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  • Review Article
  • Published:

A comprehensive guide to pilus biogenesis in Gram-negative bacteria

Key Points

  • The pili of Gram-negative bacteria are long extracellular polymers that mediate diverse functions, such as bacterial attachment, movement and substrate transport.

  • There are five classes of pili in Gram-negative bacteria: chaperone–usher pili, type IV pili, type IV secretion pili, type V pili and curli fibres.

  • Pili are assembled by multiprotein machineries that are either located in the outer membrane (chaperone–usher pili, type V pili and curli fibres) or span both the inner membrane and outer membrane (type IV pili and type IV conjugative pili).

  • Both chaperone–usher pili and type V pili use a strand exchange mechanism for assembly, whereas the double-membrane-spanning assembly platforms that assemble type IV pili and type IV conjugative pili are powered by cytoplasmic hexameric ATPases that hydrolyse ATP.

  • Recent high-resolution structures have provided a detailed picture of the pilus filament of chaperone–usher pili, type IV pili and type IV conjugative pili, which revealed distinct properties.

  • Recent cryo-electron tomography studies have revealed the structure of fully assembled pilus systems in their native environment in the bacterial cell.

  • These pilus systems could provide targets that can be used for the development of novel antibacterial compounds.

Abstract

Pili are crucial virulence factors for many Gram-negative pathogens. These surface structures provide bacteria with a link to their external environments by enabling them to interact with, and attach to, host cells, other surfaces or each other, or by providing a conduit for secretion. Recent high-resolution structures of pilus filaments and the machineries that produce them, namely chaperone–usher pili, type IV pili, conjugative type IV secretion pili and type V pili, are beginning to explain some of the intriguing biological properties that pili exhibit, such as the ability of chaperone–usher pili and type IV pili to stretch in response to external forces. By contrast, conjugative pili provide a conduit for the exchange of genetic information, and recent high-resolution structures have revealed an integral association between the pilin subunit and a phospholipid molecule, which may facilitate DNA transport. In addition, progress in the area of cryo-electron tomography has provided a glimpse of the overall architecture of the type IV pilus machinery. In this Review, we examine recent advances in our structural understanding of various Gram-negative pilus systems and discuss their functional implications.

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Figure 1: The architecture, biogenesis and structure of chaperone–usher pili.
Figure 2: Type IV pilus (T4P) architecture, biogenesis and structure.
Figure 3: F pilus architecture, biogenesis and structure.
Figure 4: Type V pilus architecture, biogenesis and structure.

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Acknowledgements

This work was funded by the UK Medical Research Council (MRC; grant 018434) and the Wellcome Trust (grant 098302 to G.W.). The authors apologize for any omissions owing to space constraints.

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Glossary

Pili

Long non-flagellar appendages at the cell surface, also referred to as fimbriae, that are present in a wide range of Gram-negative and Gram-positive bacteria and in archaea, and are involved in bacterial attachment, motility and horizontal gene transfer.

Biofilm

A community of bacterial cells that form a dense surface-associated matrix of proteins, nucleic acids and polysaccharides that provides a strong fitness advantage, such as an enhanced tolerance to antibiotics and a reduced susceptibility to host immune responses and other physical and chemical stresses.

Type IV pili

(T4P). Widespread surface appendages and important virulence factors that are used by bacteria to enable attachment, biofilm formation and both twitching and gliding motility.

Pilins

Pilus subunits, which form a pilus in their assembled state. A pilus can contain several thousand copies of a single pilin or may be composed of more than one type of pilin. In some pili, the terms 'tip pilin' and 'anchor pilin' refer to pilins that are present at the tip or the base of the pilus structure, respectively.

Lectin domain

A versatile carbohydrate-binding domain found in many Gram-negative bacterial pili that enables bacteria to attach to host tissues during infection.

SecYEG translocon

An evolutionarily conserved membrane transporter that is located in the cytoplasmic membrane of bacteria and archaea, and the membrane of the endoplasmic reticulum in eukaryotic cells. In bacteria, this machinery transports proteins into the periplasm and can insert membrane proteins into the inner membrane.

Donor-strand complementation

(DSC). A mechanism whereby an incomplete immunoglobulin-like fold in a pilus subunit is completed and stabilized in the periplasm by a donor strand from a dedicated periplasmic chaperone (either FimC or PapD).

Usher

An outer membrane-embedded protein that catalyses the assembly of chaperone–usher pili. This protein is composed of a 24-stranded β-barrel pore domain, a periplasmic amino-terminal domain (NTD), two periplasmic carboxy-terminal domains (CTD1 and CTD2) and a plug domain.

Donor-strand exchange

(DSE). A mechanism whereby an incomplete immunoglobulin-like fold in a pilus subunit is completed and stabilized by a donor strand that is provided by the amino-terminal extension of an adjacent pilus subunit. This occurs once pilus subunits are assembled into the growing pilus by the usher.

Type II secretion systems

(T2SSs). Large macromolecular nanomachines present in various pathogenic and non-pathogenic Gram-negative bacteria that are responsible for the secretion of folded proteins (including enzymes and toxins) from the periplasm to the extracellular environment.

Secretin

Large, multimeric and gated outer membrane pore-forming proteins that are found in type IV pilus systems, type II and type III secretion systems, and in some filamentous bacteriophage extrusion systems.

Pilotin

Proteins that function to ensure correct secretin localization, assembly and outer membrane insertion.

Amidase N-terminal domains

(AMIN domains). Domains that are widely distributed among bacterial peptidoglycan hydrolases and transporters located in the periplasm, and are thought to interact with the peptidoglycan cell wall. The presence of AMIN domains in secretins is thought to help anchor the type IV pilus machinery in the cell wall.

Conjugation

A mechanism of horizontal gene transfer that involves the transfer of genetic material from a donor to a recipient bacterial cell.

Donor cell

A cell that provides a conjugative genetic element, which is often a plasmid or an integrative conjugative element, that is eventually mobilized to a recipient cell at some point in the bacterial life cycle.

Integrative conjugative elements

(ICE). A large family of chromosomally encoded mobile genetic elements that encode a functional conjugative secretion system that mediates their excision, transfer and integration into a recipient cell. Integrative conjugative elements are also known as conjugative transposons.

Recipient cell

A cell that acquires a conjugative genetic element from a donor cell, which either gets incorporated into the bacterial chromosome (for example, integrative conjugative elements) or remains in the bacterial cytoplasm (for example, a plasmid).

Type IV secretion system

(T4SS). A large macromolecular nanomachine that is found in both Gram-positive and Gram-negative bacteria, as well in some archaea. These systems have evolved to deliver DNA and protein substrates into a wide range of prokaryotic and eukaryotic target cells, which promotes the spread of antibiotic resistance and bacterial pathogenesis.

Proton motive force

(PMF). An electrochemical gradient across a bacterial cell membrane that is generated by the transfer of protons or electrons across an energy-transducing membrane by an electron transport chain.

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Hospenthal, M., Costa, T. & Waksman, G. A comprehensive guide to pilus biogenesis in Gram-negative bacteria. Nat Rev Microbiol 15, 365–379 (2017). https://doi.org/10.1038/nrmicro.2017.40

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