Research

Cellular Actin Networks

Actin polymerization plays important roles in many biological processes, such as establishing cell polarization, accommodating protruding and retracting activities of motile cells, maintaining the physical integrity of the cell, and sensing environmental forces. All these processes are facilitated by the dynamical and mechanical properties of actin filaments, and their ability to exert or resist against forces generated in a cellular environment.

Remarkably little is known at the structural level about how the molecular players of the actin machinery cooperate inside cells to produce force-generating actin systems. We exploit cryo-electron tomography methodologies to elucidate the structural principles governing actin assembly and function within the native cellular environment.

Recent and on-going studies in our group include:

 

Architecture/force relationship in macrophage podosomes

In collaboration with the Poincloux group (IPBS), we explore the three-dimensional architecture of podosomes in human primary monocyte-derived macrophages. Our objective is to reveal the molecular organization of actin filaments and actin-related proteins within podosomes, and to formulate its relationship to force generation. From the mechanical and architectural insights we will build a physical model of force generation by actin polymerization in podosomes, which will be provide a unique basis for understanding nN-scale force generation by cellular actin networks.

In our latest study, we revealed the 3D architecture enabling these mechanosensitive adhesion structures to produce nanoscale forces (Jasnin et al. bioRxiv 2021). We showed that actin polymerization in the immediate membrane vicinity is not sufficient for podosome force generation. Quantitative analysis of podosome architecture revealed that the core is subjected to compressive forces and stores high elastic energy, which in turns gives rise to nanoscale force production. Our approach shows that cryo-ET provides direct access to the mechanical properties of cellular actin networks with nanoscale precision, paving the way for multiscale modelling of actin-mediated force production in situ.

 

Molecular-scale visualization of sarcomere contraction

In collaboration with the Schwille and Boettcher groups (MPIB), we exploited subtomogram averaging to reveal the polarity, structure and molecular organization of actin-based thin filaments enabling sarcomere contraction in neonatal rat cardiomyocytes (Burbaum et al. Nat Commun 2021). This work confirms that it is now possible to attain near subnanometer resolutions within the cellular interior, and demonstrates that our integrative approach can provide structural insights from the molecular scale up to the biological context of cell contraction.

The architecture of traveling actin waves

Actin waves are dynamic supramolecular structures involved in cell migration, cytokinesis, adhesion, and neurogenesis. Although actin waves are recognized as a widespread phenomenon, the architecture critical for their propagation is unknown. In collaboration with the Gerisch group (MPIB), we used in situ cryo-electron tomography of Dictyostelium cells to unveil the wave architecture in a native state. We revealed a preferred branch orientation toward the ventral membrane and actin tents at sites of branch clustering (Jasnin et al. Structure 2019). These tent structures differ from the classical dendritic organization observed in lamellipodia, and illustrate the importance to broaden our understanding of actin assembly in situ.

Figure 1. Architecture of actin waves using cryo-FIB milling and cryo-ET. (A) View of the back of a wave leaving behind a dense meshwork of actin filaments in the inner territory. (B) In situ structure of Arp2/3 complex-mediated branch junctions. (C-D) Clustered branch sites (C) generate membrane-directed tent-like structures (D).

Actin ring formation in response to a perforated surface

In collaboration with the Gerisch group (MPIB), we found that Dictyostelium cells migrating on a perforated film explore its holes by forming actin rings around their border and extending protrusions through the free space (Jasnin et al, Structure 2016). Actin rings were identified by correlative cryo-fluorescence and cryo-electron microscopy, and thin wedges of relevant regions were obtained by cryo-focused ion-beam milling. Retracting stages were distinguished from protruding ones by the accumulation of myosin-II. Early actin rings consists of filaments pointing upright from the membrane, entangled with a meshwork of filaments close to the membrane. Branches identified at later stages suggest that formin-based nucleation of filaments is followed by Arp2/3 complex-mediated network stabilization, which prevents buckling of the force-generating filaments. Additionally, we also performed cryo-electron tomography on D. discoideum cells undergoing membrane pearling upon destruction of the actin cortex (Heinrich et al, Biophys J 2014).

  

Actin cytoskeleton reorganization during bacterial infection

In collaboration with the Cossart lab (Pasteur Institute), we visualized the three-dimensional architecture of Listeria monocytogenes comet tails during intracellular motility and cell-to-cell spread (Jasnin et al., PNAS 2013). Quantitative analysis of their actin network architecture revealed the existence of bundles of hexagonally packed filaments with spacings of 12–13 nm. Similar configurations were observed in stress fibers and filopodia, suggesting that nanoscopic bundles are a generic feature of actin filament assemblies involved in motility. In collaboration with Alvaro Crevenna (LMU Munich), we also provided a quantitative description of filament branch orientation in Listeria comet tails in cell extracts (Jasnin and Crevenna, Biophys J 2016).

  

Recent Publications

Jasnin M., Hervy J., Balor S., Bouissou A.Proag A., Voituriez R., Maridonneau-Parini I.Baumeister W.Dmitrieff S.Poincloux R.: Elasticity of dense actin networks produces nanonewton protrusive forces. bioRxiv. (2021) https://doi.org/10.1101/2021.04.13.439622

Burbaum L., Schneider J., Scholze S., Böttcher R.T., Baumeister W., Schwille P., Plitzko J.M., Jasnin M.: Molecular-scale visualization of sarcomere contraction within native cardiomyocytes. Nat Commun., 12(1):4086 (2021). bioRxiv. (2020) https://doi.org/10.1101/2020.09.09.288977

Chakraborty S., Jasnin M, Baumeister W.: Three-dimensional organization of the cytoskeleton: a cryo-electron tomography perspective. Protein Sci., 29(6):1302-1320 (2020)

Jasnin M., Beck F., Ecke M., Fukuda Y., Martinez-Sanchez A., Baumeister W., Gerisch G.: The architecture of traveling actin waves revealed by cryo-electron tomography. Structure, 27(8):1211-23.e5 (2019)

Jasnin M., Ecke M., Baumeister W., Gerisch G.: Actin organization in cells responding to a perforated surface, revealed by live imaging and cryo-electron tomography. Structure, 24(7):1031-43 (2016)

Jasnin M., Crevenna A.H.: Quantitative analysis of filament branch orientation in Listeria actin comet tails. Biophys J., 110(4):817-26 (2016)

Heinrich D., Ecke M., Jasnin M., Engel U., Gerisch G.: Reversible membrane pearling in live cells upon destruction of the actin cortex. Biophys J., 106(5):1079-91 (2014)

Jasnin M., Asano S., Gouin E., Hegerl R., Plitzko J.M., Villa E., Cossart P., Baumeister W.: Three-dimentional architecture of actin filaments in Listeria monocytogenes comet tails. Proc Natl Acad Sci U S A, 110(51):20521-6 (2013)

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