Enhancement of Lipid Production under Heterotrophic Conditions by Overexpression of an Endogenous bZIP Transcription Factor in Chlorella sp. HS2.

Transcription factor engineering to regulate multiple genes has shown promise in the field of microalgae genetic engineering. Here, we report the first use of transcription factor engineering in Chlorella sp. HS2, thought to have potential for producing biofuels and bioproducts. We identified seven endogenous bZIP transcription factors in Chlorella sp. HS2 and named them HSbZIP1 through HSbZIP7. We overexpressed HSbZIP1, a C-type bZIP transcription factor, in Chlorella sp. HS2 with the goal of enhancing lipid production. Phenotype screening under heterotrophic conditions showed that all transformants exhibited increased fatty acid production. In particular, HSbZIP1 37 and 58 showed fatty acid methyl ester (FAME) yields of 859 and 1,052 mg/l, respectively, at day 10 of growth under heterotrophic conditions, and these yields were 74% and 113% higher, respectively, than that of WT. To elucidate the mechanism underlying the improved phenotypes, we identified candidate HSbZIP1-regulated genes via transcription factor binding site analysis. We then selected three genes involved in fatty acid synthesis and investigated mRNA expression levels of the genes by qRTPCR. The result revealed that the possible HSbZIP1-regulated genes involved in fatty acid synthesis were upregulated in the HSbZIP1 transformants. Taken together, our results demonstrate that HSbZIP1 can be utilized to improve lipid production in Chlorella sp. HS2 under heterotrophic conditions.