Degradation of MinD oscillator complexes by Escherichia coli ClpXP

Christopher J Labreck,Catherine E Trebino,Colby N Ferreira,Josiah J Morrison,Eric C Dibiasio,Joseph Conti,Jodi L Camberg

doi:10.1074/jbc.ra120.013866

Christopher J Labreck, Catherine E Trebino + Show 5 more

Open Access

https://doi.org/10.1074/jbc.ra120.013866

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Abstract

MinD is a cell division ATPase in Escherichia coli that oscillates from pole to pole and regulates the spatial position of the cell division machinery. Together with MinC and MinE, the Min system restricts assembly of the FtsZ-ring to midcell, oscillating between the opposite ends of the cell and preventing FtsZ-ring misassembly at the poles. Here, we show that the ATP-dependent bacterial proteasome complex ClpXP degrades MinD in reconstituted degradation reactions in vitro and in vivo through direct recognition of the MinD N-terminal region. MinD degradation is enhanced during stationary phase, suggesting that ClpXP regulates levels of MinD in cells that are not actively dividing. ClpXP is a major regulator of growth phase–dependent proteins, and these results suggest that MinD levels are also controlled during stationary phase. In vitro, MinC and MinD are known to coassemble into linear polymers; therefore, we monitored copolymers assembled in vitro after incubation with ClpXP and observed that ClpXP promotes rapid MinCD copolymer destabilization and direct MinD degradation by ClpXP. The N terminus of MinD, including residue Arg 3, which is near the ATP-binding site in sequence, is critical for degradation by ClpXP. Together, these results demonstrate that ClpXP degradation modifies conformational assemblies of MinD in vitro and depresses Min function in vivo during periods of reduced proliferation.

Highlights

The Min system of E. coli functions to spatially regulate the site of cell division by inhibiting establishment of the Z-ring near the cell poles
Deletion of either clpX or clpP from a minC deletion strain in E. coli MG1655 leads to a synthetic filamentous phenotype during exponential growth [31]
After incubation of MinD with ClpXP and ATP, we detected 45.9% less MinD after 3 h (Fig. 1A); when either ATP or ClpP was omitted from reactions, the level of MinD did not change over the course of the experiment indicating that MinD is degraded by ClpXP in an ATP-dependent manner (Fig. 1A)

Summary

Introduction

The Min system of E. coli functions to spatially regulate the site of cell division by inhibiting establishment of the Z-ring near the cell poles. MinD (6 μM) was incubated with ClpX (1.0 μM), ClpP (1.2 μM), and ATP for 3 h, and degradation was measured by monitoring the loss of full-length MinD in the reaction with time (Fig. 1A).

Results

Conclusion