Invertebrate methylomes provide insight into mechanisms of environmental tolerance and reveal methodological biases

There is a growing focus on the role of DNA methylation in the ability of marine invertebrates to rapidly respond to changing environmental factors and anthropogenic impacts. However, genome-wide DNA methylation studies in non-model organisms are currently hampered by limited understanding of methodological biases. Here we compare three methods for quantifying DNA methylation at single base pair resolution -- Whole Genome Bisulfite Sequencing (WGBS), Reduced Representation Bisulfite Sequencing (RRBS), and Methyl-CpG Binding Domain Bisulfite Sequencing (MBDBS) -- using multiple individuals from two reef-building coral species with contrasting environmental sensitivity. All methods reveal substantially greater methylation in Montipora capitata (11.4%) than the more sensitive Pocillopora acuta (2.9%). The majority of CpG methylation in both species occurs in gene bodies and flanking regions. In both species, MBDBS has the greatest capacity for detecting CpGs in coding regions at our sequencing depth, however MBDBS may be limited by intra-sample methylation heterogeneity. RRBS yields robust information for specific loci albeit without enrichment of any particular genome feature and with significantly reduced genome coverage. Relative genome size strongly influences the number and location of CpGs detected by each method when sequencing depth is limited, illuminating nuances in cross-species comparisons. These findings reinforce the role and importance of DNA methylation underlying environmental sensitivity in critical marine invertebrate taxa, and provide a genomic resource for investigating the functional role of DNA methylation in environmental tolerance.