Low temperature restoring effect on F508del-CFTR misprocessing: A proteomic approach.

Area of Science:

• Proteomics
• Cellular Biology
• Molecular Medicine

Background:

• The F508del mutation is the most common cause of cystic fibrosis (CF), leading to misfolded CFTR protein.
• Low-temperature treatment is known to partially rescue F508del-CFTR function, but the underlying protein mechanisms are not fully understood.

Purpose of the Study:

• To identify proteins involved in the low-temperature-induced rescue of F508del-CFTR.
• To elucidate the cellular responses contributing to improved F508del-CFTR processing at reduced temperatures.

Main Methods:

• Two-dimensional electrophoresis (2DE) was used to analyze the proteome of BHK cells expressing wild-type (wt) or F508del-CFTR.
• Cells were cultured at 37°C or 26°C for 24-48 hours, followed by metabolic labeling with [(35)S]-methionine.
• Mass spectrometry was employed to identify differentially expressed proteins among various treatment groups.

Main Results:

• 139 differentially expressed protein spots were identified, with 125 successfully identified by mass spectrometry.
• Cold-shock induced unfolded protein response (UPR) and repressed cell metabolism, creating a favorable environment for F508del-CFTR rescue.
• Down-regulation of proteasome regulatory PA28 and COP9 signalosome subunits, implicated in CFTR degradation, was observed.
• Cold-shock partially repaired the deregulation of RACK1, a protein crucial for CFTR plasma membrane stability.

Conclusions:

• Low-temperature treatment induces UPR and modulates cell metabolism, contributing to F508del-CFTR rescue.
• Modulation of proteins involved in CFTR degradation and stabilization, such as PA28, COP9, and RACK1, are key cold-shock effects.
• Identified proteins represent potential therapeutic targets for enhancing F508del-CFTR function in cystic fibrosis.