Methylome Profiling of a Deuterostome Invertebrate Using Oxford Nanopore Technology (ONT)

DNA methylation is crucial for genome regulation and provides key insights into the interaction between genetics and environmental factors, offering valuable perspectives for ecological research. However, knowledge of DNA methylation patterns in nonmodel invertebrates remains limited. The present study addresses this knowledge gap by conducting the first methylome profiling of the Pacific crown-of-thorns seastar (CoTS; Acanthaster cf. solaris), a coral-eating species that aggravates the decline of Indo-Pacific coral reefs. Using Oxford Nanopore Technology (ONT) we generated long-read sequences, covering over 90% of CpG dinucleotides in the CoTS genome. Our analysis revealed a mosaic methylation landscape with moderate genome-wide methylation levels of 37.7%. Comparative analysis highlights the intermediate methylation state observed in other deuterostome invertebrates, positioning them between the hypomethylated genomes of protostomes and the hypermethylated genomes of vertebrates. Methylation in CoTS was predominantly localised within gene bodies, especially in intronic regions, enabling modulation of gene expression and potentially supporting fitness in dynamic marine environments. Additionally, elevated methylation in repetitive elements suggests a role in genome defence. This study demonstrates the effectiveness of ONT for comprehensive methylome analysis in ecologically important nonmodel species and deepens our understanding of the epigenetic landscape in deuterostome invertebrates. We also present a detailed laboratory and bioinformatics workflow, including modified phenol-chloroform protocols that address the challenge of extracting high molecular weight DNA from marine invertebrates. Together with the methylome profiles, these resources serve as a foundation for future research, enabling investigations into DNA methylation functions, applications for CoTS outbreak management and comparative studies across diverse lineages.