Cryo-electron microscopy reveals two distinct type IV pili assembled by the same bacterium

Neuhaus, Alexander; Selvaraj, Manikandan; Salzer, Ralf; Langer, Julian D.; Kruse, Kerstin; Kirchner, Lennart; Sanders, Kelly; Daum, Bertram; Averhoff, Beate; Gold, Vicki A. M.

doi:10.1038/s41467-020-15650-w

Cited by 43 publications

(58 citation statements)

References 74 publications

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“…that protein is called PilE (67). In addition, on November 24, 2020 by guest http://jb.asm.org/ Downloaded from some species carry multiple copy of pilins and at least in Thermus thermophilus two major pilins (PilA4 and PilA5) are assembled into distinct filaments respectively required for natural transformation and twitching motility (68). The L. pneumophila genomes show two putative PilA homologs encoded by two consecutive genes (pilA_1, lpp1889; pilA_2, lpp1890) in a locus away from any other genes encoding Type IV pilus components.…”

Section: Major and Minor Pilins Required For Natural Transformationmentioning

confidence: 99%

Transposon Insertion Sequencing in a Clinical Isolate of Legionella pneumophila Identifies Essential Genes and Determinants of Natural Transformation

et al. 2021

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Legionella pneumophila is a Gram-negative bacterium ubiquitous in freshwater environments which, if inhaled, can cause a severe pneumonia in humans. The emergence of L. pneumophila is linked to several traits selected in the environment, the acquisition of some of which involved intra- and interkingdown horizontal gene transfer events. Transposon-insertion sequencing (TIS) is a powerful method to identify the genetic basis of selectable traits as well as to identify fitness determinants and essential genes, possible antibiotic targets. TIS has not yet been used to its full power in L. pneumophila, possibly because of difficulty to obtain a high-saturation transposon insertion library. Indeed, we found that isolates of sequence type 1 (ST1), to which belong the commonly used laboratory strains, are poorly permissive to saturating mutagenesis by conjugation-mediated transposon delivery. In contrast, we obtained high-saturation libraries in non-ST1 clinical isolates, offering the prospect of using TIS on unaltered L. pneumophila strains. Focusing on one of them, we therefore used TIS to identify essential genes in L. pneumophila. We also revealed that TIS could be used to identify genes controlling vertical transmission of mobile genetic elements. We then applied TIS to identify all the genes required for L. pneumophila to develop competence and undergo natural transformation, defining the set of major and minor Type IV pilins that are engaged in DNA uptake. This work paves the way for the functional exploration of the L. pneumophila genome by TIS and the identification of the genetic basis of other life traits of this species. IMPORTANCE Legionella pneumophila is the etiologic agent of a severe form of nosocomial and community-acquired pneumonia in humans. The environmental life traits of L. pneumophila are essential to its ability to accidentally infect humans. A comprehensive identification of their genetic basis could be obtained through the use of transposon-insertion sequencing. Yet, this powerful approach, had not been fully implemented in L. pneumophila. Here we described the successful implementation of the transposon-sequencing approach in a clinical isolate of L. pneumophila. We identify essential genes, potential drug targets, and genes required for horizontal gene transfer by natural transformation. This work represents an important step towards identifying the genetic basis of the many life traits of this environmental and pathogenic species.

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Section: Major and Minor Pilins Required For Natural Transformationmentioning

confidence: 99%

Transposon Insertion Sequencing in a Clinical Isolate of Legionella pneumophila Identifies Essential Genes and Determinants of Natural Transformation

et al. 2021

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“…Despite the importance of bacterial and archaeal T4P-like filaments, the structural details of these filaments remain sparse. However, the revolution in cryo-EM has now led to atomic structures for seven bacterial T4P 19 – 21 , three archaeal flagellar filaments 22 – 24 , one archaeal “flagellar-like” filament 25 , and one archaeal T4P 26 . Interestingly, a 4.1-Å resolution structure of the T4P from S. islandicus , a hyperthermophilic and acidophilic crenarchaeon, revealed an unprecedented level of O-glycosylation on this filament 26 .…”

