Dissecting the molecular architecture of integrin adhesion sites by cryo-electron tomography

ABSTRACT Chemoreceptor arrays are macromolecular complexes that form extended assemblies primarily at the poles of bacterial cells and mediate chemotaxis signal transduction, ultimately controlling cellular motility. We have used cryo-electron tomography to determine the spatial distribution and molecular architecture of signaling molecules that comprise chemoreceptor arrays in wild-type Caulobacter crescentus cells. We demonstrate that chemoreceptors are organized as trimers of receptor dimers, forming partially ordered hexagonally packed arrays of signaling complexes in the cytoplasmic membrane. This novel organization at the threshold between order and disorder suggests how chemoreceptors and associated molecules are arranged in signaling assemblies to respond dynamically in the activation and adaptation steps of bacterial chemotaxis.

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Molecular architecture of inner dynein arms in situ in Chlamydomonas reinhardtii flagella

The Journal of Cell Biology ◽

10.1083/jcb.200808050 ◽

2008 ◽

Vol 183 (5) ◽

pp. 923-932 ◽

Cited By ~ 121

Author(s):

Khanh Huy Bui ◽

Hitoshi Sakakibara ◽

Tandis Movassagh ◽

Kazuhiro Oiwa ◽

Takashi Ishikawa

Keyword(s):

Chlamydomonas Reinhardtii ◽

Electron Tomography ◽

Three Dimensional ◽

Dimensional Structure ◽

Molecular Architecture ◽

Heavy Chains ◽

Distal Side ◽

Cryo Electron Tomography ◽

Dynein Arms

The inner dynein arm regulates axonemal bending motion in eukaryotes. We used cryo-electron tomography to reconstruct the three-dimensional structure of inner dynein arms from Chlamydomonas reinhardtii. All the eight different heavy chains were identified in one 96-nm periodic repeat, as expected from previous biochemical studies. Based on mutants, we identified the positions of the AAA rings and the N-terminal tails of all the eight heavy chains. The dynein f dimer is located close to the surface of the A-microtubule, whereas the other six heavy chain rings are roughly colinear at a larger distance to form three dyads. Each dyad consists of two heavy chains and has a corresponding radial spoke or a similar feature. In each of the six heavy chains (dynein a, b, c, d, e, and g), the N-terminal tail extends from the distal side of the ring. To interact with the B-microtubule through stalks, the inner-arm dyneins must have either different handedness or, more probably, the opposite orientation of the AAA rings compared with the outer-arm dyneins.

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The molecular architecture of engulfment during Bacillus subtilis sporulation

eLife ◽

10.7554/elife.45257 ◽

2019 ◽

Vol 8 ◽

Cited By ~ 13

Author(s):

Kanika Khanna ◽

Javier Lopez-Garrido ◽

Ziyi Zhao ◽

Reika Watanabe ◽

Yuan Yuan ◽

...

Keyword(s):

Bacillus Subtilis ◽

Mother Cell ◽

Cell Biology ◽

Focused Ion Beam ◽

High Spatial Resolution ◽

Electron Tomography ◽

Ion Beam ◽

Molecular Architecture ◽

Native State ◽

Cryo Electron Tomography

The study of bacterial cell biology is limited by difficulties in visualizing cellular structures at high spatial resolution within their native milieu. Here, we visualize Bacillus subtilis sporulation using cryo-electron tomography coupled with cryo-focused ion beam milling, allowing the reconstruction of native-state cellular sections at molecular resolution. During sporulation, an asymmetrically-positioned septum generates a larger mother cell and a smaller forespore. Subsequently, the mother cell engulfs the forespore. We show that the septal peptidoglycan is not completely degraded at the onset of engulfment. Instead, the septum is uniformly and only slightly thinned as it curves towards the mother cell. Then, the mother cell membrane migrates around the forespore in tiny finger-like projections, whose formation requires the mother cell SpoIIDMP protein complex. We propose that a limited number of SpoIIDMP complexes tether to and degrade the peptidoglycan ahead of the engulfing membrane, generating an irregular membrane front.

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The molecular architecture of engulfment during Bacillussubtilis sporulation

10.1101/498220 ◽

2018 ◽

Author(s):

Kanika Khanna ◽

Javier Lopez-Garrido ◽

Ziyi Zhao ◽

Reika Watanabe ◽

Yuan Yuan ◽

...

Keyword(s):

Mother Cell ◽

Cell Biology ◽

Focused Ion Beam ◽

Electron Tomography ◽

Molecular Model ◽

Ion Beam ◽

Leading Edge ◽

Cellular Structures ◽

Molecular Architecture ◽

Cryo Electron Tomography

AbstractThe study of cell biology is limited by the difficulty in visualizing cellular structures at high spatial resolution within their native milieu. Here, we have visualized sporulation inBacillus subtilisusing cryo-electron tomography coupled with cryo-focused ion beam milling, a technique that allows the 3D reconstruction of cellular structures in near-native state at molecular resolution. During sporulation, an asymmetrically-positioned septum divides the cell into a larger mother cell and a smaller forespore. Subsequently, the mother cell phagocytoses the forespore in a process called engulfment, which entails a dramatic rearrangement of the peptidoglycan (PG) cell wall around the forespore. By imaging wild-type sporangia, engulfment mutants, and sporangia treated with PG synthesis inhibitors, we show that the initiation of engulfment does not entail the complete dissolution of the septal PG by the mother cell SpoIIDMP complex, as was previously thought. Instead, DMP is required to maintain a flexible septum that is uniformly and only slightly thinned at the onset of engulfment. Then, the mother cell membrane migrates around the forespore by forming tiny finger-like projections, the formation of which requires both SpoIIDMP and new PG synthesized ahead of the leading edge of the engulfing membrane. We propose a molecular model for engulfment membrane migration in which a limited number of SpoIIDMP complexes tether the membrane to and degrade the new PG ahead of the leading edge, thereby generating an irregular engulfing membrane front. Our data also reveal other structures that will provide a valuable resource for future mechanistic studies of endospore formation.

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