Cystic Fibrosis Receptor is Actively Exported within Extracellular Vesicles: A New Therapeutic Strategy

Abstract

Cystic fibrosis (CF) is a genetic disorder caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. This gene encodes an ion channel that regulates chloride (Cl^-) and water transport, and its dysfunction leads to mucus build up and multi-organ damage. While phosphorylation is known to open the CFTR channel for Cl^-transport, it remains unknown whether extracellular vesicles (EVs) traffic this active, phosphorylated form of the protein. EV transport of active CFTR channels could be a novel therapeutic for CFTR deficient cells. This study aims to determine if the p-CFTR protein is present within EVs across various cell lines. We analyzed mass spectrometry data from 3 different cell lines mesenchymal stem cell (MSC) EVs, treated ovarian carcinoma (OvCar) EVs and OvCar Human interstitial fluid (IF) Tissue EVs to detect CFTR-related protein presence in EVs. Next, we performed Western blotting on the 4T1 cell lysate and EVs using a p-CFTR(S737) antibody treated with oxidative stress (CCCP). The results demonstrate that MSC EVs and OvCar IFEVs both contain a higher abundance of CFTR-interacting proteins compared to OvCar EVs. Specifically, MSC EVs and OvCar IFEVs showed up to six commonly interacting CFTR proteins in some files, whereas OvCar EVs showed few (<1). Western blot analysis confirmed the presence of p-CFTR, PINK1, CD81, and MAPKK2 within 4T1 EVs with increased expression in CCCP treated EVs. The entire signaling cascade was within the EV. Altogether, these results represent a novel treatment plan for CFTR deficient patients receiving p-CFTR+ EVs to rescue normal CFTR.