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. 1989 Oct;9(10):4304–4311. doi: 10.1128/mcb.9.10.4304

Evolutionary conservation of homeodomain-binding sites and other sequences upstream and within the major transcription unit of the Drosophila segmentation gene engrailed.

J A Kassis 1, C Desplan 1, D K Wright 1, P H O'Farrell 1
PMCID: PMC362511  PMID: 2573829

Abstract

The engrailed (en) gene functions throughout Drosophila development and is expressed in a succession of intricate spatial patterns as development proceeds. Normal en function relies on an extremely large cis-acting regulatory region (70 kilobases). We are using evolutionary conservation to help identify en sequences important in regulating patterned expression. Sequence comparison of 2.6 kilobases upstream of the en coding region of D. melanogaster and D. virilis (estimated divergence time, 60 million years) showed that 30% of this DNA occurs in islands of near perfect sequence conservation. One of these conserved islands contains binding sites for homeodomain-containing proteins. It has been shown genetically that homeodomain-containing proteins regulate en expression. Our data suggested that this regulation may be direct. The remaining conserved islands may contain binding sites for other regulatory proteins.

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Selected References

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  1. Bender A., Sprague G. F., Jr MAT alpha 1 protein, a yeast transcription activator, binds synergistically with a second protein to a set of cell-type-specific genes. Cell. 1987 Aug 28;50(5):681–691. doi: 10.1016/0092-8674(87)90326-6. [DOI] [PubMed] [Google Scholar]
  2. Beverley S. M., Wilson A. C. Molecular evolution in Drosophila and the higher Diptera II. A time scale for fly evolution. J Mol Evol. 1984;21(1):1–13. doi: 10.1007/BF02100622. [DOI] [PubMed] [Google Scholar]
  3. Biggin M. D., Tjian R. Transcription factors that activate the Ultrabithorax promoter in developmentally staged extracts. Cell. 1988 Jun 3;53(5):699–711. doi: 10.1016/0092-8674(88)90088-8. [DOI] [PubMed] [Google Scholar]
  4. Blackman R. K., Meselson M. Interspecific nucleotide sequence comparisons used to identify regulatory and structural features of the Drosophila hsp82 gene. J Mol Biol. 1986 Apr 20;188(4):499–515. doi: 10.1016/s0022-2836(86)80001-8. [DOI] [PubMed] [Google Scholar]
  5. Bray S. J., Hirsh J. The Drosophila virilis dopa decarboxylase gene is developmentally regulated when integrated into Drosophila melanogaster. EMBO J. 1986 Sep;5(9):2305–2311. doi: 10.1002/j.1460-2075.1986.tb04498.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bray S. J., Johnson W. A., Hirsh J., Heberlein U., Tjian R. A cis-acting element and associated binding factor required for CNS expression of the Drosophila melanogaster dopa decarboxylase gene. EMBO J. 1988 Jan;7(1):177–188. doi: 10.1002/j.1460-2075.1988.tb02798.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Briggs M. R., Kadonaga J. T., Bell S. P., Tjian R. Purification and biochemical characterization of the promoter-specific transcription factor, Sp1. Science. 1986 Oct 3;234(4772):47–52. doi: 10.1126/science.3529394. [DOI] [PubMed] [Google Scholar]
  8. Brower D. L. Engrailed gene expression in Drosophila imaginal discs. EMBO J. 1986 Oct;5(10):2649–2656. doi: 10.1002/j.1460-2075.1986.tb04547.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. 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]
