: 10.1126/science.3080805. [DOI] [PubMed] [Google Scholar]
  27. Kim K. S., Pfeifer K., Powell L., Guarente L. Internal deletions in the yeast transcriptional activator HAP1 have opposite effects at two sequence elements. Proc Natl Acad Sci U S A. 1990 Jun;87(12):4524–4528. doi: 10.1073/pnas.87.12.4524. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Kim Y. J., Björklund S., Li Y., Sayre M. H., Kornberg R. D. A multiprotein mediator of transcriptional activation and its interaction with the C-terminal repeat domain of RNA polymerase II. Cell. 1994 May 20;77(4):599–608. doi: 10.1016/0092-8674(94)90221-6. [DOI] [PubMed] [Google Scholar]
  29. Kodadek T. How does the GAL4 transcription factor recognize the appropriate DNA binding sites in vivo? Cell Mol Biol Res. 1993;39(4):355–360. [PubMed] [Google Scholar]
  30. Krajewska W. M. Regulation of transcription in eukaryotes by DNA-binding proteins. Int J Biochem. 1992 Dec;24(12):1885–1898. doi: 10.1016/0020-711x(92)90284-8. [DOI] [PubMed] [Google Scholar]
  31. Kulmburg P., Judewicz N., Mathieu M., Lenouvel F., Sequeval D., Felenbok B. Specific binding sites for the activator protein, ALCR, in the alcA promoter of the ethanol regulon of Aspergillus nidulans. J Biol Chem. 1992 Oct 15;267(29):21146–21153. [PubMed] [Google Scholar]
  32. Kulmburg P., Sequeval D., Lenouvel F., Mathieu M., Felenbok B. Identification of the promoter region involved in the autoregulation of the transcriptional activator ALCR in Aspergillus nidulans. Mol Cell Biol. 1992 May;12(5):1932–1939. doi: 10.1128/mcb.12.5.1932. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Lechner J. A zinc finger protein, essential for chromosome segregation, constitutes a putative DNA binding subunit of the Saccharomyces cerevisiae kinetochore complex, Cbf3. EMBO J. 1994 Nov 1;13(21):5203–5211. doi: 10.1002/j.1460-2075.1994.tb06851.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Lechner J., Carbon J. A 240 kd multisubunit protein complex, CBF3, is a major component of the budding yeast centromere. Cell. 1991 Feb 22;64(4):717–725. doi: 10.1016/0092-8674(91)90501-o. [DOI] [PubMed] [Google Scholar]
  35. Leuther K. K., Salmeron J. M., Johnston S. A. Genetic evidence that an activation domain of GAL4 does not require acidity and may form a beta sheet. Cell. 1993 Feb 26;72(4):575–585. doi: 10.1016/0092-8674(93)90076-3. [DOI] [PubMed] [Google Scholar]
  36. Lodi T., Guiard B. Complex transcriptional regulation of the Saccharomyces cerevisiae CYB2 gene encoding cytochrome b2: CYP1(HAP1) activator binds to the CYB2 upstream activation site UAS1-B2. Mol Cell Biol. 1991 Jul;11(7):3762–3772. doi: 10.1128/mcb.11.7.3762. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Luo Y., Karpichev I. V., Kohanski R. A., Small G. M. Purification, identification, and properties of a Saccharomyces cerevisiae oleate-activated upstream activating sequence-binding protein that is involved in the activation of POX1. J Biol Chem. 1996 May 17;271(20):12068–12075. doi: 10.1074/jbc.271.20.12068. [DOI] [PubMed] [Google Scholar]
  38. Lupas A., Van Dyke M., Stock J. Predicting coiled coils from protein sequences. Science. 1991 May 24;252(5009):1162–1164. doi: 10.1126/science.252.5009.1162. [DOI] [PubMed] [Google Scholar]
  39. 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]
  40. Maldonado E., Ha I., Cortes P., Weis L., Reinberg D. Factors involved in specific transcription by mammalian RNA polymerase II: role of transcription factors IIA, IID, and IIB during formation of a transcription-competent complex. Mol Cell Biol. 1990 Dec;10(12):6335–6347. doi: 10.1128/mcb.10.12.6335. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Marczak J. E., Brandriss M. C. Analysis of constitutive and noninducible mutations of the PUT3 transcriptional activator. Mol Cell Biol. 1991 May;11(5):2609–2619. doi: 10.1128/mcb.11.5.2609. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Marmorstein R., Carey M., Ptashne M., Harrison S. C. DNA recognition by GAL4: structure of a protein-DNA complex. Nature. 1992 Apr 2;356(6368):408–414. doi: 10.1038/356408a0. [DOI] [PubMed] [Google Scholar]
