Curated Optogenetic Publication Database

Abstract: Cell contraction plays an important role in many physiological and pathophysiological processes. This includes functions in skeletal, heart, and smooth muscle cells, which lead to highly coordinated contractions of multicellular assemblies, and functions in non-muscle cells, which are often highly localized in subcellular regions and transient in time. While the regulatory processes that control cell contraction in muscle cells are well understood, much less is known about cell contraction in non-muscle cells. In this review, we focus on the mechanisms that control cell contraction in space and time in non-muscle cells, and how they can be investigated by light-based methods. The review particularly focusses on signal networks and cytoskeletal components that together control subcellular contraction patterns to perform functions on the level of cells and tissues, such as directional migration and multicellular rearrangements during development. Key features of light-based methods that enable highly local and fast perturbations are highlighted, and how experimental strategies can capitalize on these features to uncover causal relationships in the complex signal networks that control cell contraction.

Abstract: Rho GTPase crosstalk is thought to play a key role in the spatio-temporal coordination of cytoskeletal dynamics during cell migration. Here, we directly investigated crosstalk between the major Rho GTPases Rho, Rac and Cdc42 by combining acute activity perturbation with activity measurements in individual, mammalian cells. As expected for their proposed mutual inhibition, we confirmed that Rho inhibits Rac activity. However, surprisingly, we found that Rac strongly stimulates Rho activity. We hypothesized that this crosstalk might play a role in mediating the tight spatio-temporal coupling between cell protrusions and retractions that are typically observed during mesenchymal cell migration. Using new, improved activity sensors for endogenous Rho GTPases, we find that Rac activation is tightly and precisely coupled to local cell protrusions, followed by Rho activation during retraction. In a screen for potential crosstalk mediators, we find that a subset of the Rho activating Lbc-type GEFs, in particular Arhgef11 and Arhgef12, are enriched at transient cell protrusions and retractions. Furthermore, via an optogenetic approach, we show that these Lbc GEFs are recruited to the plasma membrane by active Rac, suggesting that they might link cell protrusion and retraction by mediating Rac/Rho activity crosstalk. Indeed, depletion of these GEFs impaired cell protrusion-retraction dynamics, which was accompanied by an increase in migration directionality and reduced migration velocity. Thus, our study shows that Arhgef11 and Arhgef12 facilitate effective exploratory cell migration by coordinating the central cell morphogenic processes of cell protrusion and retraction by coupling the activity of the associated small GTPases Rac and Rho.

Abstract: Local cell contraction pulses play important roles in tissue and cell morphogenesis. Here, we improve a chemo-optogenetic approach and apply it to investigate the signal network that generates these pulses. We use these measurements to derive and parameterize a system of ordinary differential equations describing temporal signal network dynamics. Bifurcation analysis and numerical simulations predict a strong dependence of oscillatory system dynamics on the concentration of GEF-H1, an Lbc-type RhoGEF, which mediates the positive feedback amplification of Rho activity. This prediction is confirmed experimentally via optogenetic tuning of the effective GEF-H1 concentration in individual living cells. Numerical simulations show that pulse amplitude is most sensitive to external inputs into the myosin component at low GEF-H1 concentrations and that the spatial pulse width is dependent on GEF-H1 diffusion. Our study offers a theoretical framework to explain the emergence of local cell contraction pulses and their modulation by biochemical and mechanical signals.