Section: Introductionmentioning

confidence: 99%

The structures of two archaeal type IV pili illuminate evolutionary relationships

Wang

Baquero

et al. 2020

Nat Commun

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We have determined the cryo-electron microscopic (cryo-EM) structures of two archaeal type IV pili (T4P), from Pyrobaculum arsenaticum and Saccharolobus solfataricus, at 3.8 Å and 3.4 Å resolution, respectively. This triples the number of high resolution archaeal T4P structures, and allows us to pinpoint the evolutionary divergence of bacterial T4P, archaeal T4P and archaeal flagellar filaments. We suggest that extensive glycosylation previously observed in T4P of Sulfolobus islandicus is a response to an acidic environment, as at even higher temperatures in a neutral environment much less glycosylation is present for Pyrobaculum than for Sulfolobus and Saccharolobus pili. Consequently, the Pyrobaculum filaments do not display the remarkable stability of the Sulfolobus filaments in vitro. We identify the Saccharolobus and Pyrobaculum T4P as host receptors recognized by rudivirus SSRV1 and tristromavirus PFV2, respectively. Our results illuminate the evolutionary relationships among bacterial and archaeal T4P filaments and provide insights into archaeal virus-host interactions.

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“…The nature and subunit composition of Synechocystis thin pili have not been solved so far. However, recent structures of Thermus thermophilus wide and narrow pili have shown that T4P machinery can produce structurally distinct pili depending on the type of pilin subunit incorporated [ 18 ]. Besides PilA1, Synechocystis contains a complement of ten other known PilA variants termed minor pilins [ 11 ].…”

Section: The Type IV Pilus Machinery Conveys Twitching Motility Tomentioning

confidence: 99%

“…It has been suggested that some minor pilins may be incorporated into T4P as has been found in P. aeruginosa [ 97 ], with thin pili visible during electron microscopy potentially being composed of a different set of pilins. Neuhaus et al recently found that Thermus thermophilus produces pili with different diameters that vary in pilin composition [ 18 ]. Synechocystis thin pili may thus likewise be (partially) composed of minor pilins and play an important role in flocculation.…”

Section: The T4p Apparatus Has Structural and Secretory Roles In Cmentioning

confidence: 99%

The Role of the Cyanobacterial Type IV Pilus Machinery in Finding and Maintaining a Favourable Environment

Conradi

Mullineaux

Wilde

2020

Life

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Type IV pili (T4P) are proteinaceous filaments found on the cell surface of many prokaryotic organisms and convey twitching motility through their extension/retraction cycles, moving cells across surfaces. In cyanobacteria, twitching motility is the sole mode of motility properly characterised to date and is the means by which cells perform phototaxis, the movement towards and away from directional light sources. The wavelength and intensity of the light source determine the direction of movement and, sometimes in concert with nutrient conditions, act as signals for some cyanobacteria to form mucoid multicellular assemblages. Formation of such aggregates or flocs represents an acclimation strategy to unfavourable environmental conditions and stresses, such as harmful light conditions or predation. T4P are also involved in natural transformation by exogenous DNA, secretion processes, and in cellular adaptation and survival strategies, further cementing the role of cell surface appendages. In this way, cyanobacteria are finely tuned by external stimuli to either escape unfavourable environmental conditions via phototaxis, exchange genetic material, and to modify their surroundings to fit their needs by forming multicellular assemblies.

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Cryo-electron microscopy reveals two distinct type IV pili assembled by the same bacterium

Cited by 43 publications

References 74 publications

Transposon Insertion Sequencing in a Clinical Isolate of Legionella pneumophila Identifies Essential Genes and Determinants of Natural Transformation

Transposon Insertion Sequencing in a Clinical Isolate of Legionella pneumophila Identifies Essential Genes and Determinants of Natural Transformation

The structures of two archaeal type IV pili illuminate evolutionary relationships

The Role of the Cyanobacterial Type IV Pilus Machinery in Finding and Maintaining a Favourable Environment

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