Aromatase is a direct target of FOXL2: C134W in granulosa cell tumors via a single highly conserved binding site in the ovarian specific promoter.

Abstract

BACKGROUND:

Granulosa cell tumors (GCT) of the ovary often express aromatase and synthesize estrogen, which in turn may influence their progression. Recently a specific point mutation (C134W) in the FOXL2 protein was identified in >94% of adult-type GCT and it is likely to contribute to their development. A number of genes are known to be regulated by FOXL2, including aromatase/CYP19A1, but it is unclear which are direct targets and whether the C134W mutation alters their regulation. Recently, it has been reported that FOXL2 forms a complex with steroidogenic factor 1 (SF-1) which is a known regulator of aromatase in granulosa cells.

METHODOLOGY/PRINCIPAL FINDINGS:

In this work, the human GCT-derived cell lines KGN and COV434 were heterozygous and wildtype for the FOXL2:C134W mutation, respectively. KGN had abundant FOXL2 mRNA expression but it was not expressed in COV434. Expression of exogenous FOXL2:C134W in COV434 cells induced higher expression of a luciferase reporter for the ovarian specific aromatase promoter, promoter II (PII) (-516bp) than expression of wildtype FOXL2, but did not alter induction of a similar reporter for the steroidogenic acute regulatory protein (StAR) promoter (-1300bp). Co-immunoprecipitation confirmed that FOXL2 bound SF-1 and that it also bound its homologue, liver receptor homologue 1 (LRH-1), however, the C134W mutation did not alter these interactions or induce a selective binding of the proteins. A highly conserved putative binding site for FOXL2 was identified in PII. FOXL2 was demonstrated to bind the site by electrophoretic mobility shift assays (EMSA) and site-directed mutagenesis of this element blocked its differential induction by wildtype FOXL2 and FOXL2:C134W.

CONCLUSIONS/SIGNIFICANCE:

These findings suggest that aromatase is a direct target of FOXL2:C134W in adult-type GCT via a single distinctive and highly conserved binding site in PII and therefore provide insight into the pathogenic mechanism of this mutation.

• Authors: Nicholas I Fleming, Kevin C Knower, Kyren A Lazarus, Peter J Fuller, Evan R Simpson, Colin D Clyne
• Journal: PloS one (PLoS One) Vol. 5 Issue 12 Pg. e14389 (Dec 20 2010) ISSN: 1932-6203 [Electronic] United States
• PMID: 21188138 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, Non-P.H.S.)

Chemical References

• FOXL2 protein, human
• Forkhead Box Protein L2
• Forkhead Transcription Factors
• Phosphoproteins
• steroidogenic acute regulatory protein
• Aromatase

Topics

• Aromatase (metabolism)
• Binding Sites
• Cell Line, Tumor
• Female
• Forkhead Box Protein L2
• Forkhead Transcription Factors (genetics)
• Gene Expression Regulation, Neoplastic
• Granulosa Cell Tumor (metabolism)
• Humans
• Mutagenesis, Site-Directed
• Mutation
• Ovarian Neoplasms (genetics)
• Phosphoproteins (genetics)
• Point Mutation
• Promoter Regions, Genetic
• Regulatory Sequences, Nucleic Acid