  43. Marmorstein R., Harrison S. C. Crystal structure of a PPR1-DNA complex: DNA recognition by proteins containing a Zn2Cys6 binuclear cluster. Genes Dev. 1994 Oct 15;8(20):2504–2512. doi: 10.1101/gad.8.20.2504. [DOI] [PubMed] [Google Scholar]
  44. Nogi Y., Fukasawa T. Functional domains of a negative regulatory protein, GAL80, of Saccharomyces cerevisiae. Mol Cell Biol. 1989 Jul;9(7):3009–3017. doi: 10.1128/mcb.9.7.3009. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Pan T., Coleman J. E. GAL4 transcription factor is not a "zinc finger" but forms a Zn(II)2Cys6 binuclear cluster. Proc Natl Acad Sci U S A. 1990 Mar;87(6):2077–2081. doi: 10.1073/pnas.87.6.2077. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Parsons L. M., Davis M. A., Hynes M. J. Identification of functional regions of the positively acting regulatory gene amdR from Aspergillus nidulans. Mol Microbiol. 1992 Oct;6(20):2999–3007. doi: 10.1111/j.1365-2958.1992.tb01758.x. [DOI] [PubMed] [Google Scholar]
  47. Pfeifer K., Kim K. S., Kogan S., Guarente L. Functional dissection and sequence of yeast HAP1 activator. Cell. 1989 Jan 27;56(2):291–301. doi: 10.1016/0092-8674(89)90903-3. [DOI] [PubMed] [Google Scholar]
  48. Reece R. J., Ptashne M. Determinants of binding-site specificity among yeast C6 zinc cluster proteins. Science. 1993 Aug 13;261(5123):909–911. doi: 10.1126/science.8346441. [DOI] [PubMed] [Google Scholar]
  49. Remboutsika E., Kohlhaw G. B. Molecular architecture of a Leu3p-DNA complex in solution: a biochemical approach. Mol Cell Biol. 1994 Aug;14(8):5547–5557. doi: 10.1128/mcb.14.8.5547. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Richardson I. B., Katz M. E., Hynes M. J. Molecular characterization of the lam locus and sequences involved in regulation by the AmdR protein of Aspergillus nidulans. Mol Cell Biol. 1992 Jan;12(1):337–346. doi: 10.1128/mcb.12.1.337. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Rottensteiner H., Kal A. J., Filipits M., Binder M., Hamilton B., Tabak H. F., Ruis H. Pip2p: a transcriptional regulator of peroxisome proliferation in the yeast Saccharomyces cerevisiae. EMBO J. 1996 Jun 17;15(12):2924–2934. [PMC free article] [PubMed] [Google Scholar]
  52. Roy A., Exinger F., Losson R. cis- and trans-acting regulatory elements of the yeast URA3 promoter. Mol Cell Biol. 1990 Oct;10(10):5257–5270. doi: 10.1128/mcb.10.10.5257. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Sakurai H., Hiraoka Y., Fukasawa T. Yeast GAL11 protein is a distinctive type transcription factor that enhances basal transcription in vitro. Proc Natl Acad Sci U S A. 1993 Sep 15;90(18):8382–8386. doi: 10.1073/pnas.90.18.8382. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Sequeval D., Felenbok B. Relationship between zinc content and DNA-binding activity of the DNA-binding motif of the transcription factor ALCR in Aspergillus nidulans. Mol Gen Genet. 1994 Jan;242(1):33–39. doi: 10.1007/BF00277345. [DOI] [PubMed] [Google Scholar]
  55. Siddiqui A. H., Brandriss M. C. The Saccharomyces cerevisiae PUT3 activator protein associates with proline-specific upstream activation sequences. Mol Cell Biol. 1989 Nov;9(11):4706–4712. doi: 10.1128/mcb.9.11.4706. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Stone G., Sadowski I. GAL4 is regulated by a glucose-responsive functional domain. EMBO J. 1993 Apr;12(4):1375–1385. doi: 10.1002/j.1460-2075.1993.tb05782.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Strich R., Surosky R. T., Steber C., Dubois E., Messenguy F., Esposito R. E. UME6 is a key regulator of nitrogen repression and meiotic development. Genes Dev. 1994 Apr 1;8(7):796–810. doi: 10.1101/gad.8.7.796. [DOI] [PubMed] [Google Scholar]
