Analysis of multiple gene co-expression networks to discover interactions favoring CFTR biogenesis and ΔF508-CFTR rescue

Background We previously reported that expression of a miR-138 mimic or knockdown of SIN3A in primary cultures of cystic fibrosis (CF) airway epithelia increased Delta F508-CFTR mRNA and protein levels, and partially restored CFTR-dependent chloride transport. Global mRNA transcript profiling in Delta F508-CFBE cells treated with miR-138 mimic or SIN3A siRNA identified two genes, SYVN1 and NEDD8, whose inhibition significantly increased Delta F508-CFTR trafficking, maturation, and function. Little is known regarding the dynamic changes in the CFTR gene network during such rescue events. We hypothesized that analysis of condition-specific gene networks from transcriptomic data characterizing Delta F508-CFTR rescue could help identify dynamic gene modules associated with CFTR biogenesis. Methods We applied a computational method, termed M-module, to analyze multiple gene networks, each of which exhibited differential activity compared to a baseline condition. In doing so, we identified both unique and shared gene pathways across multiple differential networks. To construct differential networks, gene expression data from CFBE cells were divided into three groups: (1) siRNA inhibition of NEDD8 and SYVN1; (2) miR-138 mimic and SIN3A siRNA; and (3) temperature (27 degrees C for 24 h, 40 degrees C for 24 h, and 27 degrees C for 24 h followed by 40 degrees C for 24 h). Results Interrogation of individual networks (e.g., NEDD8/SYVN1 network), combinations of two networks (e.g., NEDD8/SYVN1 + temperature networks), and all three networks yielded sets of 1-modules, 2-modules, and 3-modules, respectively. Gene ontology analysis revealed significant enrichment of dynamic modules in pathways including translation, protein metabolic/catabolic processes, protein complex assembly, and endocytosis. Candidate CFTR effectors identified in the analysis included CHURC1, GZF1, and RPL15, and siRNA-mediated knockdown of these genes partially restored CFTR-dependent transepithelial chloride current to Delta F508-CFBE cells. Conclusions The ability of the M-module to identify dynamic modules involved in Delta F508 rescue provides a novel approach for studying CFTR biogenesis and identifying candidate suppressors of Delta F508.

Automated summary

The study demonstrated that miR-138 mimic or SIN3A siRNA could increase Delta F508-CFTR levels and partially restore CFTR-dependent chloride transport. Gene network analysis in CFBE cells revealed that inhibition of NEDD8 and SYVN1 significantly improved Delta F508-CFTR trafficking, maturation, and function.