  10. Desplan C., Theis J., O'Farrell P. H. The sequence specificity of homeodomain-DNA interaction. Cell. 1988 Sep 23;54(7):1081–1090. doi: 10.1016/0092-8674(88)90123-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. DiNardo S., Kuner J. M., Theis J., O'Farrell P. H. Development of embryonic pattern in D. melanogaster as revealed by accumulation of the nuclear engrailed protein. Cell. 1985 Nov;43(1):59–69. doi: 10.1016/0092-8674(85)90012-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. DiNardo S., O'Farrell P. H. Establishment and refinement of segmental pattern in the Drosophila embryo: spatial control of engrailed expression by pair-rule genes. Genes Dev. 1987 Dec;1(10):1212–1225. doi: 10.1101/gad.1.10.1212. [DOI] [PubMed] [Google Scholar]
  13. DiNardo S., Sher E., Heemskerk-Jongens J., Kassis J. A., O'Farrell P. H. Two-tiered regulation of spatially patterned engrailed gene expression during Drosophila embryogenesis. Nature. 1988 Apr 14;332(6165):604–609. doi: 10.1038/332604a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dorn A., Bollekens J., Staub A., Benoist C., Mathis D. A multiplicity of CCAAT box-binding proteins. Cell. 1987 Sep 11;50(6):863–872. doi: 10.1016/0092-8674(87)90513-7. [DOI] [PubMed] [Google Scholar]
  15. Drees B., Ali Z., Soeller W. C., Coleman K. G., Poole S. J., Kornberg T. The transcription unit of the Drosophila engrailed locus: an unusually small portion of a 70,000 bp gene. EMBO J. 1987 Sep;6(9):2803–2809. doi: 10.1002/j.1460-2075.1987.tb02576.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Duboule D., Haenlin M., Galliot B., Mohier E. DNA sequences homologous to the Drosophila opa repeat are present in murine mRNAs that are differentially expressed in fetuses and adult tissues. Mol Cell Biol. 1987 May;7(5):2003–2006. doi: 10.1128/mcb.7.5.2003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Dynan W. S., Tjian R. Control of eukaryotic messenger RNA synthesis by sequence-specific DNA-binding proteins. 1985 Aug 29-Sep 4Nature. 316(6031):774–778. doi: 10.1038/316774a0. [DOI] [PubMed] [Google Scholar]
  18. Efstratiadis A., Posakony J. W., Maniatis T., Lawn R. M., O'Connell C., Spritz R. A., DeRiel J. K., Forget B. G., Weissman S. M., Slightom J. L. The structure and evolution of the human beta-globin gene family. Cell. 1980 Oct;21(3):653–668. doi: 10.1016/0092-8674(80)90429-8. [DOI] [PubMed] [Google Scholar]
  19. Fjose A., McGinnis W. J., Gehring W. J. Isolation of a homoeo box-containing gene from the engrailed region of Drosophila and the spatial distribution of its transcripts. Nature. 1985 Jan 24;313(6000):284–289. doi: 10.1038/313284a0. [DOI] [PubMed] [Google Scholar]
  20. Fromental C., Kanno M., Nomiyama H., Chambon P. Cooperativity and hierarchical levels of functional organization in the SV40 enhancer. Cell. 1988 Sep 23;54(7):943–953. doi: 10.1016/0092-8674(88)90109-2. [DOI] [PubMed] [Google Scholar]
  21. Henikoff S., Eghtedarzadeh M. K. Conserved arrangement of nested genes at the Drosophila Gart locus. Genetics. 1987 Dec;117(4):711–725. doi: 10.1093/genetics/117.4.711. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Henikoff S. Unidirectional digestion with exonuclease III creates targeted breakpoints for DNA sequencing. Gene. 1984 Jun;28(3):351–359. doi: 10.1016/0378-1119(84)90153-7. [DOI] [PubMed] [Google Scholar]
  23. Hoey T., Levine M. Divergent homeo box proteins recognize similar DNA sequences in Drosophila. Nature. 1988 Apr 28;332(6167):858–861. doi: 10.1038/332858a0. [DOI] [PubMed] [Google Scholar]
  24. Howard K., Ingham P. Regulatory interactions between the segmentation genes fushi tarazu, hairy, and engrailed in the Drosophila blastoderm. Cell. 1986 Mar 28;44(6):949–957. doi: 10.1016/0092-8674(86)90018-8. [DOI] [PubMed] [Google Scholar]
