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. 2025 Jun 17;10(14):e189636.
doi: 10.1172/jci.insight.189636. eCollection 2025 Jul 22.

The MUC5B promoter variant results in proteomic changes in the nonfibrotic lung

Jeremy A Herrera  1 , Mark Maslanka  2 , Rachel Z Blumhagen  3 , Rachel Blomberg  4 , Nyan Ye Lwin  1 , Janna Brancato  1 , Carlyne D Cool  5 , Jonathan P Huber  1 , Jonathan S Kurche  1   6 , Chelsea M Magin  1   4   7 , Kirk C Hansen  2 , Ivana V Yang  1   8 , David A Schwartz  1   6   9
Affiliations

The MUC5B promoter variant results in proteomic changes in the nonfibrotic lung

Jeremy A Herrera et al. JCI Insight. .

Abstract

The gain-of-function MUC5B promoter variant is the dominant risk factor for the development of idiopathic pulmonary fibrosis (IPF). However, its impact on protein expression in both nonfibrotic control and IPF lung specimens has not been well characterized. Utilizing laser capture microdissection coupled to mass spectrometry, we investigated the proteomic profiles of airway and alveolar epithelium in nonfibrotic controls (n = 12) and IPF specimens (n = 12), stratified by the MUC5B promoter variant. Through qualitative and quantitative analyses, as well as pathway analysis and immunohistological validation, we have identified a distinct MUC5B-associated protein profile. Notably, the nonfibrotic control alveoli exhibited substantial MUC5B-associated protein changes, with an increase in IL-3 signaling. Additionally, we found that epithelial cells overlying IPF fibroblastic foci clustered closely to alveolar epithelia and expressed proteins associated with cellular stress pathways. In conclusion, our findings suggest that the MUC5B promoter variant leads to protein changes in alveolar and airway epithelium that appear to be associated with initiation and progression of lung fibrosis.

Keywords: Fibrosis; Genetics; Proteomics; Pulmonology.

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Conflict of interest statement

Conflict of interest: DAS is the founder and unpaid chief scientific officer of Eleven P15, Inc., a company dedicated to the early diagnosis and treatment of lung fibrosis. DAS also serves as a consultant for Vertex Pharmaceuticals. IVY and CDC report consulting fees from Eleven P15, Inc. RZB, JPH, and JB report salary support on a sponsored research agreement from Eleven P15, Inc., outside the submitted work. CMM is a member of the board of directors for the Colorado BioScience Institute.

Figures

Figure 1
Figure 1. Impact of the MUC5B promoter variant on nonfibrotic control lung.
(A and B) Volcano plots comparing the MUC5B promoter variant and WT allele in nonfibrotic control (A) small airways and (B) alveoli, showing the negative natural log of adjusted P values (0.05) for each protein. (C) Representative images for the IHC against phosphorylated STAT5A at Tyr694 on nonfibrotic control lungs. Scale bars: 50 μm. (D) Quantification of C showing the percentage of positive cells (n = 12, balanced for MUC5B; 5 fields of view per patient with a total of 22,999 counted nuclei). **P < 0.005 by unpaired, 2-tailed t test.
Figure 2
Figure 2. The histopathological lesions of IPF uniquely cluster.
A hierarchical cluster analysis based on the top 1,000 variable proteins. For IPF specimens (n = 12), we collected 12 small airway, 11 honeycomb cyst, 12 epithelia overlying fibroblastic foci (FF), 12 alveoli, and 12 FF samples from the same 12 IPF specimens. We further collected an additional 8 FF samples (a total of 20 FF samples) from 8 additional IPF specimens. For nonfibrotic control specimens (n = 12), we collected 12 small airway and 12 alveoli samples from the same specimen. The samples were balanced for the MUC5B promoter variant.
Figure 3
Figure 3. The unaffected epithelia (airway and alveoli) of IPF are abnormal.
(A and C) Volcano plots comparing (A) IPF small airway and nonfibrotic control small airways and (C) IPF alveoli and nonfibrotic control alveoli, showing the negative natural log of adjusted P values plotted against the log2(fold change) for each protein. (B and D) Focusing on the significantly changed proteins from A and C, we reanalyzed the data for the MUC5B promoter variant and show volcano plots for (B) IPF small airways and (D) IPF alveoli. n = 12 per group (6 WT and 6 MUC5B promoter variant).
Figure 4
Figure 4. IPF honeycomb cysts are defined by remodeling of epithelial adherens junctions.
(A) Volcano plot comparing IPF honeycomb cyst (HC) and IPF small airways. (B) The subset of proteins associated with “remodeling of epithelial adherens junctions,” showing the negative natural log of adjusted P values plotted against the log2(fold change) for each protein. (C) A heatmap displaying z scores for the significantly changed “remodeling of epithelial adherens junctions” proteins, comparing IPF HC, IPF small airways, and nonfibrotic control small airways. (D and E) Volcano plots comparing MUC5B promoter variant and WT allele in (D) the subset of significantly changed IPF HC proteins in A and (E) the subset of all significantly changed airway proteins in IPF and nonfibrotic controls while controlling for region. n = 12 per group (6 WT and 6 MUC5B promoter variant).
Figure 5
Figure 5. Airway expression of adhesion-associated proteins CD9 and myoferlin.
Representative IHC staining for mucin 5B (MUC5B), CD9, and myoferlin (MYOF) in (A) IPF honeycomb cyst (red arrows), (B) IPF small airway, and (C) nonfibrotic control small airway. Scale bars: 20 μm. Quantification of the IHC staining for (D) CD9 and (E) MYOF positivity per airway (n = 6 nonfibrotic control and n = 6 IPF specimens). **P < 0.005; ***P < 0.0005; ****P < 0.0001 by 1-way ANOVA.
Figure 6
Figure 6. IPF fibroblastic foci (FF) are defined by increased translational control.
(A, B, and D) Volcano plots comparing (A) IPF FF versus IPF alveoli, (B) the subset of proteins associated with “eukaryotic translation initiation,” and (D) the subset of significantly changed proteins from A reanalyzed for the MUC5B promoter variant showing the negative natural log of adjusted P values plotted against the log2(fold change) for each protein. (C) A heatmap displaying z scores of the significantly changed “eukaryotic translation initiation” proteins comparing IPF FF, IPF alveoli, and nonfibrotic control alveoli. n = 12 per group (n = 20 IPF FF, balanced for the MUC5B promoter variant).
Figure 7
Figure 7. IPF epithelia overlying the fibroblastic foci (FF) are defined by cell stress pathways.
(A) Venn diagram of detected proteins in the IPF epithelia overlying FF as compared with IPF and nonfibrotic control alveoli. (B, C, and E) Volcano plots comparing (B) IPF epithelia overlying the FF and IPF alveoli, (C) the subset of proteins associated with “eIF2 signaling,” and (E) the subset of significantly changed proteins in B reassessed for the MUC5B promoter variant, showing the negative natural log of adjusted P values plotted against the log2(fold change) for each protein. (D) A heatmap displaying z scores of the significantly changed “eIF2 signaling” proteins comparing IPF epithelia overlying FF, IPF alveoli, and nonfibrotic control alveoli. n = 12 per group (6 WT and 6 MUC5B promoter variant).
Figure 8
Figure 8. IPF epithelia overlying fibroblastic foci (FF) express stress markers.
Representative images of IHC staining for cytokeratin 17, p-eIF2α (serine 51), ATF4, and ATF6 in (A) IPF epithelia (black arrows) overlying a fibroblastic foci (red asterisk), (B) IPF alveoli, and (C) nonfibrotic control alveoli. Scale bars: 50 μm.
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