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. 1993 Feb 15;90(4):1460–1464. doi: 10.1073/pnas.90.4.1460

Identification of a functional silencer element involved in neuron-specific expression of the synapsin I gene.

L Li 1, T Suzuki 1, N Mori 1, P Greengard 1
PMCID: PMC45893  PMID: 8381968

Abstract

We have identified a functional silencer element (positions -231 to -211) in the human synapsin I gene that selectively represses its transcription in nonneuronal cells. Transfection experiments using synapsin I-luciferase constructs show that site-specific mutations or deletion of this silencer sequence results in expression of the reporter gene in nonneuronal cells. Moreover, the silencer element is capable of conferring repression on a heterologous promoter in nonneuronal cells. Gel-shift assays reveal the presence of a sequence-specific synapsin I silencer-binding protein in nonneuronal cell extracts but not in neuronal cell extracts. Mutagenesis studies of the silencer sequence demonstrate that formation of the specific silencer-protein complex in vitro correlates well with repression of transcription in vivo. These data indicate that the interaction between synapsin I silencer and its binding protein is involved in tissue-specific expression of the synapsin I gene. In addition, our results suggest the existence of at least one additional cis-acting element within the promoter-proximal region (positions -233 to +20) that also contributes to the neuron-specific expression of the synapsin I gene.

