Sex-stratified and interaction multi-omics analysis of DNA methylation and gene expression in asthma
- 1Indiana University school of medicine
Abstract
BackgroundSex differences in asthma are well recognized, but sex-associated airway DNA methylation (DNAm) patterns and their relationships with gene expression remain incompletely characterized. We therefore conducted cis- and trans-expression quantitative trait methylation (eQTM) analyses using sex-stratified and CpG-by-sex interaction models.
MethodsWe analyzed nasal airway epithelial DNAm and transcriptomic data from 298 participants in the CAAPA discovery cohort. Asthma-associated differentially methylated CpGs (DMCs) were identified using sex-combined and sex-stratified models. Cis- and trans-eQTM analyses were then performed in participants with asthma using sex-combined, sex-stratified, and CpG-by-sex interaction models. To prioritize asthma-associated regulatory relationships, eQTMs that were also significant in non-asthmatic controls were excluded from downstream analyses. Replication in an independent bronchial epithelial cohort was defined by concordant effect direction and P < 0.05. Replicated targets underwent pathway and network analyses.
FindingsAsthma-associated DMCs were predominantly hypomethylated in females and hypermethylated in males. Among trans-eQTMs, 379 associations were shared across all four models, whereas the majority were model-specific, indicating extensive sex-dependent regulatory architecture. We replicated 3, 1, 9, and 2 cis-eQTMs and 96, 84, 216, and 41 trans-eQTMs in the sex-combined, female-stratified, male-stratified, and interaction models, respectively. Sex-combined targets implicated innate immune and inflammatory signaling (TNF, IRF8); female-stratified targets implicated cytoskeletal remodeling, focal adhesion, and PI3K-Akt signaling (ITGA10); male-stratified targets implicated transcriptional regulation and post-translational modification (MOV10, HDAC7, CSF1, KAT2A); and interaction targets implicated lipid-mediator metabolism and inflammatory signaling and resolution (CYP2C9, LTB4R, CCL19). Our sex-dependent analyses showed that the majority of eQTMs resided within the trans-regulatory architecture, suggesting that sex-dependent gene expression differences are associated predominantly with trans rather than cis regulatory mechanisms. Pathway analyses revealed enrichment of innate immunity in the sex-combined model, structural remodeling in females, transcriptional control in males, and inflammatory resolution in the sex-interaction model.
InterpretationSex-specific and sex-dependent airway methylation-expression associations were largely undetected by sex-combined analysis. Sex-aware analytical models may help resolve epigenetic mechanisms underlying sex differences in asthma biology.
Research in context Evidence before this studyAsthma shows well-recognized sex differences in prevalence, severity, and clinical outcomes, yet the molecular mechanisms underlying these differences remain incompletely understood. Airway DNAm has been implicated in asthma pathogenesis and may contribute to sex-related disease heterogeneity. However, most epigenomic studies have analyzed females and males together, adjusting for sex as a covariate, an approach that may leave sex-specific and sex-dependent methylation-expression relationships undetected. Consequently, whether airway DNA methylation regulates gene expression differently in females and males, through cis- or trans-acting mechanisms, remains largely unknown.
Added value of this studyWe integrated airway epithelial DNAm and transcriptomic data to perform cis- and trans-eQTM analyses using sex-combined, sex-stratified, and CpG-by-sex interaction models. Asthma-associated CpGs showed distinct sex-specific methylation patterns, with predominantly hypomethylated CpGs in females and hypermethylated CpGs in males. Most eQTMs were model-specific, with only 379 trans-eQTMs shared across all analytical models, indicating extensive sex-dependent regulatory architecture. Replication in an independent bronchial epithelial cohort confirmed both shared and sex-specific eQTMs, with a substantially larger number of trans than cis associations. Replicated targets identified model-specific biological pathways, including innate immunity and inflammatory signaling in the sex-combined model, structural remodeling in females, transcriptional regulation in males, and lipid-mediator metabolism and inflammatory resolution in the sex-interaction model. Importantly, many of these regulatory relationships were not identified by sex-adjusted analyses.
Implications of all the available evidenceOur findings demonstrate that sex-specific and sex-dependent methylation-expression associations represent an important component of airway epigenetic regulation in asthma and are largely undetected by sex-adjusted analyses. Incorporating sex-aware analytical frameworks into epigenomic and transcriptomic studies may improve the identification of biologically relevant regulatory mechanisms, sex-specific biomarkers, and therapeutic targets. Replication of key findings in bronchial epithelium further supports their biological relevance and potential translational value for precision medicine approaches in asthma.
