Leif Dehmelt

Selforganization of the cytoskeleton



Contact

Phone:+49 (231) 755 7057
Fax:+49 (231) 133 - 2299

Research concept

The “Cell Morphodynamics” group studies how cells develop their complex shape. We particularly focus on the mechanisms that underlie the organization of dynamic, fibrous structures that are collectively called the cytoskeleton. To gain insight into those mechanisms, we first identify key system components via screening technologies. We then interrogate causalities between those components by combining acute activity perturbation and live-cell imaging. On the basis of those experiments, we build mathematical models of the spatio-temporal system dynamics, which help us to generate new, testable hypotheses.




Current research

Video 1: Propagation of self-amplified and self-inhibited activity that controls the contractility of the cell’s plasma membrane. Here, the signal molecule is the small GTPase Rho, which can act as a molecular switch. Specifically, Rho regulates cell contraction by activation of a molecular motor called myosin. The video shows a time-lapse of a single, human cancer cell with 100 times increased speed. Bright and warm colors represent high activity levels of Rho.

Self-organization of cytoskeletal dynamics

The emergence of cell shape can be perceived as a self-organizing process, in which dynamic, local interactions between molecules lead to pattern formation at the scale of cells. The cytoskeleton takes an important role in this process, due to its ability to translate patterns of signal network activity into patterns of intracellular forces to shape the cell. In our studies, we find that the cytoskeleton does not only transduce patterns, but is instead a central component of pattern formation based on reciprocal interplay with its regulators.

Spatio-temporal organization of cell contraction: "A sense of touch for individual cells"

In the context of our studies on cell contractility, we uncovered a self-organizing mechanism that leads to the spontaneous emergence of local pulses and propagating waves of the cytoskeletal regulator Rho (Video 1) [1]. Our experimental analysis showed that Rho amplifies its own activity by recruiting its activator GEF-H1 and that it inhibits its activity via time-delayed activation of myosins and associated RhoGAPs. Furthermore, Rho activity oscillations were modulated by matrix elasticity, showing that extracellular mechanical cues are coupled with signal network dynamics to control cell contractility. Thus, cells use contractility pulses to locally squeeze the plasma membrane to probe the elasticity of their surroundings and they use this information to modulate their behavior. Individual cells therefore have a sense of touch that uses an active probing mechanism that is based on the local, subcellular control of signal network activity.

Relevant Literature:
[1] Graessl M, Koch J, Calderon A, Kamps D, Banerjee S, Mazel T, Schulze N, Jungkurth JK, Patwardhan R, Solouk D, Hampe N, Hoffmann B, Dehmelt L, Nalbant P (2017). An excitable Rho GTPase signaling network generates dynamic subcellular contraction patterns. J Cell Biol 21(14):5311-6.
doi: 10.1083/jcb.201706052.


Self-organization of neurite formation

Video 2: Direct observation of microtubule (green, middle panel) pushing by dynein complexes (red, bottom panel) that are immobilized near the outer cell membrane. Top panel: merged movies.

The development of neurons is an illustrative example for the role of the cytoskeleton in the emergence of complex cell shapes. To gain insight into key system components, we first analyzed the role of all known microtubule regulators during this process. In this study, we identified several regulators that co-operate to stabilize and push microtubules towards the cell periphery [2,3]. Via a self-organizing mechanism that is based on directional microtubule movements and cell geometry-based protrusion amplification, those regulators can rearrange cellular microtubule arrays and induce thin, neurite-like cell protrusions [4-6]. Our previous work suggested that the microtubule motor cytoplasmic dynein plays an important role in this process [7]. To study the role of this molecule in more detail, we used single molecule imaging to directly investigate its interaction with the cell cortex and microtubules. This study revealed direct microtubule pushing by cortical dynein complexes, and a highly dynamic turnover of those complexes (Video 2). Simulations of microtubule dynamics (Video 3) show that this rapid turnover facilitates search and capture of microtubules for effective pushing [8].

Video 3: Simulation of dynein mediated microtubule motility in cells. Left: Stochastic simulation of microtubule (yellow) movements powered by cortical dynein speckles (blue). Right: Movements of newly formed microtubules inside a living cell. 

Relevant Literature:
[2] Arens J, Duong TT, Dehmelt L (2013). A morphometric screen identifies specific roles for microtubule-regulating genes in neuronal development of P19 stem cells. PLoS One 8(11):e79796.
doi: 10.1371/journal.pone.0079796.

[3] Dehmelt L, Poplawski G, Hwang E, Halpain S (2011). NeuriteQuant: an open source toolkit for high content screens of neuronal morphogenesis. BMC Neurosci 12:100.
doi: 10.1186/1471-2202-12-100.

