Skip to main content
The EMBO Journal logoLink to The EMBO Journal
. 1989 May;8(5):1559–1565. doi: 10.1002/j.1460-2075.1989.tb03540.x

A 14 bp promoter element directs the testis specificity of the Drosophila beta 2 tubulin gene.

F Michiels 1, A Gasch 1, B Kaltschmidt 1, R Renkawitz-Pohl 1
PMCID: PMC400987  PMID: 2504583

Abstract

To analyze the regulation of gene expression during male germ cell development, we investigated the testis-specific expression of the Drosophila beta 2 tubulin gene. Germ line transformation experiments with the upstream region of the D.melanogaster beta 2 tubulin gene fused to the Escherichia coli lacZ gene resulted in the correct tissue specific expression of the reporter gene. Furthermore, we showed that the upstream sequences of the beta 2 tubulin gene of the distantly related species D.hydei can drive the expression of the lacZ gene testis specifically in D.melanogaster flies. A detailed deletion analysis showed that 53 bp of upstream and 23 bp (D.melanogaster) or 29 bp (D.hydei) of leader sequences are sufficient to confer tissue specificity. The short promoter regions contain a 14 bp motif at identical positions in both species, which acts as a position-dependent promoter element. In vitro mutagenesis and subsequent germline transformation experiments revealed that this sequence is the only element necessary for the testis-specific transcription of the beta 2 tubulin gene in Drosophila.