  25. Ingham P. W. The molecular genetics of embryonic pattern formation in Drosophila. Nature. 1988 Sep 1;335(6185):25–34. doi: 10.1038/335025a0. [DOI] [PubMed] [Google Scholar]
  26. Jaynes J. B., O'Farrell P. H. Activation and repression of transcription by homoeodomain-containing proteins that bind a common site. Nature. 1988 Dec 22;336(6201):744–749. doi: 10.1038/336744a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Kassis J. A., Poole S. J., Wright D. K., O'Farrell P. H. Sequence conservation in the protein coding and intron regions of the engrailed transcription unit. EMBO J. 1986 Dec 20;5(13):3583–3589. doi: 10.1002/j.1460-2075.1986.tb04686.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Kassis J. A., Wong M. L., O'Farrell P. H. Electron microscopic heteroduplex mapping identifies regions of the engrailed locus that are conserved between Drosophila melanogaster and Drosophila virilis. Mol Cell Biol. 1985 Dec;5(12):3600–3609. doi: 10.1128/mcb.5.12.3600. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Kidd S., Lockett T. J., Young M. W. The Notch locus of Drosophila melanogaster. Cell. 1983 Sep;34(2):421–433. doi: 10.1016/0092-8674(83)90376-8. [DOI] [PubMed] [Google Scholar]
  30. Kimmel A. R., Firtel R. A. Sequence organization and developmental expression of an interspersed, repetitive element and associated single-copy DNA sequences in Dictyostelium discoideum. Mol Cell Biol. 1985 Aug;5(8):2123–2130. doi: 10.1128/mcb.5.8.2123. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Ko H. S., Fast P., McBride W., Staudt L. M. A human protein specific for the immunoglobulin octamer DNA motif contains a functional homeobox domain. Cell. 1988 Oct 7;55(1):135–144. doi: 10.1016/0092-8674(88)90016-5. [DOI] [PubMed] [Google Scholar]
  32. Kornberg T., Sidén I., O'Farrell P., Simon M. The engrailed locus of Drosophila: in situ localization of transcripts reveals compartment-specific expression. Cell. 1985 Jan;40(1):45–53. doi: 10.1016/0092-8674(85)90307-1. [DOI] [PubMed] [Google Scholar]
  33. Kuner J. M., Nakanishi M., Ali Z., Drees B., Gustavson E., Theis J., Kauvar L., Kornberg T., O'Farrell P. H. Molecular cloning of engrailed: a gene involved in the development of pattern in Drosophila melanogaster. Cell. 1985 Aug;42(1):309–316. doi: 10.1016/s0092-8674(85)80126-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Laughon A., Boulet A. M., Bermingham J. R., Jr, Laymon R. A., Scott M. P. Structure of transcripts from the homeotic Antennapedia gene of Drosophila melanogaster: two promoters control the major protein-coding region. Mol Cell Biol. 1986 Dec;6(12):4676–4689. doi: 10.1128/mcb.6.12.4676. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. McGinnis W., Levine M. S., Hafen E., Kuroiwa A., Gehring W. J. A conserved DNA sequence in homoeotic genes of the Drosophila Antennapedia and bithorax complexes. 1984 Mar 29-Apr 4Nature. 308(5958):428–433. doi: 10.1038/308428a0. [DOI] [PubMed] [Google Scholar]
  36. Minty A., Kedes L. Upstream regions of the human cardiac actin gene that modulate its transcription in muscle cells: presence of an evolutionarily conserved repeated motif. Mol Cell Biol. 1986 Jun;6(6):2125–2136. doi: 10.1128/mcb.6.6.2125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Norrander J., Kempe T., Messing J. Construction of improved M13 vectors using oligodeoxynucleotide-directed mutagenesis. Gene. 1983 Dec;26(1):101–106. doi: 10.1016/0378-1119(83)90040-9. [DOI] [PubMed] [Google Scholar]
