Identification and functional analysis of class 3 lipases associated with triacylglycerol metabolism in Nannochloropsis oceanica (Eustigmatophyceae).

Microalgae accumulate triacylglycerols (TAGs) as storage lipids in response to nutrient limitation, yet the mechanisms governing TAG turnover remain incompletely understood. In the oleaginous microalga Nannochloropsis oceanica, functional studies of TAG degradation have focused on lipases containing patatin domains, whereas the roles of other lipase families remain poorly characterized. To address this shortcoming, we focused on class 3 lipases, a group distinct from patatin-domain TAG lipases, and examined their roles in TAG metabolism in N. oceanica. Based on genome sequencing information and domain-based screening, 23 putative class 3 lipases were identified and classified into two major orthogroups, including a lineage-specific group unique to Nannochloropsis. Transcriptomic data obtained under nutrient-deficient conditions were used to prioritize candidate genes for functional analysis. Targeted gene-disruption mutants were examined under phosphate-deficient conditions, which induce TAG accumulation while causing only relatively mild growth inhibition. Disruption of the No3LIP6 or No3LIP14 gene was consistently associated with increased TAG accumulation without severe growth defects, whereas other class 3 lipase mutants exhibited growth-linked or condition-dependent effects on TAG accumulation. In addition, disruption of selected class 3 lipases resulted in opposing changes in TAG-associated fatty-acid composition, suggesting differential contributions to TAG remodeling rather than uniform effects on TAG hydrolysis. Together, these results indicate that TAG turnover in N. oceanica grown under phosphate-deficient conditions is associated with lipases beyond the patatin-domain family, with a specific subset of class 3 lipases implicated in this process. This study highlights the functional relevance of non-patatin TAG lipases in microalgal TAG metabolism.