Best practices for methylome characterization in novel species: a case study in the microalgae Nannochloropsis

Abstract

Microalgae remain an important feedstock for the production of biofuels and bioproducts. Discovery of new species drives innovation for biotransformation, where bioengineering and other technological advances can significantly optimize performance. Production is predicated on deep knowledge of algal behavior predicted from genomic and phenotypic studies. However, prediction and manipulation of behavior, particularly for scale up, remains a challenge. Understanding the contribution of epigenetic processes to algal function provides another piece of this complex puzzle for achieving bioeconomy goals. Utilizing Nannochloropsis species as a model, we provide a methodological framework for investigating epigenetic processes, analysis of the limitations of state-of-the-art techniques, and best practices for discerning novel modifications, specifically focusing on variants of DNA methylation in new species. Further, we demonstrate specific forms of DNA methylation can be overlooked by traditional epigenetic analysis strategies. Using high-throughput, lower cost techniques, we provide several pieces of evidence demonstrating Nannochloropsis gaditana and N. salina lack the most ubiquitous forms of eukaryotic DNA methylation (5mC and 5hmC) and instead employ N6-adenine methylation (6mA), commonly found in bacteria, in their genomes. Interestingly, transcriptionally diverse physiological conditions do not elicit differential 6mA methylation in Nannochloropsis spp. Thus, the presence of 6mA may provide stability and protection of the genome. These collective discoveries illuminate not only a new, exciting avenue for improving feedstock genetic drift, stability, and culture health for bioproduction scale up but also an ideal model species to study other epigenetic processes.