Rat liver c-erb A beta 1 thyroid hormone receptor is a constitutive activator in yeast (Saccharomyces cerevisiae): essential role of domains D,E and F in hormone-independent transcription.
To assess thyroid hormone receptor (TR)-mediated activation of transcription in yeast in the presence or absence of thyroid hormone (T3), we developed a co-expression system using a TR-beta 1 expression vector and a reporter plasmid containing a 16 base pair palindromic thyroid hormone response element (TRE) upstream from a proximal CYC1 promoter that was fused to the beta-galactosidase lac Z gene of Escherichia coli. Although TR-beta 1 functions as a repressor in most mammalian systems, using our system we observed a unique thyroid hormone-independent transcriptional response indicating that wild TR-beta 1 acted as a constitutive activator in yeast; the addition of 1 microM T3 induced a moderate but significant (p less than 0.01) 25-40% further increase in transcriptional activity. Using a series of rat TR-beta 1 mutant constructs, we found that deletion of domain D and portions of E completely eliminated transcriptional activity, whereas truncations of domain F and E permitted a partial (20-40%) response compared to wild TR-beta 1 in the presence or absence of T3. These observations indicate that TR-beta 1 functions as an activator in yeast and that domains D,E and F play important interactive roles in its hormone-independent gene activation with the D domain likely being the most essential. Furthermore, our results suggest that the different transcriptional property of TR-beta 1 in yeast compared to mammalian cells i.e. activator vs repressor function, is likely determined by transcriptional factor differences which are dependent upon cellular origin.
Ohashi
H
H
Thyroid Research Laboratory, Samuel Lunenfeld Research Institute, Mount Sinai Hospital, University of Toronto, Ontario, Canada.
Yang
Y F
YF
Walfish
P G
PG
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics; D015966:Gene Expression Regulation, Fungal / Q000187:drug effects; D005798:Genes, Bacterial; D005822:Genetic Vectors; D008099:Liver / Q000502:physiology*; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011988:Receptors, Thyroid Hormone / Q000235:genetics / Q000378:metabolism*; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D013963:Thyroid Hormones / Q000494:pharmacology*; D014158:Transcription, Genetic / Q000187:drug effects; D015533:Transcriptional Activation; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D015345:Oligonucleotide Probes; D011518:Proto-Oncogene Proteins; D011988:Receptors, Thyroid Hormone; D013963:Thyroid Hormones; D001616:beta-Galactosidase