Abstract
BackgroundEstrogen receptor alpha (ERa/ESR1) expression is regulated by alternative splicing. Its most frequently detectable exon7 skipping isoform (ERaD7) is a dominant negative variant. Elevated expression of ERaD7 was already detected in endometrial cancer (EC), while its potential prognostic significance has not been characterized so far. Exon7 contains potential binding sites for the two functional splicing regulatory opponents, HNRNPG and HTRA2-BETA1 known to trigger opposite effects on EC outcome.This study served to elucidate the influence of HNRNPG and HTRA2-BETA1 on ERa exon7 splicing regulation and the impact of ERaD7 concentration on type 1 EC outcome.MethodsFunctional in vitro experiments for HNRNPG and HTRA2-BETA1 in regard to the regulatory impact on endogenous and exogenous ERaD7 splicing were performed. Additionally, real-time PCR determined mRNA levels of ERaD7, HNRNPG and HTRA2-BETA1 in 116 type 1 EC patients.ResultsHNRNPG and HTRA2-BETA1 were found to be specific regulators of ERa exon7 splicing. While HTRA2-BETA1 promoted exon7 inclusion, HNRNPG antagonized this effect by inducing exon7 skipping (p = 0.004). ERaD7 was detected in 71 out of 116 type 1 EC specimens. Statistical analyses revealed an inverse correlation between ERaD7 mRNA levels and tumor grading (p = 0.029), FIGO stage (p = 0.033) as well as lymph node metastases (p = 0.032), respectively. Furthermore, higher ERaD7 expression could be correlated to an improved disease-specific survival (p = 0.034).ConclusionsOur study demonstrates antagonistic regulatory effects of HNRNPG and HTRA2-BETA1 on ERa exon7 splicing with potential impact on type 1 EC clinical outcome due to the consecutively variable expression levels of the ERa isoform D7.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-015-1088-1) contains supplementary material, which is available to authorized users.
Highlights
Estrogen receptor alpha (ERa/Estrogen receptor 1 (ESR1)) expression is regulated by alternative splicing
HTRA2-BETA1 and HNRNPG as antagonistic regulators of ERa exon7 splicing Functional experiments in Ishikawa endometrial cancer cells with transient transfection of HTRA2-BETA1 and HNRNPG expression plasmids revealed that endogenous exon7 inclusion was induced by HTRA2BETA1
Induced Estrogen receptor alpha delta 7 (ERaD7) expression is correlated to favorable clinico-pathological parameters In type 1 endometrial cancer (EC), ERa mRNA expression could be detected in 87 samples (75%) of which 71 (61.2% of the complete cohort) expressed the ERaD7 isoform
Summary
Estrogen receptor alpha (ERa/ESR1) expression is regulated by alternative splicing. Its most frequently detectable exon skipping isoform (ERaD7) is a dominant negative variant. The ERa exon skipping (ERaD7) isoform has been identified as the most common phenotype in EC and breast cancer and encodes for a protein lacking a portion of the hormone binding domain [7,10] This isoform represents a dominant negative variant for both ERa and ER beta [7,10]. Induced ERaD7 expression has been detected in the proliferative compared to the secretory phase of endometrial tissue [11] and in well to moderately differentiated EC in comparison to poorly differentiated EC [12] Besides these findings and an influence on estrogen therapy sensitivity in schizophrenic patients [13], the clinical significance of ERaD7 in estrogen related cancer has not been elucidated yet. Their antagonizing effects on alternative splicing processes were directly reflected by their opposite effects on EC biology
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