  58. Suárez T., de Queiroz M. V., Oestreicher N., Scazzocchio C. The sequence and binding specificity of UaY, the specific regulator of the purine utilization pathway in Aspergillus nidulans, suggest an evolutionary relationship with the PPR1 protein of Saccharomyces cerevisiae. EMBO J. 1995 Apr 3;14(7):1453–1467. doi: 10.1002/j.1460-2075.1995.tb07132.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Sze J. Y., Kohlhaw G. B. Purification and structural characterization of transcriptional regulator Leu3 of yeast. J Biol Chem. 1993 Feb 5;268(4):2505–2512. [PubMed] [Google Scholar]
  60. Sze J. Y., Remboutsika E., Kohlhaw G. B. Transcriptional regulator Leu3 of Saccharomyces cerevisiae: separation of activator and repressor functions. Mol Cell Biol. 1993 Sep;13(9):5702–5709. doi: 10.1128/mcb.13.9.5702. [DOI] [PMC free article] [PubMed] [Google Scholar]
  61. Timmerman J. E., Guiard B., Shechter E., Delsuc M. A., Lallemand J. Y., Gervais M. The DNA-binding domain of the yeast Saccharomyces cerevisiae CYP1(HAP1) transcription factor possesses two zinc ions which are complexed in a zinc cluster. Eur J Biochem. 1994 Oct 15;225(2):593–599. doi: 10.1111/j.1432-1033.1994.00593.x. [DOI] [PubMed] [Google Scholar]
  62. Van Hoy M., Leuther K. K., Kodadek T., Johnston S. A. The acidic activation domains of the GCN4 and GAL4 proteins are not alpha helical but form beta sheets. Cell. 1993 Feb 26;72(4):587–594. doi: 10.1016/0092-8674(93)90077-4. [DOI] [PubMed] [Google Scholar]
  63. Vashee S., Xu H., Johnston S. A., Kodadek T. How do "Zn2 cys6" proteins distinguish between similar upstream activation sites? Comparison of the DNA-binding specificity of the GAL4 protein in vitro and in vivo. J Biol Chem. 1993 Nov 25;268(33):24699–24706. [PubMed] [Google Scholar]
  64. Witte M. M., Dickson R. C. Cysteine residues in the zinc finger and amino acids adjacent to the finger are necessary for DNA binding by the LAC9 regulatory protein of Kluyveromyces lactis. Mol Cell Biol. 1988 Sep;8(9):3726–3733. doi: 10.1128/mcb.8.9.3726. [DOI] [PMC free article] [PubMed] [Google Scholar]
  65. Witte M. M., Dickson R. C. The C6 zinc finger and adjacent amino acids determine DNA-binding specificity and affinity in the yeast activator proteins LAC9 and PPR1. Mol Cell Biol. 1990 Oct;10(10):5128–5137. doi: 10.1128/mcb.10.10.5128. [DOI] [PMC free article] [PubMed] [Google Scholar]
  66. Zhang L., Bermingham-McDonogh O., Turcotte B., Guarente L. Antibody-promoted dimerization bypasses the regulation of DNA binding by the heme domain of the yeast transcriptional activator HAP1. Proc Natl Acad Sci U S A. 1993 Apr 1;90(7):2851–2855. doi: 10.1073/pnas.90.7.2851. [DOI] [PMC free article] [PubMed] [Google Scholar]
  67. Zhang L., Guarente L. The yeast activator HAP1--a GAL4 family member--binds DNA in a directly repeated orientation. Genes Dev. 1994 Sep 1;8(17):2110–2119. doi: 10.1101/gad.8.17.2110. [DOI] [PubMed] [Google Scholar]
  68. Zhou K. M., Bai Y. L., Kohlhaw G. B. Yeast regulatory protein LEU3: a structure-function analysis. Nucleic Acids Res. 1990 Jan 25;18(2):291–298. doi: 10.1093/nar/18.2.291. [DOI] [PMC free article] [PubMed] [Google Scholar]
  69. Zhou K. M., Kohlhaw G. B. Transcriptional activator LEU3 of yeast. Mapping of the transcriptional activation function and significance of activation domain tryptophans. J Biol Chem. 1990 Oct 15;265(29):17409–17412. [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES