3 - Epigenetic Landscape Of 5-hydroxymethylcytosine And Associations With Gene Expression In Placental Tissue.

Poster Number: 3
Publication Number: 3.109

Michael Mortillo, Rollins School of Public Health, Emory University, Atlanta, GA, United States; Karen Hermetz, Emory University School of Public Health, Atlanta, GA, United States; Amber Burt, Rollins School of Public Health at Emory University, Atlanta, GA, United States; Carmen Marsit, Emory University Rollins School of Public Health, Atlanta, GA, United States

Background: 5-hydroxymethylcystosine (5hmC) is produced through enzymatic oxidation of 5-methylcytosine (5mC). 5hmC is an intermediary in the DNA demethylation pathway, though recent evidence suggests that it also acts as a functional epigenetic modification. The landscape of 5hmC and its role in gene expression in the placenta has thus far been poorly understood.

Objective: We aim to characterize placental 5hmC distribution and investigate its role in gene expression.

Design/Methods: Using 5hmC levels at CG dinucleotide (CpG) probes along with RNA-sequencing (RNA-seq) data, we assessed 5hmC distribution and its impact on gene expression across 197 placenta samples from participants in the Rhode Island Child Health Study (RICHS). 5hmC distribution was described in a genomic and CpG island-context. To identify individual loci that may be driving expression, we implemented a genome-wide expression quantitative trait hydroxymethylation (eQTHM) analysis, which performs linear regression to measure probe-transcript associations. We regressed log2-transformed RNA-seq counts on Z-score-transformed 5hmC, controlling for infant sex, gestational age, and cell-type heterogeneity. We performed a cis and trans-eQTHM analysis, which outputs cis CpG-gene pairs for probes that lie ± 1 Mb from the transcription start site (TSS) of their target gene, with all other probes classified as trans-acting.

Results: Although 5hmC levels across the placental epigenome were generally low, we identified ~48,000 loci with consistently elevated 5hmC levels. Significant (p < 0.0001) depletion of 5hmC was observed at CpG islands and within the first exon, while enrichment was found at probes far from CpG islands and within the gene body. Our eQTHM analysis produced 376 and 716 significant cis and trans pairs, respectively (cis, p < 1e-5; trans, p < 1e-7). Both cis and trans analyses showed a greater proportion of pairs with positive correlation between 5hmC and expression (cis, 71.3%, p < 0.0001; trans, 62.4%, p < 0.0001), particularly among probes closer to the TSS of their target gene (p = 0.06). The largest proportion of cis and trans probes were found in the gene body and regions further away from CpG islands (p < 0.0001). Cis probes act locally to the enhancer start site (ESS) of their target gene, while trans probes act distally downstream.Conclusion(s): Collectively, our results suggest that 5hmC is uniquely distributed across the placenta and positively regulates gene expression. This study provides the most complete epigenetic landscape of the human placenta and will be useful for future studies of the placental epigenome.