Unraveling Vitamin B₁₂-Responsive Gene Regulation in Algae

Abstract

Photosynthetic microalgae play a vital role in primary productivity and biogeochemical cycling in both marine and freshwater systems across the globe. However, the growth of these cosmopolitan organisms depends on the bioavailability of nutrients such as vitamins. Approximately one-half of all microalgal species requires vitamin B 12 as a growth supplement. The major determinant of algal B 12 requirements is defined by the isoform of methionine synthase possessed by an alga, such that the presence of the B 12 -independent methionine synthase (METE) enables growth without this vitamin. Moreover, the widespread but phylogenetically unrelated distribution of B 12 auxotrophy across the algal lineages suggests that the METE gene has been lost multiple times in evolution. Given that METE expression is repressed by the presence of B 12 , prolonged repression by a reliable source of the vitamin could lead to the accumulation of mutations and eventually gene loss. Here, we probe METE gene regulation by B 12 and methionine/folate cycle metabolites in both marine and freshwater microalgal species. In addition, we identify a B 12 -responsive element of Chlamydomonas reinhardtii METE using a reporter gene approach. We show that complete repression of the reporter occurs via a region spanning 2574 to 290 bp upstream of the METE start codon. A proteomics study reveals that two other genes (S-Adenosylhomocysteine hydrolase and Serine hydroxymethyltransferase2) involved in the methionine-folate cycle are also repressed by B 12 in C. reinhardtii. The strong repressible nature and high sensitivity of the B 12 -responsive element has promising biotechnological applications as a cost-effective regulatory gene expression tool.

Green genes: bioinformatics and systems-biology innovations drive algal biotechnology

“…In this context, there are several well-established systems for inducible gene expression in C. reinhardtii, most notably promoters that are regulated in response to nitrate (NIT1 or NIA1) [65] or copper (CYC6) [66]. More recently, vitamin-responsive cis elements have been identified, namely a cobalamin (vitamin B 12 )-responsive promoter [67] as well as a thiamine (vitamin B 1 )-responsive riboswitch [68], and these have been demonstrated to be useful regulatory tools. Vitamins have the advantages of being benign, cheap, and effective at low concentrations.…”

The Synthetic Biology Toolkit for Photosynthetic Microorganisms

“…Numerous genetic parts are available for the control of gene expression including diverse inducible, repressible and constitutive promoters and multiple 39untranslated region (3'UTR)/terminators (Ohresser et al, 1997;Schroda et al, 2000;Fischer and Rochaix, 2001;Ferrante et al, 2008;Helliwell et al, 2014;Sawyer et al, 2015;Baek et al, 2016;Lee et al, 2018;Watanabe et al, 2018;Beltran-Aguilar et al, 2019). The promoters are detailed in Box 2.…”

“…Most of the genetic resources cited above were standardized, validated, and are openly distributed to the community (Crozet et al, 2018). The kit In Chlamydomonas, these other gene parts used to influence expression include regulatory sequences from the PSAD gene (Fischer and Rochaix, 2001) and the chimeric promoter HSP70A-RBCS2 and the RBCS2 terminator (Schroda et al, 2000) for strong constitutive expression, as well as the NIT1 (Ohresser et al, 1997) or METE (Helliwell et al, 2014) promoters, which are repressible by ammonium and vitamin B12, respectively.…”

Enhancement of lipid production and fatty acid profiling in Chlamydomonas reinhardtii, CC1010 for biodiesel production

“…Cyanocobalamin (Vitamin B12) is the key growth promoter for living organism and about 50% of microalgal population requires vitamin B12 for its growth (Droop, 2007;Helliwell et al, 2014). In order to evaluate the effects of vitamin B12, TAP media was supplemented with different concentrations of vitamin B12 (0.001%, 0.002% and 0.003%).…”