Reconstitution of signaling in bacterial chemotaxis

Abstract

Strains missing several genes required for chemotaxis toward amino acids, peptides, and certain sugars were tethered and their rotational behavior was analyzed. Null strains (called gutted) were deleted for genes that code for the transducers Tsr, Tar, Tap, and Trg and for the cytoplasmic proteins CheA, CheW, CheR, CheB, CheY, and CheZ. Motor switch components were wild type, flaAII(cheC), orflaBll(cheV). Gutted cells with wild-type motors spun exclusively counterclockwise, while those with mutant motors changed their directions of rotation. CheY reduced the bias (the fraction of time that cells spun counterclockwise) in either case. CheZ offset the effect of CheY to an extent that varied with switch allele but did not change the bias when tested alone. Transducers also increased the bias in the presence of CheY but not when tested alone. However, cells containing transducers and CheY failed to respond to attractants or repellents normally detected in the periplasm. This sensitivity was restored by addition of CheA and CheW. Thus, CheY both enhances clockwise rotation and couples the transducers to the flagella. CheZ acts, at the level of the motor, as a CheY antagonist. CheA or CheW or both are required to complete the signal pathway. A model is presented that explains these results and is consistent with other data found in the literature.Sensory transduction in bacterial chemotaxis involves receipt of information about the external environment, passage of this information across the cytoplasmic membrane, generation of signals that converge on the flagellar motors, and activation of mechanisms that permit adaptation. The concentrations of certain chemicals are sensed by transmembrane receptors, also called transducers or methylaccepting chemotaxis proteins (1, 11,39; for a review, see reference 10). Four transducers are known, one sensitive to aspartate, maltose, and certain repellents (Tar), a second sensitive to serine and certain other repellents (Tsr), a third sensitive to galactose and ribose (Trg), and a fourth sensitive to dipeptides (Tap [17]). Changes in receptor occupancy, through a series of intermediate events that we hope to understand, alter the probability that the flagella spin clockwise (CW) or counterclockwise (CCW). If they spin CW, the cells move erratically with little net displacement (they tumble); if they spin CCW, the cells swim smoothly (they run) (15,16). If a run happens to carry the cell up a spatial gradient of an attractant (e.g., of aspartate), the probability of CW rotation decreases and the probability of CCW rotation increases (4, 5), extending the run. This enables the cell to move toward a more favorable environment (3).How is sensory information transferred from the receptors to the flagella? From measurements of signal propagation in filamentous cells, Segall et al. (31) showed that there is an internal signal but that its range is short, only a few micrometers. To explain this limited range, they suggested that the signal is a small protein or ligand that is inactivated ...

Chemotactic‐like response of Escherichia coli cells lacking the known chemotaxis machinery but containing overexpressed CheY

“…However, the occurrence of this phenomenon in other two-component regulatory systems taken together with the common knowledge that extremely biased flagellar rotation per se can result in a non-chemotactic phenotype raises the question whether the extreme bias of null chemotaxis mutants may be a cause of their lack of response in chemotaxis assays. Flagellar rotation can be made less counterclockwise (CCW) biased by intracellular overproduction of the response regulator CheY (Clegg and Koshland, 1984;Wolfe et al, 1987). Here, we demonstrate that null chemotaxis mutants, made less CCW biased in this manner, exhibit a chemotactic-like response to chemoeffectors.…”

“…Wolfe (personal communication) found that the gutted strain HCB437 that overexpresses CheY responds to galactose by increasing CCW rotation. (c) Wolfe et al (1987) did not find a response of tethered cells of a Y þ -gutted strain to 10 ¹4 M aspartate. This finding is in agreement with our observation that the net response to aspartate at this concentration is zero (Fig.…”

“…In such strains (unlike strains containing the plasmid pRL22⌬PvuII), the level of CheY produced without induction is relatively low ( Fig. 2; Wolfe et al, 1987). Even when induced with arabinose, the level of CheY was lower than that in noninduced, pRL22⌬Pvu II-containing strains (Fig.…”

“…receptors (Parkinson and Houts, 1982;Wolfe et al, 1987). These gutted strains, which lack the entire conventional chemotaxis system, rotate their flagella exclusively CCW (Parkinson and Houts, 1982;Wolfe et al, 1987), which is the default direction of rotation (for a review, see Eisenbach, 1996).…”

“…The plasmid produces large quantities of CheY with and without induction (Ϸ 12-fold and Ϸ fourfold higher than the wild-type level of CheY respectively; Kuo and Koshland, 1987;Wolfe et al, 1987). We worked without induction so as to avoid formation of filamentous cells (Clegg and Koshland, 1984).…”