Native CFTR codon bias controls translation rate to balance off-pathway aggregation and channel function by conformational imprinting.

Protein folding in vivo is biologically tuned to minimize off-pathway events and optimize native folding outcomes. A key factor in this process is biased synonymous codon usage, in which synonymous codons modulate local translation rate while maintaining the native amino acid sequence. Here, we demonstrate that native codon usage within the first nucleotide-binding domain (NBD1) of the cystic fibrosis transmembrane conductance regulator (CFTR) induces a translational pause during a critical window of synthesis that affects CFTR folding, processing, and function. Eliminating this pause by substituting synonymous codons increased the aggregation propensity of immature CFTR and induced conformational and functional changes that persisted during CFTR processing and plasma membrane expression. Interestingly, the resulting mature CFTR protein at the plasma membrane exhibited enhanced ATP-dependent chloride channel gating. Thus, during protein synthesis, cotranslational events dictated by codon usage can imprint persistent conformational and functional properties upon CFTR. Our findings suggest that CFTR codon usage has evolved and adapted to balance a compromise between protein aggregation and a modest loss of channel function.