Repurposing of a DNA segregation machinery into a cytoskeletal system controlling cell shape.

Bacteria, like eukaryotes, use conserved cytoskeletal systems for intracellular organization. The plasmid-encoded ParMRC system forms actin-like filaments that segregate low–copy number plasmids. In multicellular cyanobacteria such as Anabaena sp., we found that a chromosomally encoded ParMR system has evolved into a cytoskeletal system named CorMR with a function in cell shape control rather than DNA segregation. Live-cell imaging, in vitro reconstitution, and cryo–electron microscopy revealed that CorM formed dynamically unstable, antiparallel double-stranded filaments that were recruited to the membrane by CorR through an amphipathic helix conserved in multicellular cyanobacteria. CorMR filaments were regulated by MinC, which excluded them from the poles and division plane. Comparative genomics indicated that the repurposing of ParMR and Min systems coevolved with cyanobacterial multicellularity, highlighting the evolutionary plasticity of cytoskeletal systems in bacteria. Editor's summary: How do complex cellular structures evolve? Springstein et al. found that DNA segregation machinery has been re-engineered into a cytoskeletal system controlling cell shape in multicellular cyanobacteria. This system forms dynamic filaments on the cell membrane, and its organization is guided by the Min system, a well-known regulator of cell division in bacteria. The pairing of these two ancient modules reveals how existing protein machineries can be combined and repurposed to create new cellular functions. These findings highlight the flexibility of bacterial cell biology and provide clues about the origin of multicellularity. —Stella M. Hurtley INTRODUCTION: Cytoskeletal systems provide internal protein scaffolds that give cells their shape and organize their contents. In bacteria, one such system, the plasmid-encoded ParMR module, forms actin-like filaments that push low–copy number plasmids to opposite sides of a dividing cell, ensuring that each daughter cell inherits equal DNA. Multicellular cyanobacteria such as the model cyanobacterium Anabaena sp. PCC 7120 contain an unusual, chromosome-encoded ParMR system with a function that has remained unclear, raising the possibility that this protein machinery has been evolutionarily repurposed for new cellular functions. RATIONALE: Anabaena cells carry several copies of their chromosome, and no dedicated segregation system has been described. Because ParMR systems typically partition DNA, we initially investigated whether the chromosomal ParMR in Anabaena helps to distribute chromosomes during cell division. When we deleted the corresponding genes, however, DNA distribution looked normal, but cells became enlarged and more spherical, similar to mutants lacking the shape-determining actin-like protein MreB. This unexpected phenotype prompted us to test whether the chromosomal ParMR had evolved into a cytoskeletal system that controls cell shape instead of DNA segregation. RESULTS: Using comparative genomics, we found that the chromosomal ParMR system is restricted to multicellular cyanobacteria. Live-cell high-resolution fluorescence microscopy and cryo–electron tomography showed that these filaments form cortical arrays just under the inner membrane oriented mainly perpendicular to the long cell axis. These unexpected findings prompted us to rename the chromosomal ParMR system of Anabaena CorMR (for cortical ParMR) because it functions as a new cytoskeletal system involved in cell shape maintenance instead of segregating plasmid DNA. Biochemical experiments and in vitro reconstitution on artificial membranes demonstrated that CorR can recruit CorM to lipid bilayers, where CorM filaments grow and then suddenly collapse, a behavior known as dynamic instability. Cryo–electron microscopy revealed that CorM filaments have a previously undescribed architecture for this protein family: two antiparallel strands forming a bipolar helical filament. To identify regulators of this system, we combined structural prediction, interaction assays, and in vitro biochemistry, and discovered that the well-known cell division inhibitor MinC directly binds CorM. In multicellular cyanobacteria, MinC has gained an extended N-terminal region that occupies the same binding pocket on CorM as CorR. MinC efficiently disassembles CorM filaments in vitro, and its oscillation in living cells efficiently excludes CorM polymerization from the poles and division sites. CONCLUSION: Our study uncovers an excellent example of an evolutionary repurposing of the ancient plasmid DNA-partitioning ParMR system into the membrane-associated CorMR cytoskeleton controlling the cell shape of multicellular cyanobacteria. The emergence of the CorMR system involved three linked innovations: the evolution of a bipolar CorM filament while retaining dynamic instability, the acquisition of membrane-binding by CorR, and use of the oscillating Min system as a spatial regulator that controls CorM localization. Our findings highlight the plasticity of bacterial cytoskeletal systems and illustrate how existing biochemical modules can be coopted to support new functions during the evolution of multicellularity. The repurposing of a preexisting polymeric system illustrates how cytoskeletal innovation can contribute to the emergence of cellular complexity and multicellularity in bacteria.: (Top) Evolutionary diversification and functional transformation of the ParMR plasmid segregation system into the cyanobacterial CorMR cytoskeleton. (Bottom left) CorMR forms dynamically unstable, cortical filaments that control cell shape. (Bottom right) MinC oscillations exclude CorMR filaments from cell-to-cell contacts and the division plane. [ABSTRACT FROM AUTHOR]