Distinct Chemotaxis Protein Paralogs Assemble into Chemoreceptor Signaling Arrays To Coordinate Signaling Output

Abstract

Most chemotactic motile bacteria possess multiple chemotaxis signaling systems, the functions of which are not well characterized. Chemotaxis signaling is initiated by chemoreceptors that assemble as large arrays, together with chemotaxis coupling proteins (CheW) and histidine kinase proteins (CheA), which form a baseplate with the cytoplasmic tips of receptors. These cell pole-localized arrays mediate sensing, signaling, and signal amplification during chemotaxis responses. Membrane-bound chemoreceptors with different cytoplasmic domain lengths segregate into distinct arrays. Here, we show that a bacterium, Azospirillum brasilense, which utilizes two chemotaxis signaling systems controlling distinct motility parameters, coordinates its chemotactic responses through the production of two separate membrane-bound chemoreceptor arrays by mixing paralogs within chemotaxis baseplates. The polar localization of chemoreceptors of different length classes is maintained in strains that had baseplate signaling proteins from either chemotaxis system but was lost when both systems were deleted. Chemotaxis proteins (CheA and CheW) from each of the chemotaxis signaling systems (Che1 and Che4) could physically interact with one another, and chemoreceptors from both classes present in A. brasilense could interact with Che1 and Che4 proteins. The assembly of paralogs from distinct chemotaxis pathways into baseplates provides a straightforward mechanism for coordinating signaling from distinct pathways, which we predict is not unique to this system given the propensity of chemotaxis systems for horizontal gene transfer. IMPORTANCE The assembly of chemotaxis receptors and signaling proteins into polar arrays is universal in motile chemotactic bacteria. Comparative genome analyses indicate that most motile bacteria possess multiple chemotaxis signaling systems, and experimental evidence suggests that signaling from distinct chemotaxis systems is integrated. Here, we identify one such mechanism. We show that paralogs from two chemotaxis systems assemble together into chemoreceptor arrays, forming baseplates comprised of proteins from both chemotaxis systems. These mixed arrays provide a straightforward mechanism for signal integration and coordinated response output from distinct chemotaxis systems. Given that most chemotactic bacteria encode multiple chemotaxis systems and the propensity for these systems to be laterally transferred, this mechanism may be common to ensure chemotaxis signal integration occurs.

Specific Root Exudate Compounds Sensed by Dedicated Chemoreceptors Shape Azospirillum brasilense Chemotaxis in the Rhizosphere

“…One possibility may lie in the organization of chemoreceptors in large ordered arrays that are clustered at the cell poles (48). A. brasilense possesses two spatially distinct membrane-bound chemotaxis arrays at the cell poles with chemoreceptors segregating into arrays based on heptad repeat length (36H and 38H) (49,50). The genome of A. brasilense encodes 6 PilZ domaincontaining chemoreceptors, which all belong to the 38H length class and are thus expected to be clustered together into a single array, together with other 38H chemoreceptors and spatially segregated from the 36H chemotaxis signaling array (49).…”

“…We hypothesize that in the absence of c-di-GMP, the PilZ-containing chemoreceptors adopt a conformation that abolishes signaling competence by the chemotaxis array. Given that the majority of A. brasilense chemoreceptors belong to the 38H class (49), it is likely that such a conformational change would alter chemoreceptor packing in such a way as to render cells unable to integrate and respond to most chemical cue gradients. Our previous observations that the lack of c-di-GMP binding to Tlp1 (38,42) and Aer (31) appears to inhibit their ability to signal during chemotaxis are consistent with this possibility.…”

“…Given the allosteric packing and interaction of chemoreceptors in chemotaxis signaling arrays, it is conceivable that the binding of c-di-GMP to PilZ domain chemoreceptors could influence the receptor signaling state as well as that of allosterically coupled neighboring chemoreceptors. This hypothesis predicts that cells should be able to process some cues through the 36H chemoreceptor array, which would lack PilZ domain-containing chemoreceptors, although the sensing versatility of 36H cluster-associated chemoreceptors is expected to be significantly limited given that only three 36H chemoreceptors are predicted to comprise this cluster (49).…”

Electrolocation? The evidence for redox‐mediated taxis in Shewanella oneidensis

“…Double‐deletion mutants, each lacking SO_2240 and one of the remaining four genes, however, showed greater defects than the SO_2240 deletion alone. These results may be rationalized by considering that MCPs can be bundled into heterogeneous arrays, combining inputs from different receptors (Briegel et al, 2012; O’Neal et al, 2019; Seitz et al, 2014). MCPs with the same number of seven amino acid repeats (heptads) in their cytoplasmic domain have the same cytoplasmic domain length, and can be coupled together into MCP arrays (O’Neal et al, 2019; Seitz et al, 2014).…”

Multiple CheY Proteins Control Surface-Associated Lifestyles of Azospirillum brasilense

“…The alphaproteobacterium A. brasilense are soil motile diazotrophic bacteria able to colonize the roots of diverse plants and promote their growth through phytohormones production and nitrogen fixation ( Steenhoudt and Vanderleyden, 2000 ). A. brasilense motility and chemotaxis are important for plant root colonization ( Zhulin and Armitage, 1992 ; Greer-Phillips et al, 2004 ; O’Neal et al, 2019 , 2020 ). A. brasilense cells are motile using a single polar flagellum that allows the cells to swim in liquid media and when the viscosity of the media increases, cells produce multiple lateral flagella, structurally distinct from the polar flagellum, that permit translocation across surfaces by swarming ( Moens et al, 1996 ).…”