Novel application of luciferase assay for the in vitro functional assessment of KAL1 variants in three females with septo-optic dysplasia (SOD).

Abstract

KAL1 is implicated in 5% of Kallmann syndrome cases, a disorder which genotypically overlaps with septo-optic dysplasia (SOD). To date, a reporter-based assay to assess the functional consequences of KAL1 mutations is lacking. We aimed to develop a luciferase assay for novel application to functional assessment of rare KAL1 mutations detected in a screen of 422 patients with SOD. Quantitative analysis was performed using L6-myoblasts stably expressing FGFR1, transfected with a luciferase-reporter vector containing elements of the FGF-responsive osteocalcin promoter. The two variants assayed [p.K185N, p.P291T], were detected in three females with SOD (presenting with optic nerve hypoplasia, midline and pituitary defects). Our novel assay revealed significant decreases in transcriptional activity [p.K185N: 21% (p < 0.01); p.P291T: 40% (p < 0.001)]. Our luciferase-reporter assay, developed for assessment of KAL1 mutations, determined that two variants in females with hypopituitarism/SOD are loss-of-function; demonstrating that this assay is suitable for quantitative assessment of mutations in this gene.

• Authors: Mark J McCabe, Youli Hu, Louise C Gregory, Carles Gaston-Massuet, Kyriaki S Alatzoglou, José W Saldanha, Angelica Gualtieri, Ajay Thankamony, Ieuan Hughes, Sharron Townshend, Juan-Pedro Martinez-Barbera, Pierre-Marc Bouloux, Mehul T Dattani
• Journal: Molecular and cellular endocrinology (Mol Cell Endocrinol) Vol. 417 Pg. 63-72 (Dec 05 2015) ISSN: 1872-8057 [Electronic] Ireland
• PMID: 26375424 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2015 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

Chemical References

• ANOS1 protein, human
• Extracellular Matrix Proteins
• Nerve Tissue Proteins
• Luciferases

Topics

• Animals
• COS Cells
• Chlorocebus aethiops
• Extracellular Matrix Proteins (genetics, metabolism)
• Female
• Humans
• In Vitro Techniques
• Luciferases (metabolism)
• Models, Molecular
• Nerve Tissue Proteins (genetics, metabolism)
• Pedigree
• Pituitary Gland (metabolism)
• Polymorphism, Single Nucleotide
• Septo-Optic Dysplasia (genetics, metabolism, pathology)