Wildfire Smoke Exposure Is Associated With Decreased Methylation Of The Pdl2 Gene

Abstract Objective To investigate the association between wildfire smoke exposure and DNA methylation of Foxp3-related genes using targeted bisulfite specific methylation sequencing assays recently developed by EpigenDx. Methods Subjects (n=16) from the San Francisco Bay area were recruited and gave blood samples during wildfire smoke exposure from the Paradise, CA wildfire 165 miles away. Subjects had their blood re-drawn 3 months later when there was no smoke exposure. An immunology/Foxp3 methylation panel was developed for 20 genes covering 150 CpG sites, targeting genes known to exhibit distinctive methylation profiles in regulatory T cells. Multiplex PCR amplified the regions of interest, using minimal sample DNA and generates inserts ready for NGS adaptor ligation. By initially amplifying specific regions of bisulfite treated DNA, library preparation is enriched with a population of molecules that have potential as indicators or affecters of gene regulation. Using our EpigenDx Ion Torrent library preparation kit, we can ligate these amplified regions to barcoded adaptors in one step. Results Paired t-tests revealed less methylation of the PDL2 gene during wildfire exposure at sites immediately upstream of the transcriptional start sites (CpG #-85: p=.002; χ̄ wildfire= 26.3 +14.0; χ̄ no wildfire= 30.8+15.9 and CpG #-86:p=.004; χ̄ wildfire = 22.2+13.4; χ̄ no wildfire= 25.7+15.6), but not in the 5′UTR region (CpG #-84; p=.53). No significant differences were found for the other genes. Conclusion Wildfire smoke is associated with decreased PDL2 gene methylation. PDL2, a programmed death ligand, downregulates T cell responses and has previously been associated with airway inflammation and cancer, but not pollution exposure.