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.
Full text
PDF




Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]
- 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]