Imaging the nanoscale organization of peptidoglycan in living Lactococcus lactis cells

André, Guillaume; Kulakauskas, Saulius; Chapot‐Chartier, Marie‐Pierre; Navet, Benjamine; Deghorain, Marie; Bernard, Elvis; Hols, Pascal; Dufrêne, Yves F.

doi:10.1038/ncomms1027

Cited by 131 publications

(124 citation statements)

References 45 publications

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“…Recent dramatic improvements in AFM have opened up new avenues for analyzing cellular structures, such as isolated membranes 26 and cell envelopes, 8,9,32 and for probing their spatial organization, interaction, and elasticity at a molecular resolution under physiologic conditions. The temporal resolution of conventional AFM, however, is not sufficient to analyze the molecular dynamics on living cell membranes.…”

Section: Discussionmentioning

confidence: 99%

“…Surface structures on various types of cells were visualized using conventional AFM. High-resolution images were achieved mainly on microbial cells, [8][9][10] while high-resolution images of animal cells remain challenging. 36 This difference may be due to the softness of animal cells.…”

Section: Discussionmentioning

confidence: 99%

“…6,7 In particular, visualization of the architecture of a living cell is a novel powerful approach to unveil the molecular organization in vivo. For example, the hexagonal S-layer of Corynebacterium glutamicum, 8 the peptidoglycan organization of Lactococcus lactis, 9 and the germinating spores of Bacillus atrophaeus 10 have been directly visualized in living microbial cells using AFM.…”

Section: Introductionmentioning

confidence: 99%

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Single-Molecule Imaging on Living Bacterial Cell Surface by High-Speed AFM

Yamashita

Taoka

Uchihashi

et al. 2012

Journal of Molecular Biology

119

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Advances in microscopy have contributed to many biologic discoveries. Electron microscopic techniques such as cryo-electron tomography are remarkable tools for imaging the interiors of bacterial cells in the near-native state, whereas optical microscopic techniques such as fluorescence imaging are useful for following the dynamics of specific single molecules in living cells. Neither technique, however, can be used to visualize the structural dynamics of a single molecule at high resolution in living cells.In the present study, we used high-speed atomic force microscopy (HS-AFM) to image the molecular dynamics of living bacterial cell surfaces. HS-AFM visualizes the dynamic molecular processes of isolated proteins at sub-molecular resolution without the need for complicated sample preparation. In the present study, magnetotactic bacterial cells were anchored in liquid medium on substrate modified by poly-L-lysine and glutaraldehyde. High-resolution HS-AFM images of live cell surfaces showed that the bacterial outer membrane was covered with a net-like structure comprising holes and the hole rims framing them. Furthermore, HS-AFM captured the dynamic movement of the surface ultrastructure, showing that the holes in the net-like structure slowly diffused in the cell surface. Nano-dissection revealed that porin trimers constitute the net-like structure. Here, we report for the first time the direct observation of dynamic molecular architectures on a live cell surface using HS-AFM.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Single-Molecule Imaging on Living Bacterial Cell Surface by High-Speed AFM

Yamashita

Taoka

Uchihashi

et al. 2012

Journal of Molecular Biology

119

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“…Recent electron cryotomography and surface AFM experiments have revealed circumferential furrows in the cell-wall surface (Beeby et al 2013;Andre et al 2010;Hayhurst et al 2008) with a spacing of roughly 50 nm. While this observation is in agreement with the model of circumferential glycan strands it also suggests a higherorder three-dimensional structure, which is not understood yet.…”

Section: Necessity For Regulationmentioning

confidence: 99%

Getting into shape: How do rod-like bacteria control their geometry?

Amir¹,

Teeffelen²

2014

Syst Synth Biol

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Rod-like bacteria maintain their cylindrical shapes with remarkable precision during growth. However, they are also capable to adapt their shapes to external forces and constraints, for example by growing into narrow or curved confinements. Despite being one of the simplest morphologies, we are still far from a full understanding of how shape is robustly regulated, and how bacteria obtain their near-perfect cylindrical shapes with excellent precision. However, recent experimental and theoretical findings suggest that cell-wall geometry and mechanical stress play important roles in regulating cell shape in rod-like bacteria. We review our current understanding of the cell wall architecture and the growth dynamics, and discuss possible candidates for regulatory cues of shape regulation in the absence or presence of external constraints. Finally, we suggest further future experimental and theoretical directions which may help to shed light on this fundamental problem.

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“…Understanding these functions requires elucidation of the molecular architecture of microbial cell walls [21]. The cell wall of Trichophyton contains b-glucan, b-1,2-mannosides, chitin, phospholipomannan, glucuronoxylomannan, mannan, and galactomannan [21][22][23].…”

Section: Discussionmentioning

confidence: 99%

Trichophyton rubrum manipulates the innate immune functions of human keratinocytes

et al. 2011

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Evasion or subversion of host immune responses have been shown for a variety of microorganisms, and this might be the case for Trichophyton rubrum, the most common pathogenic fungus causing chronic dermatophytosis in humans. Keratinocytes, the main epidermal cells, have important roles as a first defense against microbial challenges in local immune reactions. Epidermal keratinocytes express several Toll-like receptors and produce host defense peptides, cytokines and chemokines in response to various stimuli. We analyzed the expression of Toll-Like receptor TLR2, TLR4, TLR6, and Human Beta Defensin (HBD)-1, HBD-2, Interleukin IL-1b and IL-8 production, when exposing primary keratinocyte cultures to T. rubrum. We observed changes in size and granularity of keratinocytes stimulated with either whole conidia or conidial homogenates compared to other treatments. Intact conidia decreased keratinocytes’ TLR2 and TLR6 expression without affecting that of TLR4, while conidial homogenates increased the expression of these three receptors. Interestingly, whole conidia decreased HBD-1 and HBD-2 production, whereas conidial homogenate increased it. No changes were observed in IL-1b and IL-8 production after stimulation with conidia or conidial homogenate. CONCLUSIONS. Our results suggest that: 1) Keratinocytes can recognize and respond to cell wall components of T. rubrum; 2) Viable intact conidia inhibit TLR-2 and TLR6 expression and decrease HBD-1 and HBD-2 production; 3) Conidial homogenate from T. rubrum increases the expression of TLR2, TLR4 and TLR6 and induces HBD-1 and HBD-2 production; 4) Therefore, innate immune functions of keratinocytes as the first level of local skin immunity are apparently manipulated by T. rubrum, likely to ensure its establishment, persistence and survival.

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Imaging the nanoscale organization of peptidoglycan in living Lactococcus lactis cells

Cited by 131 publications

References 45 publications

Single-Molecule Imaging on Living Bacterial Cell Surface by High-Speed AFM

Single-Molecule Imaging on Living Bacterial Cell Surface by High-Speed AFM

Getting into shape: How do rod-like bacteria control their geometry?

Trichophyton rubrum manipulates the innate immune functions of human keratinocytes

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