A Blue Light-Responsive Strong Synthetic Promoter Based on Rational Design in Chlamydomonas reinhardtii

Abstract

Chlamydomonas reinhardtii (C. reinhardtii) is a single-cell green alga that can be easily genetically manipulated. With its favorable characteristics of rapid growth, low cost, non-toxicity, and the ability for post-translational protein modification, C. reinhardtii has emerged as an attractive option for the biosynthesis of various valuable products. To enhance the expression level of exogenous genes and overcome the silencing of foreign genes by C. reinhardtii, synthetic promoters such as the chimeric promoter AR have been constructed and evaluated. In this study, a synthetic promoter GA was constructed by hybridizing core fragments from the natural promoters of the acyl carrier protein gene (ACP2) and the glutamate dehydrogenase gene (GDH2). The GA promoter exhibited a significant increase (7 times) in expressing GUS, over the AR promoter as positive control. The GA promoter also displayed a strong responsiveness to blue light (BL), where the GUS expression was doubled compared to the white light (WL) condition. The ability of the GA promoter was further tested in the expression of another exogenous cadA gene, responsible for catalyzing the decarboxylation of lysine to produce cadaverine. The cadaverine yield driven by the GA promoter was increased by 1–2 times under WL and 2–3 times under BL as compared to the AR promoter. This study obtained, for the first time, a blue light-responsive GDH2 minimal fragment in C. reinhardtii, which delivered a doubling effect under BL when used alone or in hybrid. Together with the strong GA synthetic promoter, this study offered useful tools of synthetic biology to the algal biotechnology field.

Harnessing genetic engineering to drive economic bioproduct production in algae

“…These come from genes like the Rubisco small subunit (RBCS2), heat shock protein 70A (HSP70A), photosystem I subunit D (PSAD), glutamate dehydrogenase gene (GDH2), and the acyl carrier protein gene (ACP2). They have also created synthetic ones like the HSP70A-RBCS2 fusion (AR) and the GDH2-ACP2 fusion (GA) ( Cerutti et al, 1997 ; Schroda et al, 2002 ; Scranton et al, 2016 ; Schroda, 2019 ; Neupert et al, 2020 ; Chen et al, 2023 ; Milito et al, 2023 ). GA showed seven times higher expression than AR, responding well to blue light.…”