Biology subjects

Devreotes, P.

Publications and source records attributed to Devreotes, P..

3 recordsLinked to original sources

bioRxiv2024

PIP5K-Ras bistability initiates plasma membrane symmetry breaking to regulate cell polarity and migration

Symmetry breaking is a fundamental process that underlies key cellular behaviors such as cell polarity and migration, but the mechanism - how a uniform plasma membrane spontaneously transitions to an asymmetric state - is still unknown. Here in this study, using a combination of Dictyostelium amoeba, multiple mammalian leukocytes, and human cancer cells and 3D organoid systems, we monitored the localization dynamics of RasGTP and PIP5K, dissected the effects of eliminating, conditionally increasing, or optogenetically manipulating membrane PIP5K levels, screened for key regulators of Ras activation, and tracked single-molecules of PIP5K. Our data converge on a core biochemical circuit involving mutually inhibitory interactions between PIP5K and RasGTP that is both necessary and sufficient for symmetry breaking. The process is initiated by stochastic, localized Ras activation coupled with a decrease in the lifetime of the association of PIP5K with the membrane. A resulting localized reduction in PI(4,5)P2 facilitates the recruitment of a RasGEF to the corresponding domain, amplifying RasGTP production through a positive feedback loop. Dissociated PIP5K relocates to other membrane regions, where it suppresses Ras activation. These events separate the membrane into distinct active and inactive zones, even when receptor inputs or cytoskeletal activities are absent. This same core biochemical circuit controls the spatial organization of downstream PI3K/Akt/Rac signaling and actin/actomyosin activities, which generate the localized protrusions to define polarity and migration mode. While many models have been forwarded to explain polarity or symmetry breaking, the PIP5K-Ras mutually inhibitory bistable circuit we present here is the first universal molecular mechanism to explain the initiation of asymmetry as well as subsequent polarity and migration.

cell biology↗

bioRxiv2023

Electrostatic modulation of signaling at cell membrane: Waveform- and time-dependent electric control of ERK dynamics

Different exogenous electric fields (EF) can guide cell migration, disrupt proliferation, and program cell development. Studies have shown that many of these processes were initiated at the cell membrane, but the mechanism has been unclear, especially for conventionally non-excitable cells. In this study, we focus on the electrostatic aspects of EF coupling with the cell membrane by eliminating Faradaic processes with dielectric-coated microelectrodes, and show that the ERK signaling pathway of epithelial cells (MCF10A) can be both inhibited and activated by AC EF with different amplitude thresholds, peaking times and refractory periods. Interestingly, the ERK responses were sensitive to the waveform and timing of EF stimulation pulses, depicting the characteristics of electrostatic and dissipative interactions. Blocker tests and correlated changes of active Ras on the cell membrane with ERK signals indicated that both EGFR and Ras were involved in the rich ERK dynamics induced by EF. We propose that the frequency-dependent dielectric relaxation process could be an important mechanism to couple EF energy to the cell membrane region and modulate membrane protein-initiated signaling pathways, which can be further explored to precisely control cell behavior and fate with high temporal and spatial resolution.

biophysics↗

bioRxiv2023

Ras-mediated homeostatic control of front-back signaling dictates cell polarity

Studies in the model systems, Dictyostelium amoebae and HL-60 neutrophils, have shown that local Ras activity directly regulates cell motility or polarity. Localized Ras activation on the membrane is spatiotemporally regulated by its activators, RasGEFs, and inhibitors, RasGAPs, which might be expected to create a stable front and back, respectively, in migrating cells. Focusing on C2GAPB in amoebae and RASAL3 in neutrophils, we investigated how Ras activity along the cortex controls polarity. Since existing gene knockout and overexpression studies can be circumvented, we chose optogenetic approaches to assess the immediate, local effects of these Ras regulators on the cell cortex. In both cellular systems, optically targeting the respective RasGAPs to the cell front extinguished existing protrusions and changed the direction of migration, as might be expected. However, when the expression of C2GAPB was induced globally, amoebae polarized within hours. Furthermore, within minutes of globally recruiting either C2GAPB in amoebae or RASAL3 in neutrophils, each cell type polarized and moved more rapidly. Targeting the RasGAPs to the cell backs exaggerated these effects on migration and polarity. Overall, in both cell types, RasGAP-mediated polarization was brought about by increased actomyosin contractility at the back and sustained, localized F-actin polymerization at the front. These experimental results were accurately captured by computational simulations in which Ras levels control front and back feedback loops. The discovery that context-dependent Ras activity on the cell cortex has counterintuitive, unanticipated effects on cell polarity can have important implications for future drug-design strategies targeting oncogenic Ras.

cell biology↗