New Insights into Bacterial Chemoreceptor Array Structure and Assembly from Electron Cryotomography

Abstract

Bacterial chemoreceptors cluster in highly ordered, cooperative, extended arrays with a conserved architecture, but the principles that govern array assembly remain unclear. Here we show images of cellular arrays as well as selected chemoreceptor complexes reconstituted in vitro that reveal new principles of array structure and assembly. First, in every case, receptors clustered in a trimers-of-dimers configuration, suggesting this is a highly favored fundamental building block. Second, these trimers-of-receptor dimers exhibited great versatility in the kinds of contacts they formed with each other and with other components of the signaling pathway, although only one architectural type occurred in native arrays. Third, the membrane, while it likely accelerates the formation of arrays, was neither necessary nor sufficient for lattice formation. Molecular crowding substituted for the stabilizing effect of the membrane and allowed cytoplasmic receptor fragments to form sandwiched lattices that strongly resemble the cytoplasmic chemoreceptor arrays found in some bacterial species. Finally, the effective determinant of array structure seemed to be CheA and CheW, which formed a “superlattice” of alternating CheA-filled and CheA-empty rings that linked receptor trimers-of-dimer units into their native hexagonal lattice. While concomitant overexpression of receptors, CheA, and CheW yielded arrays with native spacing, the CheA occupancy was lower and less ordered, suggesting that temporal and spatial coordination of gene expression driven by a single transcription factor may be vital for full order, or that array overgrowth may trigger a disassembly process. The results described here provide new insights into the assembly intermediates and assembly mechanism of this massive macromolecular complex.

A new view into prokaryotic cell biology from electron cryotomography

“…3c). Other ECT studies have provided insights into the mechanism of activation of the system (in which substrate binding at the proximal end of the chemoreceptor triggers a conformational change in the kinase bound at its distal tip) 99 and into the assembly of arrays from subunits of trimers-of-receptor-dimers 100 .…”

Signaling Consequences of Structural Lesions that Alter the Stability of Chemoreceptor Trimers of Dimers

“…Receptor trimers of dimers are obligate assembly intermediates for receptor/CheW/CheA ternary complexes and core signaling units [9, 11, 12, 19, 23]. Accordingly, we evaluated E402* and R404* receptors expressed in receptor-less strain UU2610 (R−B−) for ternary complex formation by fluorescence microscopy, using YFP-CheZ as a clustering reporter.…”

Treponema pallidum, the syphilis spirochete: making a living as a stealth pathogen

“…Near the cell poles are chemoreceptor arrays containing the T. pallidum ’s four methyl-accepting chemotaxis proteins (Mcp1 (TP0040), Mcp2 (TP0488), Mcp3 (TP0639) and Mcp4 (TP0640)) (Fig. 1G) 91 , sensors that bind exogenously derived ligands within the periplasm and relay chemotactic signals to the more distal flagellar motors 92 . A cone-like lattice at the cell tips, connected by fine fibrils to the outer membrane, could function as an organelle mediating end-on attachment (Fig.…”