Architecture and Assembly of a Divergent Member of the ParM Family of Bacterial Actin-like Proteins

Christopher R Rivera,Justin M Kollman,Jessica K Polka,David A Agard,R Dyche Mullins

doi:10.1074/jbc.m110.203828

Christopher R Rivera, Justin M Kollman + Show 3 more

Open Access

https://doi.org/10.1074/jbc.m110.203828

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Abstract

Eubacteria and archaea contain a variety of actin-like proteins (ALPs) that form filaments with surprisingly diverse architectures, assembly dynamics, and cellular functions. Although there is much data supporting differences between ALP families, there is little data regarding conservation of structure and function within these families. We asked whether the filament architecture and biochemical properties of the best-understood prokaryotic actin, ParM from plasmid R1, are conserved in a divergent member of the ParM family from plasmid pB171. Previous work demonstrated that R1 ParM assembles into filaments that are structurally distinct from actin and the other characterized ALPs. They also display three biophysical properties thought to be essential for DNA segregation: 1) rapid spontaneous nucleation, 2) symmetrical elongation, and 3) dynamic instability. We used microscopic and biophysical techniques to compare and contrast the architecture and assembly of these related proteins. Despite being only 41% identical, R1 and pB171 ParMs polymerize into nearly identical filaments with similar assembly dynamics. Conservation of the core assembly properties argues for their importance in ParM-mediated DNA segregation and suggests that divergent DNA-segregating ALPs with different assembly properties operate via different mechanisms.

Highlights

Dynamics differ significantly from each other and from conventional actin in vitro (11, 14 –16)
A paradigm emerging from this work is that, unlike the eukaryotic actin cytoskeleton, whose architecture and function are determined by accessory factors, each bacterial actin appears adapted to a specific function, with unique properties that reduce its need for accessory factors
Given the diversity of the actin-like proteins (ALPs), we asked whether the biochemical properties we proposed to be important for the cellular function of one actin-like protein, ParM from the R1 plasmid, are conserved across the entire ParM family

Summary

To whom correspondence should be addressed

Dynamics differ significantly from each other and from conventional actin in vitro (11, 14 –16). Given the diversity of the ALPs, we asked whether the biochemical properties we proposed to be important for the cellular function of one actin-like protein, ParM from the R1 plasmid, are conserved across the entire ParM family. It forms unique filaments that bundle spontaneously and lack dynamic instability [14] These findings, especially the differences in polymer assembly dynamics, invite the intriguing conclusion that different ALP families partition plasmid DNA via distinct mechanisms. These results suggest an important question; how well conserved are the biochemical and biophysical properties of more closely related ALPs, especially as individual ALP families can be more diverse than the entire family of eukaryotic actins?. Using time-resolved light scattering, as well as electron and TIRF microscopy of single filaments, we asked whether the structure and basic biophysical properties of R1 ParM are conserved in pB171 ParM

EXPERIMENTAL PROCEDURES

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DISCUSSION