Chemical regulation of Fea1 driven transgene expression in Chlamydomonas reinhardtii

Abstract

Inducible promoters can provide regulated gene expression allowing the biosynthesis of gene products at most suitable moments of cultivation. In this study, parameters of induction and deactivation of the iron-responsive Fea1 promoter (Allen et al., 2007) were investigated in the green alga Chlamydomonas reinhardtii. Our results indicate that the construct used, ble-2A-mCherry, can be expressed successfully by the Fea1 promoter under iron-deficient conditions. The fluorescence signals of the fluorescent protein mCherry obtained via flow cytometry were detectable at different intensities in response to concentrations of iron ranging from 0μM to 20μM in media. We also demonstrate that the addition of the iron chelator deferroxamine (DFO) to iron-replete media leads to promoter activation, resulting in the increase of mCherry fluorescence. Reversibility of promoter induction is detected already within 3h after transferring the cells to iron-replete chelator-free media. In this case, the progressive decrease in mCherry fluorescence can reach, within 48h, as low as 5% of the fluorescence observed in a 40h – treatment with DFO. Cell viability after DFO treatment is not affected up to a concentration of 100μM of the chelator, which enables the establishment of a cyclic process of induction and repression for the production of recombinant proteins under the control of the Fea1 inducible promoter.