1: Nucleic Acids Res. 1998 Jul 1;26(13):3215-20. 

Influence of promoter potency on the transcriptional effects of YY1, SRF and
Msx-1 in transient transfection analysis.

Lee T, Bradley ME, Walowitz JL.

Department of Biochemistry, SUNY at Buffalo, 140 Farber Hall, 3435 Main Street,
Buffalo, NY 14214-3000, USA. chunglee@acsu.buffalo.edu

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.

PMID: 9628921 [PubMed - indexed for MEDLINE]


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