ABSTRACT The overall goal of our proposed research is to identify genes and transcripts associated with MUC5B expression that contribute to the development of idiopathic pulmonary fibrosis (IPF). Rare mutations (16- 21) and common variants (22-26) are associated with IPF and account for at least 40% of the risk of developing this disease. In the past 5 years, we have found that: 1) a gain-of-function MUC5B promoter variant rs35705950 is the strongest risk factor for the development of IPF (22, 26-34); 2) epigenetic mechanisms affect the expression of MUC5B (35); 3) IPF is a complex genetic disease with rare and common variants contributing to the development of this disease, including pronounced changes in DNA methylation (36) and transcriptional subtypes (37); and 4) MUC5B appears to be involved in the pathogenesis of IPF (27, 38-40). However, there is no clear explanation for the low penetrance of the MUC5B promoter variant; while the minor allele of MUC5B is present in the heterozygous or homozygous state in ≈19% of the general population (27), IPF occurs in far less than 1% of the population (41, 42). These observations lead us to postulate that the etiology of IPF will best be understood by identifying the genes and transcripts that control MUC5B expression and contribute to the development of pulmonary fibrosis. To generate a feasible experimental model to pursue this concept, we phenotyped the response to intratracheal bleomycin in the eight founder mouse strains that were used to create the Diversity Outbred (DO) mouse population (43) and found that although there is a clear relationship between Muc5b RNA and MUC5B protein expression and bleomycin-induced lung fibrosis, this relationship is not consistent across the eight founder DO mouse strains. These preliminary findings suggest that the DO mouse population can be used to integrate genetic variation with gene expression to create multi-scale models of bleomycin-induced lung fibrosis and then use this knowledge to understand the genetic basis of MUC5B-induced IPF. Based on these observations, we hypothesize that genes and transcripts that control bleomycin- induced Muc5b/MUC5B expression contribute to the risk of developing pulmonary fibrosis. In this project, we plan to phenotype and genotype 900 DO mice for their response to bleomycin to identify the genes that control MUC5B protein expression and contribute to the development of lung fibrosis (Aim 1), while also identifying transcriptional changes (including Muc5b transcript) associated with bleomycin-induced lung fibrosis (Aim 2). We will then determine whether these candidate genes and transcriptional changes identified in mice exposed to bleomycin are generalizable to IPF (Aim 3). The successful completion of these Aims will have broad impact, resulting in specific genetic targets and biologic pathways for use in the design of future preventive, mechanistic, and intervention studies of IPF.

NARRATIVE The overall goal of our proposed research is to identify genes and transcripts associated with MUC5B expression that contribute to the development of idiopathic pulmonary fibrosis (IPF). The paradox that we have chosen to address in this proposal is that while the MUC5B promoter variant rs35705950 is the strongest risk factor for the development of IPF, there is no convincing explanation for the low penetrance of this allele in relation to disease risk.