Characterization of the transcription start point of the trout estrogen receptor-encoding gene: evidence for alternative splicing in the 5' untranslated region.

ABSTRACT

The estrogen receptor (ER)-encoding gene (ER) regulates many genes implicated in the reproductive functions. Moreover, rainbow trout ER (rtER) is itself up-regulated by its own product. We have used Northern blot, RNase protection, primer extension and reverse transcription-polymerase chain reaction (RT-PCR) to study the position of the rtER mRNA transcription start point (tsp) in liver. This analysis has revealed the presence of a tsp positioned at the beginning of the cloned rtER cDNA. Functionality of this tsp was tested in transient transfections in CHO-K1 cells. The characterization of the rtER 5' untranslated region (UTR) showed that two transcripts exist in liver which differ in their 5'-UTR. The first one is 100% homologous to the cloned rtER cDNA sequence. The other one contains a 41-bp insertion. The isolation and sequencing of the first intron showed that this insertion arises from alternative splicing, due to the use of a splicing site internal to the first intron.