  38. O'Farrell P. H., Desplan C., DiNardo S., Kassis J. A., Kuner J. M., Sher E., Theis J., Wright D. Embryonic pattern in Drosophila: the spatial distribution and sequence-specific DNA binding of engrailed protein. Cold Spring Harb Symp Quant Biol. 1985;50:235–242. doi: 10.1101/sqb.1985.050.01.030. [DOI] [PubMed] [Google Scholar]
  39. Perkins K. K., Dailey G. M., Tjian R. In vitro analysis of the Antennapedia P2 promoter: identification of a new Drosophila transcription factor. Genes Dev. 1988 Dec;2(12A):1615–1626. doi: 10.1101/gad.2.12a.1615. [DOI] [PubMed] [Google Scholar]
  40. Perler F., Efstratiadis A., Lomedico P., Gilbert W., Kolodner R., Dodgson J. The evolution of genes: the chicken preproinsulin gene. Cell. 1980 Jun;20(2):555–566. doi: 10.1016/0092-8674(80)90641-8. [DOI] [PubMed] [Google Scholar]
  41. Philippe J., Drucker D. J., Knepel W., Jepeal L., Misulovin Z., Habener J. F. Alpha-cell-specific expression of the glucagon gene is conferred to the glucagon promoter element by the interactions of DNA-binding proteins. Mol Cell Biol. 1988 Nov;8(11):4877–4888. doi: 10.1128/mcb.8.11.4877. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Poole S. J., Kauvar L. M., Drees B., Kornberg T. The engrailed locus of Drosophila: structural analysis of an embryonic transcript. Cell. 1985 Jan;40(1):37–43. doi: 10.1016/0092-8674(85)90306-x. [DOI] [PubMed] [Google Scholar]
  43. Ptashne M. How eukaryotic transcriptional activators work. Nature. 1988 Oct 20;335(6192):683–689. doi: 10.1038/335683a0. [DOI] [PubMed] [Google Scholar]
  44. Rosales R., Vigneron M., Macchi M., Davidson I., Xiao J. H., Chambon P. In vitro binding of cell-specific and ubiquitous nuclear proteins to the octamer motif of the SV40 enhancer and related motifs present in other promoters and enhancers. EMBO J. 1987 Oct;6(10):3015–3025. doi: 10.1002/j.1460-2075.1987.tb02607.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Saari G., Bienz M. The structure of the Ultrabithorax promoter of Drosophila melanogaster. EMBO J. 1987 Jun;6(6):1775–1779. doi: 10.1002/j.1460-2075.1987.tb02430.x. [DOI] [PMC free article] [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. Santoro C., Mermod N., Andrews P. C., Tjian R. A family of human CCAAT-box-binding proteins active in transcription and DNA replication: cloning and expression of multiple cDNAs. Nature. 1988 Jul 21;334(6179):218–224. doi: 10.1038/334218a0. [DOI] [PubMed] [Google Scholar]
  48. Scott M. P., Carroll S. B. The segmentation and homeotic gene network in early Drosophila development. Cell. 1987 Dec 4;51(5):689–698. doi: 10.1016/0092-8674(87)90092-4. [DOI] [PubMed] [Google Scholar]
  49. Soeller W. C., Poole S. J., Kornberg T. In vitro transcription of the Drosophila engrailed gene. Genes Dev. 1988 Jan;2(1):68–81. doi: 10.1101/gad.2.1.68. [DOI] [PubMed] [Google Scholar]
  50. Wharton K. A., Yedvobnick B., Finnerty V. G., Artavanis-Tsakonas S. opa: a novel family of transcribed repeats shared by the Notch locus and other developmentally regulated loci in D. melanogaster. Cell. 1985 Jan;40(1):55–62. doi: 10.1016/0092-8674(85)90308-3. [DOI] [PubMed] [Google Scholar]
  51. Wilde C. D., Akam M. Conserved sequence elements in the 5' region of the Ultrabithorax transcription unit. EMBO J. 1987 May;6(5):1393–1401. doi: 10.1002/j.1460-2075.1987.tb02380.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. van Ooyen A., Kwee V., Nusse R. The nucleotide sequence of the human int-1 mammary oncogene; evolutionary conservation of coding and non-coding sequences. EMBO J. 1985 Nov;4(11):2905–2909. doi: 10.1002/j.1460-2075.1985.tb04021.x. [DOI] [PMC free article] [PubMed] [Google Scholar]

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