Full text

PDF
1561

Images in this article

Selected References

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

  1. Allen R. L., O'Brien D. A., Eddy E. M. A novel hsp70-like protein (P70) is present in mouse spermatogenic cells. Mol Cell Biol. 1988 Feb;8(2):828–832. doi: 10.1128/mcb.8.2.828. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Barberis A., Superti-Furga G., Busslinger M. Mutually exclusive interaction of the CCAAT-binding factor and of a displacement protein with overlapping sequences of a histone gene promoter. Cell. 1987 Jul 31;50(3):347–359. doi: 10.1016/0092-8674(87)90489-2. [DOI] [PubMed] [Google Scholar]
  3. 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]
  4. Bialojan S., Falkenburg D., Renkawitz-Pohl R. Characterization and developmental expression of beta tubulin genes in Drosophila melanogaster. EMBO J. 1984 Nov;3(11):2543–2548. doi: 10.1002/j.1460-2075.1984.tb02170.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. 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]
  6. Busslinger M., Schümperli D., Birnstiel M. L. Regulation of histone gene expression. Cold Spring Harb Symp Quant Biol. 1985;50:665–670. doi: 10.1101/sqb.1985.050.01.081. [DOI] [PubMed] [Google Scholar]
  7. Chen E. Y., Seeburg P. H. Supercoil sequencing: a fast and simple method for sequencing plasmid DNA. DNA. 1985 Apr;4(2):165–170. doi: 10.1089/dna.1985.4.165. [DOI] [PubMed] [Google Scholar]
  8. Cleveland D. W., Sullivan K. F. Molecular biology and genetics of tubulin. Annu Rev Biochem. 1985;54:331–365. doi: 10.1146/annurev.bi.54.070185.001555. [DOI] [PubMed] [Google Scholar]
  9. Cole K. D., Kandala J. C., Kistler W. S. Isolation of the gene for the testis-specific H1 histone variant H1t. J Biol Chem. 1986 Jun 5;261(16):7178–7183. [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. Fischer J. A., Maniatis T. Drosophila Adh: a promoter element expands the tissue specificity of an enhancer. Cell. 1988 May 6;53(3):451–461. doi: 10.1016/0092-8674(88)90165-1. [DOI] [PubMed] [Google Scholar]
  12. Fuller M. T., Caulton J. H., Hutchens J. A., Kaufman T. C., Raff E. C. Genetic analysis of microtubule structure: a beta-tubulin mutation causes the formation of aberrant microtubules in vivo and in vitro. J Cell Biol. 1987 Mar;104(3):385–394. doi: 10.1083/jcb.104.3.385. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Garabedian M. J., Shepherd B. M., Wensink P. C. A tissue-specific transcription enhancer from the Drosophila yolk protein 1 gene. Cell. 1986 Jun 20;45(6):859–867. doi: 10.1016/0092-8674(86)90560-x. [DOI] [PubMed] [Google Scholar]
  14. Gasch A., Hinz U., Leiss D., Renkawitz-Pohl R. The expression of beta 1 and beta 3 tubulin genes of Drosophila melanogaster is spatially regulated during embryogenesis. Mol Gen Genet. 1988 Jan;211(1):8–16. doi: 10.1007/BF00338387. [DOI] [PubMed] [Google Scholar]
  15. Glaser R. L., Wolfner M. F., Lis J. T. Spatial and temporal pattern of hsp26 expression during normal development. EMBO J. 1986 Apr;5(4):747–754. doi: 10.1002/j.1460-2075.1986.tb04277.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Grosschedl R., Baltimore D. Cell-type specificity of immunoglobulin gene expression is regulated by at least three DNA sequence elements. Cell. 1985 Jul;41(3):885–897. doi: 10.1016/s0092-8674(85)80069-6. [DOI] [PubMed] [Google Scholar]
  17. Hackstein J. H. Spermatogenesis in Drosophila. Results Probl Cell Differ. 1987;15:63–116. doi: 10.1007/978-3-540-47184-4_2. [DOI] [PubMed] [Google Scholar]
  18. Hall C. V., Jacob P. E., Ringold G. M., Lee F. Expression and regulation of Escherichia coli lacZ gene fusions in mammalian cells. J Mol Appl Genet. 1983;2(1):101–109. [PubMed] [Google Scholar]
  19. Heberlein U., England B., Tjian R. Characterization of Drosophila transcription factors that activate the tandem promoters of the alcohol dehydrogenase gene. Cell. 1985 Jul;41(3):965–977. doi: 10.1016/s0092-8674(85)80077-5. [DOI] [PubMed] [Google Scholar]
  20. Heberlein U., Tjian R. Temporal pattern of alcohol dehydrogenase gene transcription reproduced by Drosophila stage-specific embryonic extracts. Nature. 1988 Feb 4;331(6155):410–415. doi: 10.1038/331410a0. [DOI] [PubMed] [Google Scholar]
  21. Hennig W. Untersuchungen zur Struktur und Funktion des Lampenbürsten-Y-Chromosoms in der Spermatogenese von Drosophila. Chromosoma. 1967;22(3):294–357. doi: 10.1007/BF00319879. [DOI] [PubMed] [Google Scholar]
  22. Hiromi Y., Gehring W. J. Regulation and function of the Drosophila segmentation gene fushi tarazu. Cell. 1987 Sep 11;50(6):963–974. doi: 10.1016/0092-8674(87)90523-x. [DOI] [PubMed] [Google Scholar]
  23. Kalfayan L., Wensink P. C. Developmental regulation of Drosophila alpha-tubulin genes. Cell. 1982 May;29(1):91–98. doi: 10.1016/0092-8674(82)90093-9. [DOI] [PubMed] [Google Scholar]
  24. Karess R. E., Rubin G. M. Analysis of P transposable element functions in Drosophila. Cell. 1984 Aug;38(1):135–146. doi: 10.1016/0092-8674(84)90534-8. [DOI] [PubMed] [Google Scholar]
