Abstract
Bacterial cell and chloroplast division are driven by a contractile “Z ring” composed of the tubulin-like cytoskeletal GTPase FtsZ. Unlike bacterial Z rings, which consist of a single FtsZ, the chloroplast Z ring in plants is composed of two FtsZ proteins, FtsZ1 and FtsZ2. Both are required for chloroplast division in vivo, but their biochemical relationship is poorly understood. We used GTPase assays, light scattering, transmission electron microscopy, and sedimentation assays to investigate the assembly behavior of purified Arabidopsis thaliana (At) FtsZ1 and AtFtsZ2 both individually and together. Both proteins exhibited GTPase activity. AtFtsZ2 assembled relatively quickly, forming protofilament bundles that were exceptionally stable, as indicated by their sustained assembly and slow disassembly. AtFtsZ1 did not form detectable protofilaments on its own. When mixed with AtFtsZ2, AtFtsZ1 reduced the extent and rate of AtFtsZ2 assembly, consistent with its previously demonstrated ability to promote protofilament subunit turnover in living cells. Mixing the two FtsZ proteins did not increase the overall GTPase activity, indicating that the effect of AtFtsZ1 on AtFtsZ2 assembly was not due to a stimulation of GTPase activity. However, the GTPase activity of AtFtsZ1 was required to reduce AtFtsZ2 assembly. Truncated forms of AtFtsZ1 and AtFtsZ2 consisting of only their conserved core regions largely recapitulated the behaviors of the full-length proteins. Our in vitro findings provide evidence that FtsZ1 counterbalances the stability of FtsZ2 filaments in the regulation of chloroplast Z-ring dynamics and suggest that restraining FtsZ2 self-assembly is a critical function of FtsZ1 in chloroplasts.
Highlights
Chloroplasts, the photosynthetic organelles in plants, arose from the endosymbiosis of a free-living cyanobacterium [1]
GTP-dependent assembly of a similar AtFtsZ2 construct has been reported [52], to date no in vitro study has characterized the equivalent soluble AtFtsZ1 construct alone or in combination with AtFtsZ2, which is important for fully understanding how they cooperate in Z-ring dynamics
Sedimentation assays confirmed that AtFtsZ2 assembly is GTP-dependent [52], as indicated by the substantial increase in the proportion of AtFtsZ2 in the pellet fraction in GTP versus GDP (Fig. 1, F and Galdieria sulphuraria (Gs))
Summary
Osteryoung1,* From the 1Department of Plant Biology, Michigan State University, East Lansing, Michigan, USA; 2Department of Cell Biology, Duke University School of Medicine, Durham, North Carolina, USA
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