Membrane-bound MinDE complex acts as a toggle switch that drives Min oscillation coupled to cytoplasmic depletion of MinD

Abstract

The Escherichia coli Min system self-organizes into a cell-pole to cell-pole oscillator on the membrane to prevent divisions at the cell poles. Reconstituting the Min system on a lipid bilayer has contributed to elucidating the oscillatory mechanism. However, previous in vitro patterns were attained with protein densities on the bilayer far in excess of those in vivo and failed to recapitulate the standing wave oscillations observed in vivo. Here we studied Min protein patterning at limiting MinD concentrations reflecting the in vivo conditions. We identified "burst" patterns-radially expanding and imploding binding zones of MinD, accompanied by a peripheral ring of MinE. Bursts share several features with the in vivo dynamics of the Min system including standing wave oscillations. Our data support a patterning mechanism whereby the MinD-to-MinE ratio on the membrane acts as a toggle switch: recruiting and stabilizing MinD on the membrane when the ratio is high and releasing MinD from the membrane when the ratio is low. Coupling this toggle switch behavior with MinD depletion from the cytoplasm drives a self-organized standing wave oscillator.cell division | subcellular organization | pattern formation | self-organization | intracellular positioning

MinE conformational dynamics regulate membrane binding, MinD interaction, and Min oscillation

“…In contrast, although I74M releases the MTS and causes constitutive membrane binding, it is in the 6β form and the sensing step is still required to flip the switch to the active form, something MinD M193L cannot provide. In vitro work examining MinE-MinD pattern formation on a lipid bilayer indicates that MinE lacking the MTS reacts as fast or faster with MinD than the WT protein (27). Such a mutant is more effective in supporting a wave pattern in vitro, but it is unable to produce bursts (radial regions of MinD surrounded by MinE), which are thought to most closely resemble the in vivo oscillation.…”

“…The latent conformation is a 6β-stranded structure that diffuses in the cytoplasm because the segments of MinE that interact with MinD (β1 strand at the dimer interface) and the membrane (N-terminal amphipathic helices also called membrane targeting sequences, MTS) are masked. In contrast, the active conformation consists of a 4β-stranded structure with the MTS bound to the membrane and the released β1 strand and part of the loop region (residues [11][12][13][14][15][16][17][18][19][20][21][22][23][24][25][26][27][28] converted to an alpha helix bound to MinD (Fig. 1).…”

Oscillation helps to get division right

“…A third protein, MinC, which binds and travels as a passenger with MinD, is a division inhibitor but is not required for dynamic patterning. In PNAS, Vecchiarelli et al (10) extend their analysis of MinCD behavior on supported lipid bilayers in vitro to provide much needed new mechanistic insight into the dynamic patterning mechanism, which helps direct the spatial positioning of division. The work reveals the nonlinear protein interactions that drive the observed Min oscillatory behavior and demonstrates roles of MinD and MinE in patterning additional to those identified in earlier studies.…”

“…Even though the basic biochemical features of MinDE have been known for some time, it would be a mistake to believe that these past studies of Min oscillatory behavior provide a comprehensive molecular understanding of the process. One of the many strong points of the paper by Vecchiarelli et al (10) is that the new biochemical insight that emerges could not have been gleaned from classic ensemble biochemistry, or from models based on simulations of reaction-diffusion patterning mechanisms. Importantly, the paper discusses the fact that mechanistically diverse models, using different biochemical assumptions, can capture the same self-organizing oscillatory behavior using reaction-diffusion patterning mechanisms in which there is a single nonlinear protein interaction term (11)(12)(13)(14).…”

“…Vecchiarelli et al (10) preincubated ATP with fluorescently labeled derivatives of MinD and MinE at close to physiological concentrations. Introduction of this mixture into a flowcell with a supported lipid bilayer and a ∼25-μm-deep aqueous channel along its length generated a gradient of MinD that decreased from inlet to outlet as limiting MinD bound the lipid bilayer (Fig.…”