Adenine Nucleotide-dependent Regulation of Assembly of Bacterial Tubulin-like FtsZ by a Hypermorph of Bacterial Actin-like FtsA

Tushar K Beuria,William Margolin,Mahalakshmi Sadasivam,Eugenia Mileykovskaya,Srinivas Mullapudi,William Dowhan

doi:10.1074/jbc.m808872200

Abstract

Cytokinesis in bacteria depends upon the contractile Z ring, which is composed of dynamic polymers of the tubulin homolog FtsZ as well as other membrane-associated proteins such as FtsA, a homolog of actin that is required for membrane attachment of the Z ring and its subsequent constriction. Here we show that a previously characterized hypermorphic mutant FtsA (FtsA*) partially disassembled FtsZ polymers in vitro. This effect was strictly dependent on ATP or ADP binding to FtsA* and occurred at substoichiometric levels relative to FtsZ, similar to cellular levels. Nucleotide-bound FtsA* did not affect FtsZ GTPase activity or the critical concentration for FtsZ assembly but was able to disassemble preformed FtsZ polymers, suggesting that FtsA* acts on FtsZ polymers. Microscopic examination of the inhibited FtsZ polymers revealed a transition from long, straight polymers and polymer bundles to mainly short, curved protofilaments. These results indicate that a bacterial actin, when activated by adenine nucleotides, can modify the length distribution of bacterial tubulin polymers, analogous to the effects of actin-depolymerizing factor/cofilin on F-actin.

Highlights

Bacterial cell division requires a large number of proteins that colocalize to form a putative protein machine at the cell membrane [1]
In E. coli, the FtsA:FtsZ ratio is crucial for proper cell division, with either too high or too low a ratio inhibiting septum formation [26, 27]
The R286W and E124A mutants of FtsA bypass the FtsA:FtsZ ratio rule, allowing cell division to occur at higher ratios than with WT2 FtsA

Summary

Introduction

Bacterial cell division requires a large number of proteins that colocalize to form a putative protein machine at the cell membrane [1]. We show that FtsA*, at physiological concentrations in the presence of ATP or ADP, has significant effects on the assembly of FtsZ filaments. Sedimentation Assay—Purified FtsZ (12 ␮M) was polymerized in FtsZ assembly buffer (25 mM PIPES, pH 7.4, with 10 mM MgCl2 and 100 mM potassium glutamate) plus 2 mM GTP and HT-FtsA* (0 –5 ␮M), in the absence or presence of 2 mM ATP, ADP, or AMPPNP.

Results

Conclusion