The speed of FtsZ treadmilling is tightly regulated by membrane binding

Abstract

As one of the key elements in bacterial cell division, the cytoskeletal protein FtsZ appears to be highly involved in circumferential treadmilling along the inner membrane, yielding circular vortices when transferred to flat membranes. However, it remains unclear how a membrane-targeted protein can produce these dynamics. Here, we dissect the roles of membrane binding, GTPase activity, and the unstructured c-terminal linker on the treadmilling of a chimera ftsZ protein through in vitro reconstitution of different FtsZ-YFP-mts variants on supported membranes. In summary, our results suggest substantial robustness of dynamic vortex formation, where only significant mutations, resulting in abolished membrane binding or compromised lateral interactions, are detrimental for the generation of treadmilling rings. In addition to GTPase activity, which directly affects treadmilling dynamics, we found a striking correlation of membrane binding with treadmilling speed as a result of changing the MtS on our chimera proteins. this discovery leads to the hypothesis that the in vivo existence of two alternative tether proteins for ftsZ could be a mechanism for controlling ftsZ treadmilling.

The Pneumococcal Divisome: Dynamic Control of Streptococcus pneumoniae Cell Division

“…FtsZ monomers form short protofilaments at the midcell and move circumferentially around the short axis of the cell via a rapid GTP-dependent polymerisation and depolymerisation treadmilling mechanism, similar to that of eukaryotic tubulin ( Dai and Lutkenhaus, 1991 ; Adams and Errington, 2009 ; Erickson et al, 2010 ; Meier and Goley., 2014 ; Haeusser and Margolin, 2016 ; Bisson-Filho et al, 2017 ; Yang et al, 2017 ; Perez et al, 2019 ). FtsZ treadmilling velocity appears to be controlled by the manner of membrane anchoring, and FtsZ GTPase activity ( Bisson-Filho et al, 2017 ; Yang et al, 2017 ; Perez et al, 2019 ; García-Soriano et al, 2020 ) is important for correct divisome formation ( Yang et al, 2017 ; Du and Lutkenhaus, 2019 ).…”

A conserved cell division protein directly regulates FtsZ dynamics in filamentous and unicellular actinobacteria

“…While the rate of treadmilling is set by the FtsZ GTPase activity, the overall architecture of the Z-ring is modulated by FtsZ-binding proteins that can influence its positioning, membrane interaction or stimulate FtsZ filament formation and bundling ( Caldas et al, 2019 ; García-Soriano et al, 2020 ; McQuillen and Xiao, 2020 ). The function of many of these proteins has been well characterized in the classical rod-shaped model organisms Escherichia coli and Bacillus subtilis .…”

In vitro assembly, positioning and contraction of a division ring in minimal cells

“…Next, we investigated whether FtsZ can form bundles under crowding conditions inside lipid vesicles, as previously reported 23,27,44,47 . To demonstrate this, we employed FtsZ-Venus-mts, a widely used FtsZ mutant containing the YFP variant Venus before the membrane targeting sequence (mts) domain from MinD 16,23,48,49 . This mutant is able to bind the membrane independently of the interaction with the membrane anchor FtsA, which is challenging to reconstitute, due to their aggregative nature 50 .…”

“…GTPase activity of FtsZ was measured by quantifying the inorganic phosphate with a colorimetric phosphate quanti cation assay (BIOMOL GREEN kit, ENZO life sciences, Lörrach, Germany) for 140 seconds 49 . Puri ed FtsZ-wt or FtsZ-G55-Venus-Q56 were used at 3 µM in Reaction buffer, and polymerization was triggered by 1 mM GTP.…”