Graphical Abstract
O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=100 SRC="FIGDIR/small/26365928v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): [email protected]@135cda0org.highwire.dtl.DTLVardef@14ee2c5org.highwire.dtl.DTLVardef@9ff77_HPS_FORMAT_FIGEXP M_FIG C_FIG
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BibTeX
@misc{dessie2026sexstratified,
title = {{Sex-stratified and interaction multi-omics analysis of DNA methylation and gene expression in asthma}},
author = {E. Y. Dessie and Y. Gautam and B. H. Rosen and A. Mendy and M. Kang and T. B. Mersha},
howpublished = {medRxiv},
year = {2026},
month = oct,
doi = {10.64898/2026.10.19.26365928},
url = {https://www.medrxiv.org/content/10.64898/2026.10.19.26365928v1},
note = {CC BY-NC},
}
RIS
TY - UNPB
TI - Sex-stratified and interaction multi-omics analysis of DNA methylation and gene expression in asthma
AU - E. Y. Dessie
AU - Y. Gautam
AU - B. H. Rosen
AU - A. Mendy
AU - M. Kang
AU - T. B. Mersha
PY - 2026
DA - 2026/10/19
PB - medRxiv
DO - 10.64898/2026.10.19.26365928
UR - https://www.medrxiv.org/content/10.64898/2026.10.19.26365928v1
AB - BackgroundSex differences in asthma are well recognized, but sex-associated airway DNA methylation (DNAm) patterns and their relationships with gene expression remain incompletely characterized. We therefore conducted cis- and trans-expression quantitative trait methylation (eQTM) analyses using sex-stratified and CpG-by-sex interaction models. MethodsWe analyzed nasal airway epithelial DNAm and transcriptomic data from 298 participants in the CAAPA discovery cohort. Asthma-associated differentially methylated CpGs (DMCs) were identified using sex-combined and sex-stratified models. Cis- and trans-eQTM analyses were then performed in participants with asthma using sex-combined, sex-stratified, and CpG-by-sex interaction models. To prioritize asthma-associated regulatory relationships, eQTMs that were also significant in non-asthmatic controls were excluded from downstream analyses. Replication in an independent bronchial epithelial cohort was defined by concordant effect direction and P < 0.05. Replicated targets underwent pathway and network analyses. FindingsAsthma-associated DMCs were predominantly hypomethylated in females and hypermethylated in males. Among trans-eQTMs, 379 associations were shared across all four models, whereas the majority were model-specific, indicating extensive sex-dependent regulatory architecture. We replicated 3, 1, 9, and 2 cis-eQTMs and 96, 84, 216, and 41 trans-eQTMs in the sex-combined, female-stratified, male-stratified, and interaction models, respectively. Sex-combined targets implicated innate immune and inflammatory signaling (TNF, IRF8); female-stratified targets implicated cytoskeletal remodeling, focal adhesion, and PI3K-Akt signaling (ITGA10); male-stratified targets implicated transcriptional regulation and post-translational modification (MOV10, HDAC7, CSF1, KAT2A); and interaction targets implicated lipid-mediator metabolism and inflammatory signaling and resolution (CYP2C9, LTB4R, CCL19). Our sex-dependent analyses showed that the majority of eQTMs resided within the trans-regulatory architecture, suggesting that sex-dependent gene expression differences are associated predominantly with trans rather than cis regulatory mechanisms. Pathway analyses revealed enrichment of innate immunity in the sex-combined model, structural remodeling in females, transcriptional control in males, and inflammatory resolution in the sex-interaction model. InterpretationSex-specific and sex-dependent airway methylation-expression associations were largely undetected by sex-combined analysis. Sex-aware analytical models may help resolve epigenetic mechanisms underlying sex differences in asthma biology. Research in context Evidence before this studyAsthma shows well-recognized sex differences in prevalence, severity, and clinical outcomes, yet the molecular mechanisms underlying these differences remain incompletely understood. Airway DNAm has been implicated in asthma pathogenesis and may contribute to sex-related disease heterogeneity. However, most epigenomic studies have analyzed females and males together, adjusting for sex as a covariate, an approach that may leave sex-specific and sex-dependent methylation-expression relationships undetected. Consequently, whether airway DNA methylation regulates gene expression differently in females and males, through cis- or trans-acting mechanisms, remains largely unknown. Added value of this studyWe integrated airway epithelial DNAm and transcriptomic data to perform cis- and trans-eQTM analyses using sex-combined, sex-stratified, and CpG-by-sex interaction models. Asthma-associated CpGs showed distinct sex-specific methylation patterns, with predominantly hypomethylated CpGs in females and hypermethylated CpGs in males. Most eQTMs were model-specific, with only 379 trans-eQTMs shared across all analytical models, indicating extensive sex-dependent regulatory architecture. Replication in an independent bronchial epithelial cohort confirmed both shared and sex-specific eQTMs, with a substantially larger number of trans than cis associations. Replicated targets identified model-specific biological pathways, including innate immunity and inflammatory signaling in the sex-combined model, structural remodeling in females, transcriptional regulation in males, and lipid-mediator metabolism and inflammatory resolution in the sex-interaction model. Importantly, many of these regulatory relationships were not identified by sex-adjusted analyses. Implications of all the available evidenceOur findings demonstrate that sex-specific and sex-dependent methylation-expression associations represent an important component of airway epigenetic regulation in asthma and are largely undetected by sex-adjusted analyses. Incorporating sex-aware analytical frameworks into epigenomic and transcriptomic studies may improve the identification of biologically relevant regulatory mechanisms, sex-specific biomarkers, and therapeutic targets. Replication of key findings in bronchial epithelium further supports their biological relevance and potential translational value for precision medicine approaches in asthma. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=100 SRC="FIGDIR/small/26365928v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): [email protected]@135cda0org.highwire.dtl.DTLVardef@14ee2c5org.highwire.dtl.DTLVardef@9ff77_HPS_FORMAT_FIGEXP M_FIG C_FIG
ER -