C. Tiffoche et al., Novel intronic promoter in the rat ER alpha gene responsible for the transient transcription of a variant receptor, ENDOCRINOL, 142(9), 2001, pp. 4106-4119
To analyze the molecular origin of an ER variant, the truncated ER product-
1, transiently expressed at the proestrus in lactotrope cells, we generated
a 2.5-kb sequence of a genomic region upstream and downstream the specific
sequence truncated ER product-1. Genomic Southern blot analysis showed tha
t truncated ER product-1 is spliced from a noncoding leader exon localized
within the intron 4 of the ER alpha gene. Analysis of the promoter sequence
revealed the presence of a major transcriptional start site, a canonical T
ATA box and putative cis regulatory elements for pituitary specific express
ion as well as an E-responsive element. In transient transfection, the trun
cated ER product-1 promoter was transcriptionally the most active in the la
ctotrope cell lines (MMQ). Analysis of truncated ER product-1 functionality
showed that: 1) the protein inhibited ER alpha binding to the E-responsive
element in electromobility shift assays, 2) inhibited the E2 binding to ER
alpha in binding assays, 3) the truncated ER product-1/ER alpha complex an
tagonized the transcriptional activity elicited by E2, 4) nuclear localizat
ion of green fluorescent protein-ER alpha was altered in Chinese hamster ov
ary cell lines stably expressing truncated ER product-1. Collectively, thes
e data demonstrated that the protein exerts full dominant negative activity
against ER alpha. Moreover, truncated ER product-1/ER alpha complex also r
epressed the activity of all promoters tested to date, suggesting a general
inhibitory effect toward transcription. In conclusion, the data suggest th
at truncated ER product-1 could regulate estrogen signaling via a specific
promoter in lactotrope cells.