Abstract
A region (NS1) that acts like an enhancer is located approximately 300 bp upstream of the larval cap site in theAdh gene ofD. melanogaster. When this sequence is deleted (ΔNS1), the gene fails to express ADH protein. Gene expression can be restored by placing a secondAdh gene with an intact enhancer elsewhere on the same plasmid. In these circumstances, both genes are expressed equally regardless of their orientation on the plasmid. In this report we further characterize the interactions that occur when a single enhancer activates expression from a proximal and distant promoter. We have made the following observations: (1) While the two genes are expressed equivalently, their expression relative to a plasmid carrying two intact genes is reduced by a factor of 2 to 6 depending on the orientation of the two genes. (2) The single enhancer drives expression of both genes on any given plasmid molecule. (3) The enhancer does not interact with theAdh gene from which the NS7 region (which spans the larval TATA box) is removed. (4) Expression of the ΔNS1 gene can be restored by an intact gene when both are inserted together into theDrosophila genome via P element-mediated transformation. (5) Increasing the separation between the two genes on a plasmid by up to 15 kbp does not prevent the restoration of expression of the ΔNS1 gene. We propose a model that explains how a single enhancer can stimulate equal expression from two genes.
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Atchison, M.L. & R.P. Perry, 1986. Tandem kappa immunoglobin promoters are equally active in the presence of kappa enhancer: Implications for models of enhancer function. Cell 46: 253–262.
Ausubel, F.M., R. Brent, R.E. Kingston, D.D. Moore, J.G. Seidman, J.A. Smith & K. Sthrul, 1987. Current Protocols in Molecular Biology, John Wiley & Sons, New York.
Banerji, J., S. Rusconi & W. Scnaffner, 1981. Expression of a β-globin gene is enhanced by remote SV40 sequences. Cell 27: 299–308.
Benyajati, C., A.R. Place & W. Sofer, 1983. Formaldehyde mutagenesis inDrosophila melanogaster molecular analysis of alcohol dehydrogenase negative mutants. Mutat. Res. 111: 1–8.
Carey, M., J. Leatherwood & M. Ptashne, 1990. A potent gal4 derivative activates transcription at a distance in vitro. Science 247: 710–712.
Courney, A.J., S.E. Pion & J.C. Wang, 1986. The use of psoralen-modified DNA to probe the mechanism of enhancer action. Cell 45: 567–574.
Darby, M.K., M.T. Andrews & D.D. Brown, 1988. Transcription complexes that programXenopus 5s RNA gene are stable in vivo. Proc. Natl. Acad. Sci. USA 85: 5516–5520.
Davidson, B.L., J. Egly, E.R. Mulvhill & P. Chambon, 1983. Formation of stable preinitiation complexes between eukaryotic class B transcription factors and promoter sequences. Nature 301: 680–686.
Gustafson, T.A., A. Taylor & L. Kedes, 1989. DNA bending is induced by a transcription factor that interacts with the human c-fos and alpha actin promoters. Proc. Natl. Acad. Sci. USA 86: 2162–2166.
Heuchel, R., P. Matthias & W. Schaffner, 1989. Two closely spaced promoters are equally activated by a remote enhancer: Evidence against a scanning model for enhancer action. Nucleic Acids Research 17: 8931–8947.
Jack, J., D. Dorsett, Y. Delotto & S. Liu, 1991. Expression of the cut locus in theDrosophila wing margin is required for cell type specification and is regulated by a distant enhancer. Development 113: 735–748.
Kanaar, R., D.P.P. Van & N.R. Cozzarelli, 1989. Gin-mediated recombination of catenated and knotted DNA substrates implications for the mechanism of interaction between cis-acting sites. Cell 58: 147–160.
Karess, R.E. & G.M. Rubin, 1984. Analysis of P transposable element function inDrosophila. Cell 38: 135–146.
Kunkle, T.A., J.D. Roberts & R.A. Zakour, 1987. Rapid and efficient site-directed mutagenesis without phenotypic selection. Meth. Enzymol. 154: 367–382.
Livak, K.J., 1984. Organization and mapping of a sequence on theDrosophila melanogaster X and Y chromosomes that is transcribed during spermatogenesis. Genetics 107: 611–634.
Martin, P., A. Martin, A. Osmani & W. Sofer, 1986. A transient expression assay for tissue-specific gene expression of alcohol dehydrogenase inDrosophila. Dev. Biol. 117: 574–580.
Müller, H.P., J.M. Sogo & W. Schaffner, 1989. An enhancer stimulates transcription in trans when attached to the promoter via a protein bridge. Cell 58: 767–778.
Müller, M.M., T. Gerster & W. Schaffner, 1989. Enhancer sequences and the regulation of gene transcription. Eur. J. Biochem. 176: 485–495.
Nordhelm, A. & A. Rich, 1983. Negatively supercoiled simian virus 40 DNA contains Z-DNA segments within transcriptional enhancer sequences. Nature 303: 674–679.
