Mutational probing of the forkhead domain of the transcription factor FOXL2 provides insights into the pathogenicity of naturally occurring mutations

• Published in: Human molecular genetics Vol. 20; no. 17; pp. 3376 - 3385
• Main Authors: Todeschini, Anne-Laure, Dipietromaria, Aurélie, L'Hôte, David, Boucham, Fatima Zohra, Georges, Adrien B., Pandaranayaka, P.J. Eswari, Krishnaswamy, Sankaran, Rivals, Isabelle, Bazin, Claude, Veitia, Reiner A.
• Format: Journal Article
• Language: English
• Published: Oxford Oxford University Press 01-09-2011; Oxford University Press (OUP)
• Subjects: Amino acid substitution; Animals; Biochemistry, Molecular Biology; Biological and medical sciences; Cell Line; Cell physiology; Cercopithecus aethiops; COS Cells; Forkhead Box Protein L2; Forkhead Transcription Factors - chemistry; Forkhead Transcription Factors - genetics; Forkhead Transcription Factors - metabolism; Fundamental and applied biological sciences. Psychology; Genetics; Genetics of eukaryotes. Biological and molecular evolution; Humans; Life Sciences; Molecular and cellular biology; Molecular biology; Mutation; Protein Structure, Tertiary; Signal transduction; Structure-Activity Relationship; Pathogenicity; Signal transduction; Dysgenesia gonad; Malformation; Genetics; Sexual differentiation disorder; Mutation; Transcription factor; Genital diseases
• ISSN: 0964-6906; 1460-2083
• DOI: 10.1093/hmg/ddr244

Mutations of the transcription factor FOXL2, involved in cranio-facial and ovarian development, lead to the Blepharophimosis Syndrome. Here, we have systematically replaced the amino acids of the helices of the forkhead domain (FHD) of FOXL2 by glycine residues to assess the impact of such substitutions. A number of mutations lead to protein mislocalization, aggregation and to partial or complete loss of transactivation ability on a series of luciferase reporter systems. To rationalize the results of this glycine mutation scan, we have modeled the structure of the FHD by comparison with crystallographic data available for other FHDs. We failed to detect a clear-cut correlation between protein mislocalization or aggregation and the position of the mutation. However, we found that the localization of the side chain of each amino acid strongly correlated with the impact of its mutation on FOXL2 transactivation capacity. Indeed, when the side chains of the amino acids involved in the helices of the forkhead are supposed to point towards the hydrophobic core formed by the three main helices, a loss of function was observed. On the contrary, if the side chains point outward the hydrophobic core, protein function was preserved. The extension of this analysis to natural mutants shows that a similar correlation can be found for BPES mutations associated or not with ovarian dysfunction. Our findings reveal new insights into the molecular effects of FOXL2 mutations affecting the FHD, which represent two-thirds of intragenic mutations, and provide the first  predictive tool of their effects.