Mucus hyper-secretion is a hallmark feature of asthma and other muco-obstructive airway diseases. The mucin proteins MUC5AC and MUC5B are the major glycoprotein components of mucus and have critical roles in airway defense. Despite the biomedical importance of these two proteins, the loci that regulate them in the context of natural genetic variation have not been studied. To identify genes that underlie variation in airway mucin levels, we performed genetic analyses in founder strains and incipient lines of the Collaborative Cross (CC) in a house dust mite mouse model of asthma. CC founder strains exhibited significant differences in MUC5AC and MUC5B, providing evidence of heritability. Analysis of gene and protein expression of and in incipient CC lines ( = 154) suggested that post-transcriptional events were important regulators of mucin protein content in the airways. Quantitative trait locus (QTL) mapping identified distinct, protein QTL for MUC5AC (chromosome 13) and MUC5B (chromosome 2). These two QTL explained 18 and 20% of phenotypic variance, respectively. Examination of the MUC5B QTL allele effects and subsequent phylogenetic analysis allowed us to narrow the MUC5B QTL and identify as a candidate gene. mRNA and protein expression were upregulated in parallel to MUC5B after allergen challenge, and knockout mice exhibited higher MUC5B expression. Thus, BPIFB1 is a novel regulator of MUC5B.

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Background Transcriptomic data has demonstrated utility to advance the study of physiological diversity and organisms’ responses to environmental stressors. However, a lack of genomic resources and challenges associated with collecting high-quality RNA can limit its application for many wild populations. Minimally invasive blood sampling combined with de novo transcriptomic approaches has great potential to alleviate these barriers. Here, we advance these goals for marine turtles by generating high quality de novo blood transcriptome assemblies to characterize functional diversity and compare global transcriptional profiles between tissues, species, and foraging aggregations. Results We generated high quality blood transcriptome assemblies for hawksbill (Eretmochelys imbricata), loggerhead (Caretta caretta), green (Chelonia mydas), and leatherback (Dermochelys coriacea) turtles. The functional diversity in assembled blood transcriptomes was comparable to those from more traditionally sampled tissues. A total of 31.3% of orthogroups identified were present in all four species, representing a core set of conserved genes expressed in blood and shared across marine turtle species. We observed strong species-specific expression of these genes, as well as distinct transcriptomic profiles between green turtle foraging aggregations that inhabit areas of greater or lesser anthropogenic disturbance. Conclusions Obtaining global gene expression data through non-lethal, minimally invasive sampling can greatly expand the applications of RNA-sequencing in protected long-lived species such as marine turtles. The distinct differences in gene expression signatures between species and foraging aggregations provide insight into the functional genomics underlying the diversity in this ancient vertebrate lineage. The transcriptomic resources generated here can be used in further studies examining the evolutionary ecology and anthropogenic impacts on marine turtles.

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15Airway mucociliary clearance (MCC) is required for host defense and often diminished in 16 chronic lung diseases. Effective clearance depends upon coordinated actions of the airway 17 epithelium and a mobile mucus layer. Dysregulation of the primary secreted airway mucin 18 proteins, MUC5B and MUC5AC, is associated with a reduction in the rate of MCC; however, 19 how other secreted proteins impact the integrity of the mucus layer and MCC remains unclear. 20 We previously identified the gene Bpifb1 as a regulator of the levels of MUC5B in the airways 21 using genetic approaches. Here, we show that BPIFB1 is required for normal mucociliary 22 clearance in vivo using Bpifb1 knockout (KO) mice. Reduced MCC in Bpifb1 KO mice occurred in 23 the absence of defects in sodium or chloride ion transport or changes in ciliary beat frequency. 24 We found that BPIFB1 loss resulted in airway mucus flakes with significantly increased complex 25 viscosity, a key biophysical property of mucus known to impact MCC. Finally, BPIFB1 protein 26 colocalized with MUC5B in secretory granules and was present in mucus on the airway surface. 27 Collectively, our findings demonstrate that BPIFB1 is an important component of the 28 mucociliary apparatus in mice and may be a key protein constituent of the mucus network. 29 30 31 42 particular, the concentration, organization, and post-translational modifications of MUC5AC 43 and MUC5B have been shown to alter biophysical properties of mucus that are required for 44

Acute ozone (O3) exposure is associated with multiple adverse cardiorespiratory outcomes, the severity of which varies across individuals in human populations and inbred mouse strains. However, molecular determinants of response, including susceptibility biomarkers that distinguish who will develop severe injury and inflammation, are not well characterized. We and others have demonstrated that airway macrophages (AMs) are an important resident immune cell type that are functionally and transcriptionally responsive to O3 inhalation. Here, we sought to explore influences of strain, exposure, and strain-by-O3 exposure interactions on AM gene expression and identify transcriptional correlates of O3-induced inflammation and injury across 6 mouse strains, including 5 Collaborative Cross (CC) strains. We exposed adult mice of both sexes to filtered air (FA) or 2 ppm O3 for 3 hours, and measured inflammatory and injury parameters 21 hours later. Mice exposed to O3 developed airway neutrophilia and lung injury with strain-dependent severity. In AMs, we identified a common core O3 response signature across all strains, as well as a set of genes exhibiting strain-by-O3 exposure interactions. In particular, a prominent gene expression contrast emerged between a low- (CC017/Unc) and high-responding (CC003/Unc) strain, as reflected by cellular inflammation and injury. Further inspection indicated that differences in their baseline gene expression and chromatin accessibility profiles likely contributes to their divergent post-O3 exposure transcriptional responses. Together, these results suggest that aspects of O3-induced respiratory responses are mediated through altered AM transcriptional signatures, and further confirms the importance of gene-environment interactions in mediating differential responsiveness to environmental agents.

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