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Selected References

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

  1. Baniahmad A., Steiner C., Köhne A. C., Renkawitz R. Modular structure of a chicken lysozyme silencer: involvement of an unusual thyroid hormone receptor binding site. Cell. 1990 May 4;61(3):505–514. doi: 10.1016/0092-8674(90)90532-j. [DOI] [PubMed] [Google Scholar]
  2. Camper S. A., Tilghman S. M. Postnatal repression of the alpha-fetoprotein gene is enhancer independent. Genes Dev. 1989 Apr;3(4):537–546. doi: 10.1101/gad.3.4.537. [DOI] [PubMed] [Google Scholar]
  3. De Camilli P., Benfenati F., Valtorta F., Greengard P. The synapsins. Annu Rev Cell Biol. 1990;6:433–460. doi: 10.1146/annurev.cb.06.110190.002245. [DOI] [PubMed] [Google Scholar]
  4. De Camilli P., Cameron R., Greengard P. Synapsin I (protein I), a nerve terminal-specific phosphoprotein. I. Its general distribution in synapses of the central and peripheral nervous system demonstrated by immunofluorescence in frozen and plastic sections. J Cell Biol. 1983 May;96(5):1337–1354. doi: 10.1083/jcb.96.5.1337. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Eustice D. C., Feldman P. A., Colberg-Poley A. M., Buckery R. M., Neubauer R. H. A sensitive method for the detection of beta-galactosidase in transfected mammalian cells. Biotechniques. 1991 Dec;11(6):739-40, 742-3. [PubMed] [Google Scholar]
  6. Fletcher C., Heintz N., Roeder R. G. Purification and characterization of OTF-1, a transcription factor regulating cell cycle expression of a human histone H2b gene. Cell. 1987 Dec 4;51(5):773–781. doi: 10.1016/0092-8674(87)90100-0. [DOI] [PubMed] [Google Scholar]
  7. Fujita T., Sakakibara J., Sudo Y., Miyamoto M., Kimura Y., Taniguchi T. Evidence for a nuclear factor(s), IRF-1, mediating induction and silencing properties to human IFN-beta gene regulatory elements. EMBO J. 1988 Nov;7(11):3397–3405. doi: 10.1002/j.1460-2075.1988.tb03213.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Gilman M. Z., Wilson R. N., Weinberg R. A. Multiple protein-binding sites in the 5'-flanking region regulate c-fos expression. Mol Cell Biol. 1986 Dec;6(12):4305–4316. doi: 10.1128/mcb.6.12.4305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Greene L. A., Tischler A. S. Establishment of a noradrenergic clonal line of rat adrenal pheochromocytoma cells which respond to nerve growth factor. Proc Natl Acad Sci U S A. 1976 Jul;73(7):2424–2428. doi: 10.1073/pnas.73.7.2424. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Haas C. A., DeGennaro L. J. Multiple synapsin I messenger RNAs are differentially regulated during neuronal development. J Cell Biol. 1988 Jan;106(1):195–203. doi: 10.1083/jcb.106.1.195. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Ho S. N., Hunt H. D., Horton R. M., Pullen J. K., Pease L. R. Site-directed mutagenesis by overlap extension using the polymerase chain reaction. Gene. 1989 Apr 15;77(1):51–59. doi: 10.1016/0378-1119(89)90358-2. [DOI] [PubMed] [Google Scholar]
  12. Hofmann J. F., Laroche T., Brand A. H., Gasser S. M. RAP-1 factor is necessary for DNA loop formation in vitro at the silent mating type locus HML. Cell. 1989 Jun 2;57(5):725–737. doi: 10.1016/0092-8674(89)90788-5. [DOI] [PubMed] [Google Scholar]
  13. Howland D. S., Hemmendinger L. M., Carroll P. D., Estes P. S., Melloni R. H., Jr, DeGennaro L. J. Positive- and negative-acting promoter sequences regulate cell type-specific expression of the rat synapsin I gene. Brain Res Mol Brain Res. 1991 Oct;11(3-4):345–353. doi: 10.1016/0169-328x(91)90044-x. [DOI] [PubMed] [Google Scholar]
  14. Kraner S. D., Chong J. A., Tsay H. J., Mandel G. Silencing the type II sodium channel gene: a model for neural-specific gene regulation. Neuron. 1992 Jul;9(1):37–44. doi: 10.1016/0896-6273(92)90218-3. [DOI] [PubMed] [Google Scholar]
  15. Manley J. L., Fire A., Cano A., Sharp P. A., Gefter M. L. DNA-dependent transcription of adenovirus genes in a soluble whole-cell extract. Proc Natl Acad Sci U S A. 1980 Jul;77(7):3855–3859. doi: 10.1073/pnas.77.7.3855. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Maue R. A., Kraner S. D., Goodman R. H., Mandel G. Neuron-specific expression of the rat brain type II sodium channel gene is directed by upstream regulatory elements. Neuron. 1990 Feb;4(2):223–231. doi: 10.1016/0896-6273(90)90097-y. [DOI] [PubMed] [Google Scholar]
  17. Mori N., Schoenherr C., Vandenbergh D. J., Anderson D. J. A common silencer element in the SCG10 and type II Na+ channel genes binds a factor present in nonneuronal cells but not in neuronal cells. Neuron. 1992 Jul;9(1):45–54. doi: 10.1016/0896-6273(92)90219-4. [DOI] [PubMed] [Google Scholar]
  18. Mori N., Stein R., Sigmund O., Anderson D. J. A cell type-preferred silencer element that controls the neural-specific expression of the SCG10 gene. Neuron. 1990 Apr;4(4):583–594. doi: 10.1016/0896-6273(90)90116-w. [DOI] [PubMed] [Google Scholar]
  19. Sauerwald A., Hoesche C., Oschwald R., Kilimann M. W. The 5'-flanking region of the synapsin I gene. A G+C-rich, TATA- and CAAT-less, phylogenetically conserved sequence with cell type-specific promoter function. J Biol Chem. 1990 Sep 5;265(25):14932–14937. [PubMed] [Google Scholar]
  20. Savagner P., Miyashita T., Yamada Y. Two silencers regulate the tissue-specific expression of the collagen II gene. J Biol Chem. 1990 Apr 25;265(12):6669–6674. [PubMed] [Google Scholar]
  21. Shen R. A., Goswami S. K., Mascareno E., Kumar A., Siddiqui M. A. Tissue-specific transcription of the cardiac myosin light-chain 2 gene is regulated by an upstream repressor element. Mol Cell Biol. 1991 Mar;11(3):1676–1685. doi: 10.1128/mcb.11.3.1676. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Südhof T. C., Czernik A. J., Kao H. T., Takei K., Johnston P. A., Horiuchi A., Kanazir S. D., Wagner M. A., Perin M. S., De Camilli P. Synapsins: mosaics of shared and individual domains in a family of synaptic vesicle phosphoproteins. Science. 1989 Sep 29;245(4925):1474–1480. doi: 10.1126/science.2506642. [DOI] [PubMed] [Google Scholar]
  23. Südhof T. C. The structure of the human synapsin I gene and protein. J Biol Chem. 1990 May 15;265(14):7849–7852. [PubMed] [Google Scholar]
  24. Tada H., Lashgari M. S., Khalili K. Regulation of JCVL promoter function: evidence that a pentanucleotide "silencer" repeat sequence AGGGAAGGGA down-regulates transcription of the JC virus late promoter. Virology. 1991 Jan;180(1):327–338. doi: 10.1016/0042-6822(91)90037-c. [DOI] [PubMed] [Google Scholar]
  25. Thiel G., Greengard P., Südhof T. C. Characterization of tissue-specific transcription by the human synapsin I gene promoter. Proc Natl Acad Sci U S A. 1991 Apr 15;88(8):3431–3435. doi: 10.1073/pnas.88.8.3431. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Valtorta F., Benfenati F., Greengard P. Structure and function of the synapsins. J Biol Chem. 1992 Apr 15;267(11):7195–7198. [PubMed] [Google Scholar]
  27. Weinberger J., Jat P. S., Sharp P. A. Localization of a repressive sequence contributing to B-cell specificity in the immunoglobulin heavy-chain enhancer. Mol Cell Biol. 1988 Feb;8(2):988–992. doi: 10.1128/mcb.8.2.988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Weissman J. D., Singer D. S. A complex regulatory DNA element associated with a major histocompatibility complex class I gene consists of both a silencer and an enhancer. Mol Cell Biol. 1991 Aug;11(8):4217–4227. doi: 10.1128/mcb.11.8.4217. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Winoto A., Baltimore D. Alpha beta lineage-specific expression of the alpha T cell receptor gene by nearby silencers. Cell. 1989 Nov 17;59(4):649–655. doi: 10.1016/0092-8674(89)90010-x. [DOI] [PubMed] [Google Scholar]
  30. de Wet J. R., Wood K. V., DeLuca M., Helinski D. R., Subramani S. Firefly luciferase gene: structure and expression in mammalian cells. Mol Cell Biol. 1987 Feb;7(2):725–737. doi: 10.1128/mcb.7.2.725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. van der Eb A. J., Graham F. L. Assay of transforming activity of tumor virus DNA. Methods Enzymol. 1980;65(1):826–839. doi: 10.1016/s0076-6879(80)65077-0. [DOI] [PubMed] [Google Scholar]

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