The evolutionary path of chemosensory and flagellar macromolecular machines in Campylobacterota

Abstract

The evolution of macromolecular complex is a fundamental biological question, which is related to the origin of life and also guides our practice in synthetic biology. The chemosensory system is one of the complex structures that evolved very early in bacteria and displays enormous diversity and complexity in terms of composition and array structure in modern species. However, how the diversity and complexity of the chemosensory system evolved remains unclear. Here, using the Campylobacterota phylum with a robust "eco-evo" framework, we investigated the co-evolution of the chemosensory system and one of its important signaling outputs, flagellar machinery. Our analyses show that substantial flagellar gene alterations will lead to switch of its primary chemosensory class from one to another, or result in a hybrid of two classes. Unexpectedly, we discovered that the high-torque generating flagellar motor structure of Campylobacter jejuni and Helicobacter pylori likely evolved in the last common ancestor of the Campylobacterota phylum. Later lineages that experienced significant flagellar alterations lost some key components of complex scaffolding structures, thus derived simpler structures than their ancestor. Overall, this study revealed the co-evolutionary path of the chemosensory system and flagellar system, and highlights that the evolution of flagellar structural complexity requires more investigation in the Bacteria domain based on a resolved phylogenetic framework, with no assumptions on the evolutionary direction.

Fig 1. Chemosensory system in Campylobacterota.

(A) The presence of chemosensory classes in representative species of Campylobacterota that are illustrated in linearized genomes and mapped to the species tree. The characteristics of F3/F7/F8/F9/F14 classes in terms of composition and gene order are depicted in the bottom box. Abbreviations for chemosensory genes: A, cheA; V, cheV; W, cheW; B, cheB; R, cheR; C, cheC; D, cheD; X, cheX; Y, cheY; Z, cheZ; M, chemoreceptor. “.” represents a gene that is not involved in chemosensory system or of unknown function. The background colors are used to discriminate vent specialists, generalists and host-associated species of Campylobacterota. (B) The helical heptad types of chemoreceptors in Campylobacterota. The stacking histogram shows the percentage of each chemoreceptor type in respective genera, and the F classes of these genera are marked below. The cognate chemoreceptors types of F3/F7/F8/F9/F14 classes are shown in the right box. (C) Phylogenetic tree based on F3 class components (CheY, CheZ and CheX) in Campylobacterota.

Fig 2. F class switches and flagellar alterations.

(A) The distribution of flagellar genes are illustrated by brown lines in linearized genomes of representative species from Campylobacterota. The triangles below point out the location of a chemosensory gene locus on the chromosome. Nitratiruptor and Arcobacter are highlighted in red because their chemosensory class for flagellar motility are different from the rest genera. (B) Phylogenetic tree based on concatenated alignments of conserved flagellar proteins including FlhA, FlhB, FliF, FliG, FliE, FlgB, FlgC, FlgK and FlgL. The red curves highlight close evolutionary relationships of flagellar genes between two clusters, starting from the gene source with an arrow pointing to the recipient. (C) The gene order of F14 chemosensory class and flagellar locus in species of Nitratiruptor and Persephonella. The flagellar gene loci on the linearized genomes are zoomed in to show gene orders of flagellar (orange arrow) and chemosensory genes (pink arrow).

Fig 3. Flagellar genes in Arcobacter.

(A) Ternary comparison of flagellar gene distribution in genomes of Arcobacter halophilus (red), Lebetimonas sp. JS138 (navy) and Sulfurimonas gotlandica (yellow). The same flagellar genes are linked with each two of the three chromosomes. Ochre strips below represent the flagellar genes on the linearized genome of Arcobacter halophilus beneath the ternary pattern, and the gene orders of two main flagellar loci (Cluster 1 and Cluster 2) are shown at the bottom. (B) The regulatory hierarchy of flagellar genes in C. jejuni and H. pylori. The colors represent the transcriptional programing of flagellar genes starting from Class 1, then Class 2, to Class 3. The gene clusters in Fig 3A at the bottom are shaded based on the same color scheme, except two chemosensory genes cheZY in red.

Fig 4. F9 class in the Bacteria domain.

(A) Distribution of F9 class in different bacterial taxon. Numbers in parentheses of the pie chart represent the number of the species containing partial F9 class / the number of the species containing full F9 class/ the number of species investigated in the taxon. (B) Phylogenetic tree based on concatenated DosM and CheW₃. The presence of different chemosensory classes and flagellar genes in these species are depicted around the tree. The species that have partial F9 gene set are highlighted in red.

Fig 5. F class hybrids in Arcobacter.

(A) Phylogenetic tree of all CheY homologs in Campylobacterota. CheY homologs that likely interact with the flagellar motor switch proteins are shaded by light yellow. (B) Sequence alignments of all CheY homologs in Campylobacterota, represented by sequence logos for each CheY cluster based on the phylogenetic tree of Fig 5A. The blue dots mark conserved residues for almost all CheY homologs, and the red stars tag the key residues for binding FliM. (C) Phylogenetic tree based on FliM protein from different bacterial phyla. The red curves highlight close evolutionary relationships of FliM homologs between two clusters, starting from the gene source with an arrow pointing to the recipient.

Fig 6. Co-evolution of flagellar motor and chemosensory system in Campylobacterota.

(A) The presence or absence of 20 flagellar components mapped to the phylogenetic tree of Campylobacterota. Dot represents the presence of the specific gene and circle means absence. B, Basal disk; M, Medial disk; P, Proximal disk; H: H-ring. (B) The electron cryotomography pictures of flagellar motors in representative species including H. pylori [50], Wolinella succinogenes [32], C. jejuni [48], Arcobacter butzleri [32], and a proposed flagellar model for the ancestor of Campylobacterota. (C) Working model for the evolution of chemosensory system and flagella in Campylobacterota.