Abstract
The expression of both proliferation-associated and cell type-specific genes is a hallmark of both cancer cells and tumor endothelial cells. The possibility to combine both features in a single transcriptional control unit would greatly increase the selectivity of vectors used for cancer gene therapy. Previous studies by our laboratory have shown that the transcription of several cell cycle genes is regulated by a novel cell cycle-regulated repressor, termed CDF-1. This repressor functions by blocking in resting cells the transcriptional activation by specific factors binding to the upstream activating sequence (UAS), most notably the CCAAT-box binding factor NF-Y/CBF. Based on this work we have developed a dual specificity promoter system that combines cell type specificity with cell cycle regulation. A chimeric transcription factor (Gal4/NF-Y) consisting of the transactivation domain of NF-Y and the DNA-binding domain of Gal4 is expressed from a tissue-specific promoter. Gal4/NF-Y can bind to a second promoter consisting of a minimal cyclin A promoter with multiple Gal4 binding sites replacing the normal UAS. This leads to the tissue-specific expression of Gal4/NF-Y whose stimulatory activity on the promoter is restrained in resting cells by the recruitment of the CDF-1 repressor to the promoter. The functionality of this system is demonstrated for the specific transcriptional targeting of proliferating melanoma cells, where cell cycle regulation was >20-fold and cell type specificity was >50-fold.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 12 print issues and online access
$259.00 per year
only $21.58 per issue
Rent or buy this article
Prices vary by article type
from$1.95
to$39.95
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Sikora K . Gene therapy for cancer Trends Biotech 1993 11: 197–201
Miller N, Whelan J . Progress in transcriptionally targeted and regulatable vectors for genetic therapy Hum Gene Ther 1997 8: 803–815
Rigg A, Sikora K . Genetic prodrug activation therapy Mol Med Today 1997 3: 359–366
Müller R . Transcriptional regulation during the mammalian cell cycle Trends Genet 1995 11: 173–178
Lucibello FC et al. Periodic cdc25C transcription is mediated by a novel cell cycle-regulated repressor element (CDE) EMBO J 1995 14: 132–142
Zwicker J et al. Cell cycle regulation of the cyclin A, cdc25C and cdc2 genes is based on a common mechanism of transcriptional repression EMBO J 1995 14: 4514–4522
Liu N et al. CDF-1, a novel E2F-unrelated factor, interacts with cell cycle-regulated repressor elements in multiple promoters Nucleic Acids Res 1997 25: 4915–4920
Zwicker J et al. Cell cycle regulation of cdc25C tanscription is mediated by the periodic repression of the glutamine-rich activators NF-Y and Sp1 Nucleic Acids Res 1995 23: 3822–3820
Dorn A et al. A multiplicity of CCAAT box-binding proteins Cell 1987 50: 863–872
Maity SM, Golumbek PT, Karsenty G, de Crombrugghe B . Selective activation of transcription by a novel CCAAT binding factor Science 1988 241: 582–585
van Hudisduijnen HR et al. Co-evolution from yeast to mouse: cDNA cloning of the two NF-Y (CP-1/CBF) subunits EMBO J 1990 9: 3119–3127
Maity SN, Vuorio T, Crombrugghe B . The B subunit of a rat heteromeric CCAAT-binding transcription factor shows a striking sequence identity with the yeast Hap2 transcription factor Proc Natl Acad Sci USA 1990 87: 5378–5382
Maity SN, Sinha S, Ruteshouse EC, de Crombrugghe B . Three different polypeptides are necessary for DNA binding of the mammalian heteromeric CCAAT binding factor J Biol Chem 1992 267: 16574–16580
Li X-Y et al. Intron–exon organization of the NF-Y genes. Tissue-specific splicing modifies an activation domain J Biol Chem 1992 267: 8984–8990
Mantovani R et al. Monoclonal antibodies to NF-Y define its function in MHC class II and albumin gene transcription EMBO J 1992 11: 3315–3322
Mantovani R et al. Dominant negative analogs of NF-YA J Biol Chem 1994 269: 20340–20346
