Molecular architecture, polar targeting and biogenesis of the Legionella Dot/Icm T4SS

Ghosal, Debnath; Jeong, Kwangcheol C.; Chang, Yi‐Wei; Gyore, Jacob; Teng, Lin; Gardner, Adam; Vogel, Joseph P.; Jensen, Grant J.

doi:10.1038/s41564-019-0427-4

Cited by 87 publications

(166 citation statements)

References 48 publications

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“…The S. oneidensis MR-1, for instance, were grown under different growth conditions for the purpose of studying the nanowires formed by these cells (Subramanian et al, 2018). Legionella pneumophila tomograms were also used to study the bacterial type IV secretion system (Ghosal et al, 2017(Ghosal et al, , 2019a and the bacterial type II secretion system (preprint: Ghosal et al, 2019b). For further details, please see the Materials and Methods section of reference (Kaplan et al, 2019).…”

Section: Strains and Growth Conditionsmentioning

confidence: 99%

In situ imaging of the bacterial flagellar motor disassembly and assembly processes

et al. 2019

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The self‐assembly of cellular macromolecular machines such as the bacterial flagellar motor requires the spatio‐temporal synchronization of gene expression with proper protein localization and association of dozens of protein components. In Salmonella and Escherichia coli, a sequential, outward assembly mechanism has been proposed for the flagellar motor starting from the inner membrane, with the addition of each new component stabilizing the previous one. However, very little is known about flagellar disassembly. Here, using electron cryo‐tomography and sub‐tomogram averaging of intact Legionella pneumophila, Pseudomonas aeruginosa, and Shewanella oneidensis cells, we study flagellar motor disassembly and assembly in situ. We first show that motor disassembly results in stable outer membrane‐embedded sub‐complexes. These sub‐complexes consist of the periplasmic embellished P‐ and L‐rings, and bend the membrane inward while it remains apparently sealed. Additionally, we also observe various intermediates of the assembly process including an inner‐membrane sub‐complex consisting of the C‐ring, MS‐ring, and export apparatus. Finally, we show that the L‐ring is responsible for reshaping the outer membrane, a crucial step in the flagellar assembly process.

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Section: Strains and Growth Conditionsmentioning

confidence: 99%

In situ imaging of the bacterial flagellar motor disassembly and assembly processes

et al. 2019

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“…Visualization of the Dot/Icm machine in L. pneumophila revealed the OMCC as a triangular WIFI structure (see Fig. S1A in the supplemental material) (15)(16)(17). This structure presumably includes the OM-associated DotC, DotD, and DotH proteins and IM-associated proteins DotG and DotF.…”

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confidence: 98%

Analysis of Dot/Icm Type IVB Secretion System Subassemblies by Cryoelectron Tomography Reveals Conformational Changes Induced by DotB Binding

Park

Chetrit

et al. 2020

mBio

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Type IV secretion systems (T4SSs) are sophisticated nanomachines used by many bacterial pathogens to translocate protein and DNA substrates across a host cell membrane. Although T4SSs have important roles in promoting bacterial infections, little is known about the biogenesis of the apparatus and the mechanism of substrate transfer. Here, high-throughput cryoelectron tomography (cryo-ET) was used to visualize Legionella pneumophila T4SSs (also known as Dot/Icm secretion machines) in both the whole-cell context and at the cell pole. These data revealed the distribution patterns of individual Dot/Icm machines in the bacterial cell and identified five distinct subassembled intermediates. High-resolution in situ structures of the Dot/Icm machine derived from subtomogram averaging revealed that docking of the cytoplasmic DotB (VirB11-related) ATPase complex onto the DotO (VirB4-related) ATPase complex promotes a conformational change in the secretion system that results in the opening of a channel in the bacterial inner membrane. A model is presented for how the Dot/Icm apparatus is assembled and for how this machine may initiate the transport of cytoplasmic substrates across the inner membrane. IMPORTANCE Many bacteria use type IV secretion systems (T4SSs) to translocate proteins and nucleic acids into target cells, which promotes DNA transfer and host infection. The Dot/Icm T4SS in Legionella pneumophila is a multiprotein nanomachine that is known to translocate over 300 different protein effectors into eukaryotic host cells. Here, advanced cryoelectron tomography and subtomogram analysis were used to visualize the Dot/Icm machine assembly and distribution in a single L. pneumophila cell. Extensive classification and averaging revealed five distinct intermediates of the Dot/Icm machine at high resolution. Comparative analysis of the Dot/Icm machine and subassemblies derived from wild-type cells and several mutants provided a structural basis for understanding mechanisms that underlie the assembly and activation of the Dot/Icm machine.

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“…Joseph Vogel (Washington University, St Louis, USA) combined cryo‐EM with immunofluorescence microscopy to address the assembly process of the L. pneumophila Dot‐Icm T4SS, revealing that early‐stage assembly process begins with the targeting of Dot/Icm components to the bacterial poles. Interestingly, polar targeting is mediated by two T4 components, DotU and IcmF, which have homologues in T6SS, raising the possibility that these elements may have been recruited by a T6SS (Ghosal et al, ).…”

Section: Structure and Function Of T3 T4 And T6ssmentioning

confidence: 99%

“…Interestingly, polar targeting is mediated by two T4 components, DotU and IcmF, which have homologues in T6SS, raising the possibility that these elements may have been recruited by a T6SS (Ghosal et al, 2019).…”

Section: Structure and Function Of T3 T4 And T6ssmentioning

confidence: 99%

Bacterial injection machines: Evolutionary diverse but functionally convergent

Bleves

Galán

Llosa

2020

Cellular Microbiology

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Many human pathogens use Type III, Type IV, and Type VI secretion systems to deliver effectors into their target cells. The contribution of these secretion systems to microbial virulence was the main focus of a workshop organised by the International University of Andalusia in Spain. The meeting addressed structure–function, substrate recruitment, and translocation processes, which differ widely on the different secretion machineries, as well as the nature of the translocated effectors and their roles in subverting the host cell. An excellent panel of worldwide speakers presented the state of the art of the field, highlighting the involvement of bacterial secretion in human disease and discussing mechanistic aspects of bacterial pathogenicity, which can provide the bases for the development of novel antivirulence strategies.

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Molecular architecture, polar targeting and biogenesis of the Legionella Dot/Icm T4SS

Cited by 87 publications

References 48 publications

In situ imaging of the bacterial flagellar motor disassembly and assembly processes

In situ imaging of the bacterial flagellar motor disassembly and assembly processes

Analysis of Dot/Icm Type IVB Secretion System Subassemblies by Cryoelectron Tomography Reveals Conformational Changes Induced by DotB Binding

Bacterial injection machines: Evolutionary diverse but functionally convergent

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