Experimental analysis of diverse actin-like proteins from various magnetotactic bacteria by functional expression in Magnetospirillum gryphiswaldense

Magnetotactic bacteria (MTB) produce magnetosomes, which are sensory organelles consisting of nanocrystals of a magnetic iron mineral enclosed by membranes. In the well-characterized Magnetospirillum species of the Alphaproteobacteria, magnetosomes align and form highly ordered chains along filaments that consist of the bacterial actin homolog MamK. The MamK protein is part of a multi-component “magnetoskeleton” that controls the concatenation, positioning, and partitioning of the magnetosome chains (MCs) which serve as cellular compass for efficient navigation in the Earth’s magnetic field. MamK is highly conserved in all MTB; however, it is unknown whether its magnetoskeletal function is preserved, especially in those MTB which exhibit distinct and more complex architectures of MCs and often contain additional putative magnetoskeletal constituents such as the actin-like protein Mad28 with as yet-unknown functions. Here, we studied the ability of magnetosome-associated actins from a wide range of diverse MTB to rescue well-characterized magnetoskeleton mutants of the model Magnetospirillum gryphiswaldense. We found that MamK orthologs from Alpha-, Delta-, Candidatus Etaproteo-, and Nitrospirota-MTB as well as a resurrected MamK LUCA version restored MC assembly to varying degrees and exhibited filamentous localization in M. gryphiswaldense and E. coli. We also identified a novel magnetosome-related protein from the magnetotactic alphaproteobacterium Magnetovibrio blakemorei that substitutes the function of the well-characterized MamJ protein as a molecular adaptor tethering magnetosomes to MamK filaments. Moreover, we demonstrate that Mad28 orthologs from Thermodesulfobacteriota and Nitrospirota are actin-like proteins that can functionally complement mamK mutants of M. gryphiswaldense and which form filamentous structures in vivo and in vitro. IMPORTANCE: To efficiently navigate within the geomagnetic field, magnetotactic bacteria (MTB) align their magnetosome organelles into chains, which are organized by the actin-like MamK protein. Although MamK is the most highly conserved magnetosome protein common to all MTB, its analysis has been confined to a small subgroup owing to the inaccessibility of most MTB. Our study takes advantage of a genetically tractable host where expression of diverse MamK orthologs together with a resurrected MamK LUCA and uncharacterized actin-like Mad28 proteins from deep-branching MTB resulted in gradual restoration of magnetosome chains in various mutants. Our results further indicate the existence of species-specific MamK interactors and shed light on the evolutionary relationships of one of the key proteins associated with bacterial magnetotaxis.