Single-cell RNA-Sequencing Co-Expression Analysis with CFTR in Lung Tissue

Abstract

Background: While cystic fibrosis is caused by loss-of-function variants in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR), other modifier genes have been shown to associate with disease severity. Co-expression of modifiers with CFTR in normal tissue indicates a cooperative relationship and suggests the potential for compensation in the presence of CFTR dysfunction. We examined the co-expression relationships with CFTR in the lung using single cell RNA sequencing to pinpoint cell types and their modifiers involved in the forced expiratory volume in 1 second (FEV1)-based cystic fibrosis lung phenotype and support target cell-type prioritization for therapy 1. Methods: SmartSeq2 single cell RNA sequencing data from non-cystic fibrosis lung tissue was used for evaluation of co-expression with CFTR and modifier genes. Zero-inflated negative binomial model was used to formally test the co-expression association. 10X Chromium based single cell RNA sequencing data from both cystic fibrosis and non-cystic fibrosis studies were assessed graphically to confirm conclusions from the SmartSeq2 primary analysis. Results: Differentiating basal, club and alveolar epithelial type 2 cells were found to have high proportions of cells expressing CFTR as well as the greatest number of significant co-expression relationships with the modifiers. In particular, among alveolar epithelial type 2 cells, we observed a strong co-expression trio relationship between CFTR, SLC6A14 and SLC26A9 (p < 0.05). Conclusions: CFTR-modifier gene co-expression suggests basal, club and alveolar epithelial type 2 cells show coordinated expression. Alveolar epithelial type 2 cells showed strong co-expression evidence with two of the most established cystic fibrosis modifier genes.