  25. Kemphues K. J., Kaufman T. C., Raff R. A., Raff E. C. The testis-specific beta-tubulin subunit in Drosophila melanogaster has multiple functions in spermatogenesis. Cell. 1982 Dec;31(3 Pt 2):655–670. doi: 10.1016/0092-8674(82)90321-x. [DOI] [PubMed] [Google Scholar]
  26. Kemphues K. J., Raff R. A., Kaufman T. C., Raff E. C. Mutation in a structural gene for a beta-tubulin specific to testis in Drosophila melanogaster. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3991–3995. doi: 10.1073/pnas.76.8.3991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Klemenz R., Weber U., Gehring W. J. The white gene as a marker in a new P-element vector for gene transfer in Drosophila. Nucleic Acids Res. 1987 May 26;15(10):3947–3959. doi: 10.1093/nar/15.10.3947. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Krawczyk Z., Wiśniewski J., Biesiada E. A hsp70-related gene is constitutively highly expressed in testis of rat and mouse. Mol Biol Rep. 1987;12(1):27–34. doi: 10.1007/BF00580647. [DOI] [PubMed] [Google Scholar]
  29. Kuhn R., Schäfer U., Schäfer M. Cis-acting regions sufficient for spermatocyte-specific transcriptional and spermatid-specific translational control of the Drosophila melanogaster gene mst(3)gl-9. EMBO J. 1988 Feb;7(2):447–454. doi: 10.1002/j.1460-2075.1988.tb02832.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Leiss D., Hinz U., Gasch A., Mertz R., Renkawitz-Pohl R. Beta 3 tubulin expression characterizes the differentiating mesodermal germ layer during Drosophila embryogenesis. Development. 1988 Dec;104(4):525–531. doi: 10.1242/dev.104.4.525. [DOI] [PubMed] [Google Scholar]
  31. Lifschytz E. The developmental program of spermiogenesis in Drosophila: a genetic analysis. Int Rev Cytol. 1987;109:211–258. doi: 10.1016/s0074-7696(08)61723-4. [DOI] [PubMed] [Google Scholar]
  32. Maniatis T., Goodbourn S., Fischer J. A. Regulation of inducible and tissue-specific gene expression. Science. 1987 Jun 5;236(4806):1237–1245. doi: 10.1126/science.3296191. [DOI] [PubMed] [Google Scholar]
  33. Mathis D. J., Chambon P. The SV40 early region TATA box is required for accurate in vitro initiation of transcription. Nature. 1981 Mar 26;290(5804):310–315. doi: 10.1038/290310a0. [DOI] [PubMed] [Google Scholar]
  34. McKnight S. L., Kingsbury R. Transcriptional control signals of a eukaryotic protein-coding gene. Science. 1982 Jul 23;217(4557):316–324. doi: 10.1126/science.6283634. [DOI] [PubMed] [Google Scholar]
  35. Michiels F., Falkenburg D., Müller A. M., Hinz U., Otto U., Bellmann R., Glätzer K. H., Brand R., Bialojan S., Renkawitz-Pohl R. Testis-specific beta 2 tubulins are identical in Drosophila melanogaster and D. hydei but differ from the ubiquitous beta 1 tubulin. Chromosoma. 1987;95(6):387–395. doi: 10.1007/BF00333989. [DOI] [PubMed] [Google Scholar]
  36. Natzle J. E., McCarthy B. J. Regulation of Drosophila alpha- and beta-tubulin genes during development. Dev Biol. 1984 Jul;104(1):187–198. doi: 10.1016/0012-1606(84)90047-2. [DOI] [PubMed] [Google Scholar]
  37. Peschon J. J., Behringer R. R., Brinster R. L., Palmiter R. D. Spermatid-specific expression of protamine 1 in transgenic mice. Proc Natl Acad Sci U S A. 1987 Aug;84(15):5316–5319. doi: 10.1073/pnas.84.15.5316. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Propst F., Rosenberg M. P., Vande Woude G. F. Proto-oncogene expression in germ cell development. Trends Genet. 1988 Jul;4(7):183–187. doi: 10.1016/0168-9525(88)90073-x. [DOI] [PubMed] [Google Scholar]
  39. Rubin G. M., Spradling A. C. Genetic transformation of Drosophila with transposable element vectors. Science. 1982 Oct 22;218(4570):348–353. doi: 10.1126/science.6289436. [DOI] [PubMed] [Google Scholar]
  40. 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]
  41. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  42. Steller H., Pirrotta V. Regulated expression of genes injected into early Drosophila embryos. EMBO J. 1984 Jan;3(1):165–173. doi: 10.1002/j.1460-2075.1984.tb01778.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Theurkauf W. E., Baum H., Bo J., Wensink P. C. Tissue-specific and constitutive alpha-tubulin genes of Drosophila melanogaster code for structurally distinct proteins. Proc Natl Acad Sci U S A. 1986 Nov;83(22):8477–8481. doi: 10.1073/pnas.83.22.8477. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Wasylyk B., Derbyshire R., Guy A., Molko D., Roget A., Téoule R., Chambon P. Specific in vitro transcription of conalbumin gene is drastically decreased by single-point mutation in T-A-T-A box homology sequence. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7024–7028. doi: 10.1073/pnas.77.12.7024. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Wirth T., Staudt L., Baltimore D. An octamer oligonucleotide upstream of a TATA motif is sufficient for lymphoid-specific promoter activity. Nature. 1987 Sep 10;329(6135):174–178. doi: 10.1038/329174a0. [DOI] [PubMed] [Google Scholar]
  46. Wu L., Rosser D. S., Schmidt M. C., Berk A. A TATA box implicated in E1A transcriptional activation of a simple adenovirus 2 promoter. Nature. 1987 Apr 2;326(6112):512–515. doi: 10.1038/326512a0. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES