Posttranscriptional Gene Regulatory Networks in Chronic Airway Inflammatory Diseases: In silico Mapping of RNA-Binding Protein Expression in Airway Epithelium.

Luca Ricciardi,Annunziata Nigro,Giorgio Giurato,Alessandro Vatrella,Domenico Memoli,Mariagrazia Pietrafesa,Gaetano Caramori,Vincenzo Casolaro,Jessica Dal Col,Ilaria Salvato,Cristiana Stellato

doi:10.3389/fimmu.2020.579889

Abstract

Background: Posttranscriptional gene regulation (PTGR) contributes to inflammation through alterations in messenger RNA (mRNA) turnover and translation rates. RNA-binding proteins (RBPs) coordinate these processes but their role in lung inflammatory diseases is ill-defined. We evaluated the expression of a curated list of mRNA-binding RBPs (mRBPs) in selected Gene Expression Omnibus (GEO) transcriptomic databases of airway epithelium isolated from chronic obstructive pulmonary disease (COPD), severe asthma (SA) and matched control subjects, hypothesizing that global changes in mRBPs expression could be used to infer their pathogenetic roles and identify novel disease-related regulatory networks.Methods: A published list of 692 mRBPs [Nat Rev Genet 2014] was searched in GEO datasets originated from bronchial brushings of stable COPD patients (C), smokers (S), non-smokers (NS) controls with normal lung function (n = 6/12/12) (GEO ID: GSE5058) and of (SA) and healthy control (HC) (n = 6/12) (GSE63142). Fluorescence intensity data were extracted and normalized on the medians for fold change (FC) comparisons. FCs were set at ≥ |1.5| with a false discovery rate (FDR) of ≤ 0.05. Pearson correlation maps and heatmaps were generated using tMEV tools v4_9_0.45. DNA sequence motifs were searched using PScan-ChIP. Gene Ontology (GO) was performed with Ingenuity Pathway Analysis (IPA) tool.Results: Significant mRBP expression changes were detected for S/NS, COPD/NS and COPD/S (n = 41, 391, 382, respectively). Of those, 32% of genes changed by FC ≥ |1.5| in S/NS but more than 60% in COPD/NS and COPD/S (n = 13, 267, 257, respectively). Genes were predominantly downregulated in COPD/NS (n = 194, 73%) and COPD/S (n = 202, 79%), less so in S/NS (n = 4, 31%). Unsupervised cluster analysis identified in 4 out of 12 S the same mRBP pattern seen in C, postulating subclinical COPD. Significant DNA motifs enrichment for transcriptional regulation was found for downregulated RBPs. Correlation analysis identified five clusters of co-expressed mRBPs. GO analysis revealed significant enrichments in canonical pathways both specific and shared among comparisons. Unexpectedly, no significant mRBPs modulation was found in SA compared to controls.Conclusions: Airway epithelial mRBPs profiling reveals a COPD-specific global downregulation of RBPs shared by a subset of control smokers, the potential of functional cooperation by coexpressed RBPs and significant impact on relevant pathogenetic pathways in COPD. Elucidation of PTGR in COPD could identify disease biomarkers or pathways for therapeutic targeting.

Highlights

RNA-binding proteins (RBPs) are key regulatory factors in post-transcriptional gene regulation (PTGR) involved in the maturation, stability, transport and degradation of cellular RNAs
70% of the mRNA-binding RBPs (mRBPs) Differentially Expressed Genes (DEG) genes were included in this category when Chronic Obstructive Pulmonary Disease (COPD) samples were compared to both control samples (COPD/NS, COPD/S), while only 30% of smoking control group dataset (S/NS) DEG genes were upregulated over this threshold
The Significant FC/Differentially Expressed Genes (SDEG) profiles indicate that the majority of mRBP genes in COPD were downregulated compared to both controls (n = 194/267, 73% for COPD/NS; n = 202/257, 79% for COPD vs. S) while in S/NS only 30% (n = 4/13) were downregulated (Figure 3)

Summary

Introduction

RNA-binding proteins (RBPs) are key regulatory factors in post-transcriptional gene regulation (PTGR) involved in the maturation, stability, transport and degradation of cellular RNAs. Similar studies in human chronic inflammatory diseases are lagging behind, despite ample knowledge of deregulated PTGR in inflammatory responses by in vitro studies and the strong inflammatory phenotypes obtained in some of the knockout animal models [8, 9] This knowledge chasm is present for lung diseases, despite the strong link between chronic inflammatory diseases such as Chronic Obstructive Pulmonary Disease (COPD) and some lung cancer types [10] and the extensive overlap of RBP-regulated genes contributing to both disease process [11]. We evaluated the expression of a curated list of mRNA-binding RBPs (mRBPs) in selected Gene Expression Omnibus (GEO) transcriptomic databases of airway epithelium isolated from chronic obstructive pulmonary disease (COPD), severe asthma (SA) and matched control subjects, hypothesizing that global changes in mRBPs expression could be used to infer their pathogenetic roles and identify novel disease-related regulatory networks

Methods

Results

Conclusion