Three Complex alleles associated with N1303K mutation and their molecular consequences

Area of Science:

• Genetics and Molecular Biology
• Medical Genetics

Background:

• Cystic Fibrosis (CF) exhibits distinct CFTR gene mutation patterns in Arab Mediterranean countries compared to other populations.
• The CFTR c.3909C>G (N1303K) mutation is more prevalent in Mediterranean regions, often linked with other genetic variations forming complex alleles.
• N1303K is associated with varied pancreatic insufficiency phenotypes and co-occurs with cis-acting polymorphisms.

Purpose of the Study:

• To identify and characterize complex alleles carrying the N1303K mutation in Lebanese, Egyptian, and French CF patients.
• To investigate the functional impact of these complex alleles on CFTR gene expression and protein function.

Main Methods:

• Genomic DNA analysis using PCR amplification and automated DNA sequencing of all CFTR exons and flanking regions.
• Minigene splicing assays in various cell lines (HeLa, HT29, HEK293) to assess mRNA splicing alterations.
• RT-PCR analysis of mRNA to evaluate the quality and quantity of CFTR protein produced.

Main Results:

• Two common complex alleles (wild-type and mutated haplotypes) and two rare complex alleles were identified.
• The N1303K mutation and its associated cis-polymorphisms were found to induce subtle abnormalities in CFTR mRNA splicing.
• These splicing alterations resulted in modifications to both the quality and quantity of the CFTR protein.

Conclusions:

• The identified complex alleles carrying the N1303K mutation contribute to altered CFTR protein function through aberrant splicing.
• The specific combinations of polymorphisms associated with N1303K in cis may influence the clinical severity of Cystic Fibrosis.
• Further research into these complex alleles is warranted to understand their full impact on CF pathogenesis in the Mediterranean region.