Synonymous codon usage affects the expression of wild type and F508del CFTR.

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• Cystic fibrosis is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene.
• The most common mutation, F508del, leads to protein misfolding and endoplasmic-reticulum-associated degradation.
• Synonymous codons, which do not change the amino acid sequence, can influence gene expression.

Purpose of the Study:

• To investigate the impact of synonymous codon usage on the expression of wild-type CFTR and the F508del mutant.
• To determine if codon optimization can improve the maturation and function of F508del-CFTR.

Main Methods:

• Expressing CFTR and F508del-CFTR with synonymous codon replacements in HEK293 cells using a heterologous vector.
• Analyzing mRNA levels, protein expression, and channel activity.
• Investigating the role of nonsense-mediated decay in regulating mRNA stability.

Main Results:

• Codon usage significantly altered CFTR mRNA steady-state levels (up to 30-fold variation) without affecting promoter strength or mRNA half-life.
• Nonsense-mediated decay, independent of the exon junction complex, was implicated in regulating mRNA levels.
• Native codons were more efficient for wild-type CFTR expression, but codon optimization improved F508del-CFTR folding, reducing ER-associated degradation and restoring plasma membrane localization and channel activity.

Conclusions:

• Synonymous codon optimization impacts CFTR mRNA levels and protein expression.
• Codon optimization can enhance the folding and function of the F508del-CFTR mutant, suggesting a potential therapeutic strategy.