Pion, S.E. & J.C. Wang, 1986. Transcription of human β-globin gene is stimulated by an SV40 enhancer to which it is physically linked but topologically uncoupled. Cell 45: 575–580.
Pipkin, S.B. & N.E. Hewitt, 1972. Effect of gene dosage on level of alcohol dehydrogenase inDrosophila. Journal of Heredity 64: 331–336.
Posakony, J.W., J.A. Pischer & T. Maniatis, 1986. Identification of DNA sequences required for the regulation ofDrosophila alcohol dehydrogenase gene expression. Cold Spring Harbor Symp. Quant. Biol. 50: 515–520.
Stashne, M., 1986. Gene regulation by proteins acting nearby and at a distance. Nature 322: 697–701.
Rabinow, L. & J. Birchler, 1986. Reciprocal translocation between balancers of the second and third chromosomes carrying a dominant temperature sensitive lethal and larval markers. DIS 63: 150.
Roth, G.E., 1991. Replication analysis of plasmid DNAs injected intoDrosophila embryos. Chromosoma 100: 267–277.
Rothberg, I., E. Hotaling & W. Sofer, 1991. ADrosophila Adh gene can be activated in trans by an enhancer. Nucleic Acids Research 19: 5713–5717.
Rubin, G.M. & A.C. Spradling, 1982. Genetic transformation ofDrosophila with transposable element vectors. Science 218: 348–353.
Rubin, G.M. & A.C. Spradling, 1983. Vectors for P element-mediated gene transfer inDrosophila. Nucleic Acids Research 11: 6341–6351.
Sambrook, J., E.F. Fritsch & T. Maniatis, 1989. Molecular cloning: A laboratory manual. Cold Spring Harbor, New York: Cold Spring Harbor Laboratory.
Schwartz, M. & W. Sofer, 1976. Diet induced alterations in distribution of multiple forms of alcohol dehydrogenase inDrosophila. Nature 263: 129–131.
Schwartz, M.F. & H. Jornvall, 1976. Structural analyses of mutant and wild-type alcohol dehydrogenase fromDrosophila melanogaster. Eur. J. Biochem. 68: 159–168.
Schwartz, M., J. O'Donnell & W. Sofer, 1979. Origin of multiple forms of alcohol dehydrogenase EC-1.1.1.1 fromDrosophila melanogaster. Arch. Biochem. Biophys. 194: 365–378.
Serfling, E., M. Jasin & W. Schaffner, 1985. Enhancers and eukaryotic gene transcription. Trends Genet. 1: 224–230.
Shen, N.L.L., E. Hotaling, G. Subrahmanyam, P.F. Martin & W. Sofer, 1991. Analysis of sequences regulating expression of the Adh gene ofDrosophila melanogaster. Genetics 129: 763–771.
Shen, N.L.L., G. Subrahmanyam, W. Clark, P. Martin & W. Sofer, 1989. Analysis of Adh gene regulation inDrosophila: studies using somatic transformation. Dev. Genet. 10: 210–219.
Shore, E.M. & G.M. Guild, 1987. Closely linked DNA elements control the expression of the Sgs-5 glue protein gene inDrosophila. Genes & Development 1: 829–839.
Shuey, D.J. & C.S. Parker, 1986. Bending of promoter DNA on binding of heat shock transcription factor. Nature 323: 459–461.
Spradling, A.C. & G.M. Rubin, 1982. Transposition of cloned P elements intoDrosophila germ line chromosomes. Science 218: 341–347.
Thatcher, D.R. & L. Sawyer, 1980. The complete amino acid sequences of 3 alcohol dehydrogenase EC-1.1.1.1 allelo enzymes (Adhn11, AdhS, AdhUF) from the fruitflyDrosophila melanogaster. Biochem. J. 187: 875–86.
Viera, J. & J. Messing, 1984. Production of single-stranded plasmid DNA. Methods in Enzymology 153: 3–11.
Wang, J.C. & G.N. Giaever, 1988. Action at a distance along a DNA. Science 240: 300–304.
Wang, J.C., L.J. Peck & K. Becherer, 1983. DNA supercoling and its effects on DNA structure and function. Cold Spring Harbor Symp. Quant. Biol. 47: 85–91.
Weintraub, H., P.F. Cheng & K. Conrad, 1986. Expression of transfected DNA depends on DNA topology. Cell 46: 115–122.
Wolffe, A.P. & D.D. Brown, 1986. DNA replication in vitro erases aXenopus laevis 5s RNA gene transcription complex. Cell 47: 217–228.
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Freidman, R., Hotaling, E., Borack, L. et al. Interactions between the regulatory regions of twoAdh alleles. Genetica 97, 1–14 (1996). https://doi.org/10.1007/BF00132575
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DOI: https://doi.org/10.1007/BF00132575