LIM-homeodomain transcription factor, Lhx2, is involved in transcriptional control of brain-specific promoter/exon 1f of the mouse aromatase gene

Abstract

Neurosteroidal oestrogen has been proposed to play important roles in a variety of reproductive behaviours. Aromatase, a key enzyme in oestrogen synthesis, is localised in neural nuclei of specific brain regions and is developmentally regulated, with a transient expression peak at the perinatal period. The brain-specific promoter of the aromatase gene was analysed aiming to determine the transcriptional control mechanisms that could help explain the spatiotemporal expression. We previously reported that a 202-bp sequence, which is upstream from the transcriptional initiation site, is essential for the basal transcriptional activity. The 202-bp upstream region of brain-specific exon 1 comprises at least three types of cis-acting elements: aro-AI (Arom-Aα), aro-AII (Arom-Aβ) and aro-B (Arom-B). To identify the binding proteins for the cis-acting elements, a yeast one-hybrid screen was performed with these cis-element sequences using a mouse foetal cDNA library. Lhx2, a LIM-homeodomain protein, was identified as one of the aro-B binding proteins. The identification was further confirmed using the gel shift assay, which demonstrated binding competition of nuclear proteins to the aro-B element with a typical Lhx2-binding element. In addition, a chromatin immunoprecipitation assay with an anti-Lhx2 antibody demonstrated that Lhx2 bound to the aro-B site in vivo. A reporter assay of the brain-specific promoter demonstrated increased Lhx2-dependent promoter activity. Furthermore, the time-dependent increase in aromatase mRNA in primary cultured foetal neurones was suppressed by an small-interfering RNA-mediated knockdown of Lhx2 expression. These results show that Lhx2 is involved in the transcriptional regulation of aromatase in the rodent brain.