Abstract
Signal transduction utilizing membrane-spanning receptors and cytoplasmic regulator proteins is a fundamental process for all living organisms, but quantitative studies of the behavior of signaling proteins, such as their diffusion within a cell, are limited. In this study, we show that fluctuations in the concentration of the signaling molecule, phosphorylated CheY, constitute the basis of chemotaxis signaling. To analyze the propagation of the CheY-P signal quantitatively, we measured the coordination of directional switching between flagellar motors on the same cell. We analyzed the time lags of the switching of two motors in both CCW-to-CW and CW-to-CCW switching (∆τCCW-CW and ∆τCW-CCW). In wild-type cells, both time lags increased as a function of the relative distance of two motors from the polar receptor array. The apparent diffusion coefficient estimated for ∆τ values was ~9 µm2/s. The distance-dependency of ∆τCW-CCW disappeared upon loss of polar localization of the CheY-P phosphatase, CheZ. The distance-dependency of the response time for an instantaneously applied serine attractant signal also disappeared with the loss of polar localization of CheZ. These results were modeled by calculating the diffusion of CheY and CheY-P in cells in which phosphorylation and dephosphorylation occur in different subcellular regions. We conclude that diffusion of signaling molecules and their production and destruction through spontaneous activity of the receptor array generates fluctuations in CheY-P concentration over timescales of several hundred milliseconds. Signal fluctuation coordinates rotation among flagella and regulates steady-state run-and-tumble swimming of cells to facilitate efficient responses to environmental chemical signals.
Highlights
Diffusion of signaling proteins within a cell and their interaction with the molecules responsible for the input and output of the signal play an essential role in all biological signal transduction systems [1,2,3]
The switching time lag (∆τcorrelation) of two coordinated motors depended on their relative distance from the receptor array at the cell pole
The response time of CheA short (CheAS)− cells was independent of L2 (Figure 4C, black plots). These results indicate that, in CheAS+ cells, the polar localization of CheZ delays the decrease in CheY-P concentration depending on the distance from the receptor array, whereas in CheAS− cells, the CheY-P concentration decreases at the same rate throughout the cell because CheZ is distributed uniformly throughout the cytoplasm of the cell
Summary
Diffusion of signaling proteins within a cell and their interaction with the molecules responsible for the input and output of the signal play an essential role in all biological signal transduction systems [1,2,3]. In both prokaryotes and eukaryotes, cells use transmembrane receptors to sense environmental stimuli and to generate intracellular signals in the form of messenger molecules. These intracellular messenger molecules diffuse through the cytoplasm to their targets, where they regulate cellular functions such as gene expression and locomotion. Precise knowledge of the in vivo kinetics and localization of the reactions that generate and degrade signals, and of the diffusion parameters for signaling proteins, are essential to characterize sensory input-output pathways fully
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