CFTR Modulators Restore Acidification of Autophago-Lysosomes and Bacterial Clearance in Cystic Fibrosis Macrophages.

Asmaa Badr,Estelle Cormet-Boyaka,Xiaoli Zhang,Youssra Saqr,Mikhail A Gavrilin,Arwa Abu Khweek,Santiago Partida-Sanchez,Frank Robledo-Avila,Ahmad Hegazi,Mostafa Eltobgy,Kathrin Krause,Kylene P Daily,Amal O Amer,Cierra Carafice,Shady Estfanous,Stephanie Seveau,Tracey L Bonfield,Midhun N K Anne,Kaitlin Hamilton

doi:10.3389/fcimb.2022.819554

Abstract

Cystic fibrosis (CF) human and mouse macrophages are defective in their ability to clear bacteria such as Burkholderia cenocepacia. The autophagy process in CF (F508del) macrophages is halted, and the underlying mechanism remains unclear. Furthermore, the role of CFTR in maintaining the acidification of endosomal and lysosomal compartments in CF cells has been a subject of debate. Using 3D reconstruction of z-stack confocal images, we show that CFTR is recruited to LC3-labeled autophagosomes harboring B. cenocepacia. Using several complementary approaches, we report that CF macrophages display defective lysosomal acidification and degradative function for cargos destined to autophagosomes, whereas non-autophagosomal cargos are effectively degraded within acidic compartments. Notably, treatment of CF macrophages with CFTR modulators (tezacaftor/ivacaftor) improved the autophagy flux, lysosomal acidification and function, and bacterial clearance. In addition, CFTR modulators improved CFTR function as demonstrated by patch-clamp. In conclusion, CFTR regulates the acidification of a specific subset of lysosomes that specifically fuse with autophagosomes. Therefore, our study describes a new biological location and function for CFTR in autophago-lysosomes and clarifies the long-standing discrepancies in the field.

Highlights

Cystic fibrosis (CF) is one of the most common lethal autosomal recessive diseases (Paranjape and Mogayzel, 2014; Byrne et al, 2015)
We found that CFTR colocalized with LC3-labeled puncta in non-infected non-CF macrophages (Figure 1A)
The percent volume of LC3 colocalized with CFTR in non-infected cells had a mean of 48.6 and a standard error of the mean (SEM) of 5.46, and it increased significantly after 2 h of B. cenocepacia infection to reach a mean of 72.5 and a SEM of 7.68 (Supplemental Figure 1E)

Summary

Introduction

Cystic fibrosis (CF) is one of the most common lethal autosomal recessive diseases (Paranjape and Mogayzel, 2014; Byrne et al, 2015). CF patients carrying the F508del mutation experience infection with different respiratory pathogens including Pseudomonas aeruginosa (Hart and Winstanley, 2002; Del Porto et al, 2011; Bonfield et al, 2012), Staphylococcus aureus (Hart and Winstanley, 2002; Li et al, 2017), and Burkholderia cenocepacia (Scoffone et al, 2017). In healthy individuals, these pathogens can be cleared by robust autophagy activity within healthy immune cells (Abdulrahman et al, 2011) but can cause fatal infections in CF patients

Methods

Results

Conclusion