Sinha S, Maity SN, Lu J, de Crombrugghe B . Recombinant rat CGF-C, the third subunit of CGF/NFY, allows formation of a protein–DNA complex with CBF-A and CBF-B and yeast HAP2 and HAP3 Proc Natl Acad Sci USA 1995 92: 1624–1628
Sinha S et al. Three classes of mutations in the A subunit of the CCAAT-binding factor CBF delineate functional domains involved in the three-step assembly of the CBF-DNA complex Mol Cell Biol 1996 16: 328–337
Coustry F, Maity SN, de Crombrugghe B . Studies on transcription activation by the multimeric CCAAT-binding factor CBF J Biol Chem 1995 270: 468–475
Currie RA . NF-Y is associated with the histone acetyltransferases GCN5 and P/CAF J Biol Chem 1998 273: 1430–1434
Zwicker J et al. CDF-1 mediated repression of cell cycle genes targets a specific subset of transactivators Nucleic Acids Res 1997 25: 4926–4932
Lech K, Anderson K, Brent R . DNA-bound fos proteins activate transcription in yeast Cell 1988 52: 179–184
Chasman DI, Kornberg RD . GAL4 protein: purification, association with GAL80 protein and conserved domain structure Mol Cell Biol 1990 10: 2916–2923
Webster N et al. The yeast UASG is a transcriptional enhancer in HeLa cells in the presence of the GAL4 trans-activator Cell 1988 52: 169–178
Siders WM, Halloran PJ, Fenton RG . Transcriptional targeting of recombinant adenoviruses to human and murine melanoma cells Cancer Res 1996 56: 5638–5646
Hwu P et al. Use of soluble recombinant TNF receptor to improve detection of TNF secretion in cultures of tumor infiltrating lymphocytes J Immunol Meth 1992 151: 139–147
Nettelbeck DM, Jérôme V, Müller R . A new strategy for the enhancement of weak cell type-specifc promoters Gene Therapy 1998 5: 1656–1664
Müller R, Nettelbeck D, Sedlacek HH . Chim¨are Promotormodule mit rekombinanten Transkriptionsfaktoren Patent Application 1998 DE 198 314 20.5
Bean MA et al. Cell-mediated cytotoxicity for bladder carcinoma: evaluation of a workshop Cancer Res 1975 35: 2902–2913
Sewing A et al. Human cyclin D1 encodes a labile nuclear protein whose synthesis is directly induced by growth factors and suppressed by cyclic AMP J Cell Sci 1993 104: 545–555
Lau QC, Brüsselbach S, Müller R . Abrogation of c-Raf expression induces apoptotis in tumor cells Oncogene 1998 16: 1899–1902
Henglein B et al. Structure and cell cycle-regulated transcription of the human cyclin A gene Proc Natl Acad Sci USA 1994 91: 5490–5494
Ausubel I, Frederick M . Current Protocols in Molecular Biology John Wiley: New York 1991
Acknowledgements
We are grateful to Dr S Brüsselbach for useful discussions, to Professors G Aumüller, I Hart and HH Sedlacek for tumor cell lines, to Drs M Clauss and D Männel for help with the TNFα assay and reagents and to Dr M Krause for synthesis of oligonucleotides.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Nettelbeck, D., Jérôme, V. & Müller, R. A dual specificity promoter system combining cell cycle-regulated and tissue-specific transcriptional control. Gene Ther 6, 1276–1281 (1999). https://doi.org/10.1038/sj.gt.3300943
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/sj.gt.3300943
Keywords
This article is cited by
-
Construction and Quantitative Evaluation of a Dual Specific Promoter System for Monitoring the Expression Status of Stra8 and c-kit Genes
Molecular Biotechnology (2014)
-
An Efficient and Safe Herpes Simplex Virus Type 1 Amplicon Vector for Transcriptionally Targeted Therapy of Human Hepatocellular Carcinomas
Molecular Therapy (2007)
-
Melanoma-specific expression in first-generation adenoviral vectors in vitro and in vivo — use of the human tyrosinase promoter with human enhancers
Cancer Gene Therapy (2005)
-
Combining high selectivity of replication with fiber chimerism for effective adenoviral oncolysis of CAR-negative melanoma cells
Gene Therapy (2004)
-
Expression in UVW glioma cells of the noradrenaline transporter gene, driven by the telomerase RNA promoter, induces active uptake of [131I]MIBG and clonogenic cell kill
Oncogene (2001)