Microalgal oils, depending on their degree of unsaturation, can be utilized as either nutritional supplements or fuels; thus, a feedstock with genetically designed and tunable degree of unsaturation is desirable to maximize process efficiency and product versatility. Systematic profiling of ex vivo (in yeast), in vitro, and in vivo activities of type-2 diacylglycerol acyltransferases in Nannochloropsis oceanica (NoDGAT2s or NoDGTTs), via reverse genetics, revealed that NoDGAT2A prefers saturated fatty acids (SFAs), NoDGAT2D prefers monounsaturated fatty acids (MUFAs), and NoDGAT2C exhibits the strongest activity toward polyunsaturated fatty acids (PUFAs). As NoDGAT2A, 2C, and 2D originated from the green alga, red alga, and eukaryotic host ancestral participants of secondary endosymbiosis, respectively, a mechanistic model of oleaginousness was unveiled, in which the indigenous and adopted NoDGAT2s formulated functional complementarity and specific transcript abundance ratio that underlie a rigid SFA:MUFA:PUFA hierarchy in triacylglycerol (TAG). By rationally modulating the ratio of NoDGAT2A:2C:2D transcripts, a bank of N. oceanica strains optimized for nutritional supplement or fuel production with a wide range of degree of unsaturation were created, in which proportion of SFAs, MUFAs, and PUFAs in TAG varied by 1.3-, 3.7-, and 11.2-fold, respectively. This established a novel strategy to simultaneously improve productivity and quality of oils from industrial microalgae.

Summary: The origin of phytohormones is poorly understood, and their physiological roles in microalgae remain elusive. Genome comparison of photosynthetic autotrophic eukaryotes has revealed that the biosynthetic pathways of abscisic acid (ABA) and cytokinins (CKs) emerged in unicellular algae. While ABA and CK degradation mechanisms emerged broadly in algal lineages, complete vascular plant‐type conjugation pathways emerged prior to the rise of Streptophyta. In microalgae, a complete set of proteins from the canonical ABA and CK sensing and signaling pathways is not essential, but individual components are present, suggesting stepwise recruitment of phytohormone signaling components. In the oleaginous eustigmatophyte Nannochloropsis oceanica IMET1, UHPLC‐MS/MS detected a wide array of plant hormones, despite a phytohormone profile that is very distinct from that of flowering plants. Time‐series transcriptional analysis during nitrogen depletion revealed activation of the ABA biosynthetic pathway and antagonistic transcription of CK biosynthetic genes. Correspondingly, the ABA level increases while the dominant bioactive CK forms decrease. Moreover, exogenous CKs stimulate cell‐cycle progression while exogenous ABA acts as both an algal growth repressor and a positive regulator in response to stresses. The presence of such functional flowering plant‐like phytohormone signaling systems in Nannochloropsis sp. suggests a much earlier origin of phytohormone biosynthesis and degradation than previously believed, and supports the presence in microalgae of as yet unknown conjugation and sensing/signaling systems that may be exploited for microalgal feedstock development.