Mapping the genetic landscape of biomineralization in Magnetospirillum magneticum AMB-1 with RB-Tnseq

Abstract

Magnetotactic bacteria (MTB) are a phylogenetically diverse group of bacteria remarkable for their ability to biomineralize magnetite (Fe3O4) or greigite (Fe3S4) in organelles called magnetosomes. The majority of genes required for magnetosome formation are encoded by a magnetosome gene island (MAI). Here, we conducted random barcoded transposon mutagenesis (RB-TnSeq) in Magnetospirillum magneticum AMB-1 to identify the global genetic requirements for magnetosome formation under different growth conditions. We generated a library of 184,710 unique strains in a wild-type background, generating ~34 mutant strains for each gene. RB-TnSeq also allowed us to determine the essential gene set of AMB-1 under standard laboratory growth conditions. To pinpoint novel genes that are important for magnetosome formation, we subjected the library to magnetic selection screens in varied growth conditions. We compared biomineralization in standard growth conditions to biomineralization in high iron and anaerobic conditions, respectively. Strains with transposon insertions in the MAI gene mamT had an exacerbated biomineralization defect under both high iron and anerobic conditions compared to standard conditions, adding to our knowledge of the role of MamT in magnetosome formation. Mutants in amb4151, a gene outside of the MAI, are more magnetic than wild-type cells under anaerobic conditions. All three of these phenotypes were validated by creating a markerless deletion strain of the gene and evaluating with TEM imaging. Overall, our results indicate that growth conditions affect which genes are required for biomineralization and that some MAI genes may have more nuanced functions than was previously understood.

Effect of oxic and anoxic conditions on intracellular storage of polyhydroxyalkanoate and polyphosphate in Magnetospirillum magneticum strain AMB-1

“…O 2 concentrations in the media were manually controlled by monitoring with PyroScience noninvasive optical oxygen sensors (Germany) and injection of pure O 2 . As AMB-1 is typically grown with a low O 2 concentration (2-10%) in the culture headspace (McCausland et al, 2021), O 2 concentrations were controlled at 5.1 ± 1.2 μM during oxic conditions (n = 50). During anoxic conditions, O 2 concentrations were below the detection limit of the oxygen sensors (~0.0052 μM) (data not shown).…”