Nucleic Acids Research, Vol 26, Issue 13 3215-3220, Copyright © 1998 by Oxford University Press
T Lee, ME Bradley and JL Walowitz
Potent viral promoters/enhancers are often used to achieve high level
expression of ectopic genes in transient transfection analysis. By using a
GAL4-responsive transcription assay system, we show that the use of potent
eukaryotic expression vectors can lead to biased transcriptional effects.
Three functionally diverse transcription factors, YY1, SRF and Msx-1, were
examined and each was found to exhibit a strong transrepression function in
the context of the DNA binding domain of GAL4 when expressed from the
cytomegalovirus (pCMV) or simian virus 40 (pSV) promoters/enhancers. An
internal 15 amino acid domain of YY1 mediating transrepression in the viral
promoter setting was identified. This GAL4-mediated transcriptional
repression could, however, be completely relieved by using the yeast
alcohol dehydrogenase promoter (pADH) to drive gene expression, which is
approximately 100-fold weaker than canonical pCMV and pSV in cultured
mammalian cells. In addition, low level expression achieved with the pADH
vector unveiled the intrinsic transactivation functions of YY1 and SRF
previously not observed with the GAL4 assay system. Our results highlight a
potential pitfall in conventional pCMV- and pSV-based transfection assays
and suggest that the use of a low level expression system may be preferable
in most transcriptional analysis.
ARTICLES
Influence of promoter potency on the transcriptional effects of YY1, SRF and Msx-1 in transient transfection analysis
Department of Biochemistry, SUNY at Buffalo, 140 Farber Hall, 3435 Main Street, Buffalo, NY 14214-3000, USA. chunglee@acsu.buffalo.edu
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  H. Lin, J. McGrath, P. Wang, and T. Lee Cellular Toxicity Induced by SRF-Mediated Transcriptional Squelching Toxicol. Sci., March 1, 2007; 96(1): 83 - 91. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Osborne, H. Zhang, W.-M. Yang, E. Seto, and G. Blanck Histone Deacetylase Activity Represses Gamma Interferon-Inducible HLA-DR Gene Expression following the Establishment of a DNase I-Hypersensitive Chromatin Conformation Mol. Cell. Biol., October 1, 2001; 21(19): 6495 - 6506. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. M. Foti, A. Welihinda, R. J. Kaufman, and A. S. Lee Conservation and Divergence of the Yeast and Mammalian Unfolded Protein Response. ACTIVATION OF SPECIFIC MAMMALIAN ENDOPLASMIC RETICULUM STRESS ELEMENT OF THE grp78/BiP PROMOTER BY YEAST Hac1 J. Biol. Chem., October 22, 1999; 274(43): 30402 - 30409. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 W. Zhu, C. TomHon, M. Mason, T. Campbell, E. Shelden, N. Richards, M. Goodman, and D. L. Gumucio Analysis of Linked Human {varepsilon} and {gamma} Transgenes: Effect of Locus Control Region Hypersensitive Sites 2 and 3 or a Distal YY1 Mutation on Stage-Specific Expression Patterns Blood, May 15, 1999; 93(10): 3540 - 3549. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Wu and A. S. Lee YY1 as a Regulator of Replication-dependent Hamster Histone H3.2 Promoter and an Interactive Partner of AP-2 J. Biol. Chem., January 5, 2001; 276(1): 28 - 34. [Abstract] [Full Text] [PDF]
|
|