Enhancing carbohydrate repartitioning into lipid and carotenoid by disruption of microalgae starch debranching enzyme

Yuichi Kato,Akihiko Kondo,Jo-Shu Chang,Mami Matsuda,Tomoki Oyama,Kentaro Inokuma,Katsuya Satoh,Ryota Hidese,Tomohisa Hasunuma,Yutaka Oono,Christopher J Vavricka

doi:10.1038/s42003-021-01976-8

Abstract

Light/dark cycling is an inherent condition of outdoor microalgae cultivation, but is often unfavorable for lipid accumulation. This study aims to identify promising targets for metabolic engineering of improved lipid accumulation under outdoor conditions. Consequently, the lipid-rich mutant Chlamydomonas sp. KOR1 was developed through light/dark-conditioned screening. During dark periods with depressed CO2 fixation, KOR1 shows rapid carbohydrate degradation together with increased lipid and carotenoid contents. KOR1 was subsequently characterized with extensive mutation of the ISA1 gene encoding a starch debranching enzyme (DBE). Dynamic time-course profiling and metabolomics reveal dramatic changes in KOR1 metabolism throughout light/dark cycles. During light periods, increased flux from CO2 through glycolytic intermediates is directly observed to accompany enhanced formation of small starch-like particles, which are then efficiently repartitioned in the next dark cycle. This study demonstrates that disruption of DBE can improve biofuel production under light/dark conditions, through accelerated carbohydrate repartitioning into lipid and carotenoid.

Highlights

Light/dark cycling is an inherent condition of outdoor microalgae cultivation, but is often unfavorable for lipid accumulation
To identify a promising metabolic engineering target that can be modified to improve lipid content, lipid-rich mutants were first screened under light/ dark conditions
JSC4 was selected as the parental strain for mutagenesis and light/dark-conditioned screening because lipid accumulation significantly decreases under the conditions in this microalga[10]

Summary

Introduction

Light/dark cycling is an inherent condition of outdoor microalgae cultivation, but is often unfavorable for lipid accumulation. In the model green microalga Chlamydomonas reinhardtii, genome-wide gene expression of central carbon metabolism is regulated under diurnal periodicity, with starch content increasing during light periods and peaking in the middle of dark periods[6,7]. Other cell compositions such as lipid content are reported to be influenced by light/dark cycling. Lipid-rich mutants in diverse microalgal species, including C. reinhardtii, Chlorococcum littorale, and Euglena gracilis, were previously obtained by the selective breeding approach using FACS14,15,17–19 These screenings have only been conducted under the optimal conditions for lipid accumulation (for example, continuous illumination), and did not simulate unfavorable environmental factors that will be faced in outdoor cultivation (for example, light/dark cycling). These beneficial features of JSC4 are related to starch-to-lipid biosynthesis switching mechanisms[22], lipid content of JSC4 significantly decreases under light/dark conditions, compared to that of continuous illumination[10]

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