A novel HESX1 splice mutation causes isolated GH deficiency by interfering with mRNA processing

Area of Science:

• Genetics
• Molecular Biology
• Endocrinology

Background:

• Mutations in the HESX1 gene are a rare cause of growth hormone deficiency (GHD).
• HESX1 mutations can be associated with various other developmental anomalies.
• Investigating HESX1 is crucial for understanding complex GHD etiologies.

Purpose of the Study:

• To identify causative mutations in the HESX1 gene within a cohort of Italian patients with GHD.
• To investigate the functional consequences of identified HESX1 mutations on mRNA splicing and protein expression.
• To elucidate the regulatory mechanisms of HESX1 expression involving alternative splicing and nonsense-mediated mRNA decay (NMD).

Main Methods:

• Screening of the HESX1 gene's coding region and exon-intron boundaries using denaturing HPLC.
• In vitro and in vivo mRNA analysis to characterize splicing patterns of wild-type and mutant HESX1 alleles.
• In vitro assay to assess the stability of aberrant mRNA transcripts, specifically focusing on nonsense-mediated mRNA decay (NMD).

Main Results:

• A novel heterozygous mutation (c.357+3G>A) near the intron 2 splice site was identified in an isolated GHD patient.
• The mutation altered HESX1 splicing, preventing the generation of an exon 2-lacking isoform and leading to the production of truncated protein variants.
• The exon 2-deleted mRNA was found to be degraded via nonsense-mediated mRNA decay (NMD).

Conclusions:

• The c.357+3G>A mutation disrupts normal HESX1 alternative splicing and leads to a truncated protein, likely causing isolated GHD.
• The study demonstrates that the interplay between alternative splicing and NMD is critical for regulating HESX1 expression.
• Alterations in this regulatory mechanism can have severe consequences, underscoring the importance of HESX1 gene regulation in human development.