Sequencing reads of CC-503 and H5 mutant (PRJNA389185)

Title-"Multi-omics decipher the molecular mechanisms driving high-lipid production in an artificially-evolved Chlamydomonas mutant". Enhancing lipid accumulation in microalgae is critical for commercial viability. Commonly, lipid accumulation is accompanied by growth retardation. We previously isolated an artificially evolved mutant (H5) that matches its parental strain growth rate (CC-503) while accumulating significantly more lipids. Here, we report our results of the systems-level interrogations of the underlying perturbations that enable the H5 mutant to accumulate elevated lipid levels. Whole-genome sequencing revealed over 3,000 mutations in the H5 genome, including 45 in protein-coding sequences (e.g., phosphofructokinase, acyl- carrier protein, glycerol kinase). We found that six corresponding mutants from the Chlamydomonas Library Project had higher lipid accumulation than the control CC-5325. Key genes responsible for lipid accumulation, glycolysis, nutrient uptake, and cell proliferation (e.g., pyruvate carboxylase and carbonic anhydrase) were expressed significantly higher in H5 under nutrient-replete conditions. Metabolomics revealed H5 exceptional levels of malonate, crucial for lipid production and cell proliferation. Epigenomics showed significant hypomethylation in triacylglycerol (TAG) production-related genes and hypermethylation in cellular energy balance regulators. These multi-omic datasets help explain H5 phenotype, with accelerated glycolysis and the loss of several non-essential processes fueling lipid accumulation during exponential growth. The obtained findings can guide genetic engineering of microalgae for elevated lipid accumulation.