[4] Dehmelt L (2014). Cytoskeletal self-organization in neuromorphogenesis. Bioarchitecture 4:75 (2014)
doi: 10.4161/bioa.29070.

[5] Dehmelt L and Bastiaens PI (2011). Self-Organization in Cells. In Principles of Evolution. (eds 26 H. Meyer-Ortmanns and S. Thurner), pp. 219-238. Heidelberg: Springer.

[6] Dehmelt L, Bastiaens PI (2010). Spatial organization of intracellular communication: insights from imaging. Nat Rev Mol Cell Biol 11(6):440-52.
doi: 10.1038/nrm2903.

[7] Dehmelt L, Nalbant P, Steffen W, Halpain S (2006). A microtubule-based, dynein-dependent force induces local cell protrusions: Implications for neurite initiation. Brain Cell Biol35(1):39-56.

[8] Mazel T, Biesemann A, Krejczy M, Nowald J, Müller O, Dehmelt L (2014). Direct observation of microtubule pushing by cortical dynein in living cells. Mol Biol Cell 25(1):95-106.
doi: 10.1091/mbc.E13-07-0376.



Acute perturbation and activity measurements in living cells

<strong>Figure 1: Protein arrays inside living cells.</strong> Bait presenting artificial receptor constructs (bait-PARCs) transfer an antibody surface pattern into an ordered array of intracellular bait proteins. The interaction of a labeled prey protein with multiple bait proteins is monitored inside living cells via microscopy. Zoom Image
Figure 1: Protein arrays inside living cells. Bait presenting artificial receptor constructs (bait-PARCs) transfer an antibody surface pattern into an ordered array of intracellular bait proteins. The interaction of a labeled prey protein with multiple bait proteins is monitored inside living cells via microscopy. [less]

To uncover mechanisms, how cellular structures are organized in space and time, methods are required that enable direct monitoring and acute perturbation of regulatory activities. To reach this goal, we developed novel technologies to analyze and modulate biochemical reactions inside living cells.

Protein interaction arrays in living cells

In particular, relations between multiple protein reactions have to be measured simultaneously inside individual cells to untangle complex signal networks. However, current technologies to analyze protein reactions in cells are limited by the small number of markers that can be distinguished via microscopy. To break this barrier, we developed miniaturized protein arrays that allow simultaneous monitoring of multiple protein interactions inside individual living cells (Figure 1) [9,10]. We are currently applying this technology to study signal networks that control cell shape changes.

Relevant Literature:

[9] Gandor S, Reisewitz S, Venkatachalapathy M, Arrabito G, Reibner M, Schröder H, Ruf K, Niemeyer CM, Bastiaens PI, Dehmelt L (2013). A protein-interaction array inside a living cell. Angew Chem Int Ed Engl 52(18):4790-4.
doi: 10.1002/anie.201209127.

[10] Arrabito G, Schroeder H, Schröder K, Filips C, Marggraf U, Dopp C, Venkatachalapathy M, Dehmelt L, Bastiaens PI, Neyer A, Niemeyer CM (2014). Configurable low-cost plotter device for fabrication of multi-color sub-cellular scale microarrays. Small 10(14):2870-6.
doi: 10.4161/bioa.29070.

Video 4: Molecular activity painting of the letter “N” via ~1µm wide lines of the Rho activitor GEF-H1. Plasma membrane localization of GEF-H1 induced the new formation of dynamic, myosin-based contractile structures.

"Molecular Activity Painting": Switch-like, light-controlled perturbations inside living cells

To induce acute and prolonged perturbations of protein activities in the plasma membrane we developed methods based on chemically-induced dimerization and photochemically-induced targeting to immobilized artificial receptors to directly “paint” stable network perturbations in living cells (Video 4) [11]. To combine those perturbations with activity measurements, we developed TIRF-based methods to measure the activity of the major Rho GTPases Rac1, Cdc42 and RhoA. Using these tools, we directly investigated perturbation response relationships in the spatio-temporal processing of cell contractility signaling.

Relevant Literature:
[11] Chen X, Venkatachalapathy M, Kamps D, Weigel S, Kumar R, Orlich M, Garrecht R, Hirtz M, Niemeyer CM, Wu YW, Dehmelt L. (2017). "Molecular-Activity Painting": Switch-like, Light-Controlled Perturbations inside Living Cells. Angew Chem Int Ed Engl 21(14):5311-6.
doi: 10.1002/anie.201611432





We are always looking for talented co-workers and students who would like to join our lab. Please contact us via email if you are interested.




 
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