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Four naturally occurring mutations in the human GnRH receptor affect ligand binding and receptor function
Grégoy Y Bédécarrats1, Katja D Linher, Jo Ann Janovick
1Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, 221 Longwood Avenue, Boston, MA 02115, USA.
Molecular and Cellular Endocrinology
|August 2, 2003
Summary
Four GnRH receptor mutations cause idiopathic hypogonadotropic hypogonadism by impairing GnRH binding and signaling. These findings advance understanding of GnRHR function and related disorders.
Area of Science:
- Endocrinology
- Molecular Biology
- Genetics
Background:
- Idiopathic hypogonadotropic hypogonadism (IHH) is a disorder characterized by absent or delayed puberty.
- Mutations in the human GnRH receptor (GnRHR) gene are a known cause of IHH.
- Understanding the functional impact of these mutations is crucial for diagnosing and potentially treating IHH.
Purpose of the Study:
- To functionally characterize four novel GnRHR mutations found in IHH patients.
- To determine the effects of these mutations on GnRH binding, signal transduction, and gene promoter activity.
Main Methods:
- Site-directed mutagenesis to create GnRHR variants.
- Immunocytochemistry to assess receptor localization.
- GnRH binding assays.
- Inositol phosphate accumulation assays.
- Reporter gene assays for gonadotropin subunit and GnRHR promoter activity.
- ERK1 activation and CRE-luciferase assays.
Main Results:
- All four mutant GnRHRs were expressed on the cell surface.
- Mutant receptors showed impaired GnRH binding.
- Most mutants failed to induce significant inositol phosphate accumulation; Thr32Ile showed partial retention.
- Leu266Arg and Cys279Tyr mutants did not stimulate promoter activity; Cys200Tyr and Thr32Ile showed reduced maximal responses and increased EC50 values.
- Thr32Ile partially retained ERK1 activation and CRE-luciferase activity.
Conclusions:
- The Thr32Ile mutation impairs GnRHR function by reducing ligand binding affinity.
- The Cys200Tyr mutation affects GnRHR cell surface expression and signaling.
- All four identified GnRH receptor mutations interfere with GnRH signaling pathways, contributing to IHH pathogenesis.