The Non-Flagellar Type III Secretion System Evolved from the Bacterial Flagellum and Diversified into Host-Cell Adapted Systems

Abstract

Type 3 secretion systems (T3SSs) are essential components of two complex bacterial machineries: the flagellum, which drives cell motility, and the non-flagellar T3SS (NF-T3SS), which delivers effectors into eukaryotic cells. Yet the origin, specialization, and diversification of these machineries remained unclear. We developed computational tools to identify homologous components of the two systems and to discriminate between them. Our analysis of >1,000 genomes identified 921 T3SSs, including 222 NF-T3SSs. Phylogenomic and comparative analyses of these systems argue that the NF-T3SS arose from an exaptation of the flagellum, i.e. the recruitment of part of the flagellum structure for the evolution of the new protein delivery function. This reconstructed chronology of the exaptation process proceeded in at least two steps. An intermediate ancestral form of NF-T3SS, whose descendants still exist in Myxococcales, lacked elements that are essential for motility and included a subset of NF-T3SS features. We argue that this ancestral version was involved in protein translocation. A second major step in the evolution of NF-T3SSs occurred via recruitment of secretins to the NF-T3SS, an event that occurred at least three times from different systems. In rhizobiales, a partial homologous gene replacement of the secretin resulted in two genes of complementary function. Acquisition of a secretin was followed by the rapid adaptation of the resulting NF-T3SSs to multiple, distinct eukaryotic cell envelopes where they became key in parasitic and mutualistic associations between prokaryotes and eukaryotes. Our work elucidates major steps of the evolutionary scenario leading to extant NF-T3SSs. It demonstrates how molecular evolution can convert one complex molecular machine into a second, equally complex machine by successive deletions, innovations, and recruitment from other molecular systems.

Author Summary

Most motile bacteria use a flagellum to move. The extracellular components of flagella are secreted by their own Type 3 Secretion System (T3SS). The non-flagellar T3SS (NF-T3SS), also named injectisome, includes many proteins that are homologous to flagellar components. NF-T3SSs are employed by many plant and animal pathogens to deliver effectors to host cells, including toxins. NF-T3SSs are complex protein machineries with >15 components that connect bacterial cell envelopes to eukaryotic cell membranes, including the intervening extracellular space. In this study, we designed computational tools to distinguish flagella and NF-T3SSs from other bacterial protein sequences. We show that NF-T3SSs evolved from the flagellum by a series of genetic deletions, innovations, and recruitments of components from other cellular structures. Our evolutionary analysis suggests that NF-T3SSs then quickly adapted to different eukaryotic cells while maintaining a core structure that remains highly similar to the flagellum. This is an example of evolutionary tinkering where a complex structure arises by exaptation, the recruitment of elements that evolved initially for other functions in other cellular structures.