The tissue-specific mammalian transcription factor, Pit-1, activates transcription in Saccharomyces cerevisiae.
Pit-1 is a tissue-specific transcription factor which binds to specific DNA sequences within 5' flanking regions of the PRL and GH genes and activates the transcription of these genes. Previous studies have shown that expression of Pit-1 is necessary to activate transcription from the PRL or GH promoters in heterologous mammalian cells. In the present study the ability of Pit-1 expression vectors to activate expression of reporter genes in Saccharomyces cerevisiae was examined. The test system used Pit-1 expression vectors and an indicator plasmid containing multiple copies of a Pit1-binding site as a replacement for the upstream activator sequence of the CYC1 promoter. Significant activation of indicator plasmid expression was detected only in the presence of functional Pit-1 expression vectors. In both mammalian and yeast cells, amino-terminal deletions of the Pit-1 coding sequence produced similar and gradual loss of transcriptional activation. This finding indicates that similar or identical regions of Pit-1 are required for transcriptional activation in mammalian and yeast cells. Although synthetic DNA elements containing multiple copies of a single Pit-1-binding site were sufficient to permit Pit-1-mediated transcriptional activation in both yeast and mammalian cells, DNA fragments representing the proximal region or distal enhancer region of the PRL gene were transcriptionally active only in mammalian cells. These studies establish the ability of Pit-1 to stimulate transcription in the absence of other tissue-specific factors and provide a system for further genetic studies of Pit-1 structure/function relationships as well as evaluation of target sequences necessary for Pit-1 action.
Ding
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Department of Physiology and Biophysics, University of Iowa, Iowa City 52242.
Lu
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Roberson
M S
MS
Moye-Rowley
W S
WS
Maurer
R A
RA
D000818:Animals; D004252:DNA Mutational Analysis; D005809:Genes, Regulator / Q000235:genetics*; D005822:Genetic Vectors; D008156:Luciferases; D010903:Pituitary Gland, Anterior / Q000201:enzymology*; D051381:Rats; D011993:Recombinant Fusion Proteins; D012045:Regulatory Sequences, Nucleic Acid; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D013045:Species Specificity; D013329:Structure-Activity Relationship; D014157:Transcription Factors / Q000235:genetics*; D014158:Transcription, Genetic; D015533:Transcriptional Activation / Q000235:genetics*; D001616:beta-Galactosidase
D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D008156:Luciferases; D001616:beta-Galactosidase