Abstract
The bloom-forming cyanobacterium Microcystis aeruginosa is known for its global distribution and for the production of toxic compounds. In the genome of M. aeruginosa PCC 7806, we discovered that the gene coding for MaOC1, a caspase homolog protease, is followed by a toxin-antitoxin module, flanked on each side by a direct repeat. We therefore investigated their possible interaction at the protein level. Our results suggest that this module belongs to the ParE/ParD-like superfamily of type II toxin-antitoxin systems. In solution, the antitoxin is predominantly alpha-helical and dimeric. When coexpressed with its cognate toxin and isolated from Escherichia coli, it forms a complex, as revealed by light scattering and affinity purification. The active site of the toxin is restricted to the C-terminus of the molecule. Its truncation led to normal cell growth, while the wild-type form prevented bacterial growth in liquid medium. The orthocaspase MaOC1 was able to cleave the antitoxin so that it could no longer block the toxin activity. The most likely target of the protease was the C-terminus of the antitoxin with two sections of basic amino acid residues. E. coli cells in which MaOC1 was expressed simultaneously with the toxin-antitoxin pair were unable to grow. In contrast, no effect on cell growth was found when using a proteolytically inactive MaOC1 mutant. We thus present the first case of a cysteine protease that regulates the activity of a toxin-antitoxin module, since all currently known activating proteases are of the serine type.
Highlights
Microcystis aeruginosa is a cyanobacterium that occupies various ecological niches and is characterized by a large genetic diversity between strains (Humbert et al, 2013)
We show that the RelE/ParE-like toxin is toxic to E. coli cells and forms a heterotrimer with its antitoxin in a ratio of 1:2 in solution
We show that the orthocaspase MaOC1, a cysteine protease encoded adjacent to the TA pair on the M. aeruginosa genome, cleaves the antitoxin in its free form but not when in complex with the toxin, thereby regulating the availability of the toxin in the cell
Summary
Microcystis aeruginosa is a cyanobacterium that occupies various ecological niches and is characterized by a large genetic diversity between strains (Humbert et al, 2013). Among the genetic elements that enable bacteria to thrive despite environmental perturbations are Toxin-Antitoxin (TA) modules, which play a crucial role in bacterial immunity and adaptation, as. TA systems were first identified as plasmid addiction modules, exerting postsegregational killing of cells that have not received the plasmid (Tsang, 2017). These modules are common on the chromosomes of most free-living bacteria. They all consist of a toxin that inhibits cell growth and an antitoxin that counteracts the activity of the toxin. In type I and III, the antitoxin molecules are small non-coding RNAs, while in all other types the antitoxin molecules are proteins (Wang et al, 2020)
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