Principles of ribosome-associated protein quality control during the synthesis of CFTR

Area of Science:

• Molecular Biology
• Cell Biology
• Protein Quality Control

Background:

• The ribosome-associated quality control (RQC) pathway identifies and degrades proteins with prolonged translational arrests.
• Multipass transmembrane proteins are frequently targeted by the RQC, suggesting potential issues during their synthesis and membrane insertion.
• Understanding RQC's role in quality control for complex proteins like CFTR is crucial for cystic fibrosis research.

Purpose of the Study:

• To investigate the role of the RQC pathway in the quality control of CFTR, a large transmembrane protein.
• To determine if defects in transmembrane domain insertion and assembly of CFTR trigger RQC-mediated translational arrests.
• To explore the mechanisms by which RQC responds to challenges in synthesizing transmembrane proteins.

Main Methods:

• Utilized reporter assays in HEK293 cells to monitor CFTR translation and RQC activation.
• Manipulated CFTR folding and membrane insertion to assess their impact on RQC response.
• Investigated the influence of translation kinetics regulators (codon usage, GCN1, SRP) on CFTR translation arrest.

Main Results:

• A subset of nascent CFTR initiated RQC activation during translation in HEK293 cells.
• Interventions disrupting CFTR folding or membrane insertion did not increase RQC activation.
• CFTR translation arrest was largely independent of codon usage, GCN1, and the SRP complex.

Conclusions:

• The RQC pathway is activated by inherent difficulties during transmembrane segment synthesis, not solely by protein misfolding or insertion defects.
• RQC plays a novel physiological role in managing elongation-arrested transmembrane proteins.
• This finding offers new insights into protein quality control mechanisms for membrane proteins.