Only One of the Five CheY Homologs in Vibrio cholerae Directly Switches Flagellar Rotation

Abstract

Vibrio cholerae has three sets of chemotaxis (Che) proteins, including three histidine kinases (CheA) and four response regulators (CheY) that are encoded by three che gene clusters. We deleted the cheY genes individually or in combination and found that only the cheY3 deletion impaired chemotaxis, reinforcing the previous conclusion that che cluster II is involved in chemotaxis. However, this does not exclude the involvement of the other clusters in chemotaxis. In other bacteria, phospho-CheY binds directly to the flagellar motor to modulate its rotation, and CheY overexpression, even without CheA, causes extremely biased swimming behavior. We reasoned that a V. cholerae CheY homolog, if it directly controls flagellar rotation, should also induce extreme swimming behavior when overproduced. This was the case for CheY3 (che cluster II). However, no other CheY homolog, including the putative CheY (CheY0) protein encoded outside the che clusters, affected swimming, demonstrating that these CheY homologs cannot act directly on the flagellar motor. CheY4 very slightly enhanced the spreading of an Escherichia coli cheZ mutant in semisolid agar, raising the possibility that it can affect chemotaxis by removing a phosphoryl group from CheY3. We also found that V. cholerae CheY3 and E. coli CheY are only partially exchangeable. Mutagenic analyses suggested that this may come from coevolution of the interacting pair of proteins, CheY and the motor protein FliM. Taken together, it is likely that the principal roles of che clusters I and III as well as cheY0 are to control functions other than chemotaxis.

The CheYs of Rhodobacter sphaeroides

“…Why are a minimum of two CheYs required for R. sphaeroides chemotaxis when many bacteria, e.g. E. coli, Bacillus subtilis, and Vibrio cholerae, can manage with only one (45,46)? Sinorhizobium meliloti and R. sphaeroides lack the CheZ, CheC, CheX, and FliY signal terminating CheY phosphatases that have been described in other bacteria (47)(48)(49).…”

Vibrio cholerae Flagellar Synthesis and Virulence

“…The net result of these effects on flagellar rotation is net swimming towards chemoattractants and away from chemorepellants (Falke, Bass et al 1997;Armitage 1999). V. cholerae encodes three clusters of chemotaxis proteins (Heidelberg, Eisen et al 2000), but the cluster that is embedded within the flagellar gene cluster (within the Class II flhA operon: cheY3, cheZ, cheA2, cheB2, and cheW1) appears to be the major chemotaxis machinery that controls flagellar rotation under most conditions (Camilli and Mekalanos 1995;Hyakutake, Homma et al 2005). Methyl-accepting chemotaxis proteins (MCPs) in the cytoplasmic membrane interact with chemoattractant/repellants and the signal is transmitted through CheA to CheY via phosphorylation.…”

“…Phospho-CheY then interacts with the C-ring of the flagellum, which causes a reversion from CCW to CW rotation, resulting in a change of swimming direction. CheB and CheW are involved in modulating the signal transduction pathway (Freter and O'Brien 1981;Alm and Manning 1990;Everiss, Hughes et al 1994;Harkey, Everiss et al 1994;Lee, Butler et al 2001;Banerjee, Das et al 2002;Hyakutake, Homma et al 2005). …”

Integration of Global Regulatory Mechanisms Controlling Vibrio Cholerae Behavior

“…FliG, FliM and FliN, also required for torque generation, form the switch complex at the base of the flagellum basal body (Boles & McCarter, 2000;McCarter, 2001). The direction of flagellum rotation (clockwise or counterclockwise) is dictated by the interaction between the response regulator CheY3 and the FliM component of the motor (Berg, 2003;Boin et al, 2008;Hyakutake et al, 2005). The V. cholerae genome contains numerous chemotaxis-related genes, including multiple methyl-accepting chemotaxis proteins (MCP), methyltransferases (CheR), methylesterases (CheB), linker proteins (CheW), histidine kinases (CheA), and response regulators (CheY), mostly located in three clusters (Boin et al, 2008).…”