Ras – A critical regulator in chemotaxis and basal cell motility

How is Ras involved in chemotaxis signaling?

Specific migration in response to chemotaxis occurs in many biological processes, including immune response and patterning formation during development; furthermore, it is also prevalent in pathophysiological settings like cancer metastasis. The process of chemotaxis is comprised of three events, including the extension of stereotypical pseudopodia, the establishment of polarity, and directional sensing². Broadly speaking, cells sense extracellular gradients and move toward higher concentrations through activation of intracellular signaling cascades and pseudopod extension. The molecular events that occur during chemotaxis have been elucidated in model systems like Dictyostelium discoideum. For example, it was discovered that shallow cAMP gradients promote chemotaxis through activation of cAMP receptors and associated heterotrimeric G-proteins⁵. Downstream of these heterotrimeric G-proteins lies Ras, which is a key player during chemotaxis^6,7. Sasaki et al. showed that in response to chemoattractants, Ras is transiently localized and activated at the leading edge of the cell⁶.Downstream targets of Ras in chemotaxis include PI3K, TorC2, sGC, and PLA. Interestingly, a study by Kortholt et al. showed that deletion of these four proteins did not prevent Ras activation by cAMP gradients, and still promoted chemotaxis in response to steep cAMP gradients; however, these four downstream effectors provide a “memory of direction” and can increase the sensitivity for chemotaxis by 150-fold in shallow cAMP gradients⁸.

What is symmetry breaking and why is it needed?

Not only is Ras found at the leading edge of chemotaxing cells, but there is evidence that it is important in symmetry breaking, which is the initiation of polarity, an important event in cell polarization and migration. In 2013, a report showed that during chemotaxis, Ras activation was critical for symmetry breaking, and that distinct Ras GEFs and GAPs were active during the different phases of this mechanism⁹. Similarly, another group identified RapGEF as a downstream regulator of the GPCR, Gα which functioned by balancing Ras and Rap signaling to control chemotaxis¹⁰. The idea that Ras GEFs and GAPs play a critical role in Ras-mediated chemotaxis is supported by two new studies showing that modifying RasGEF or RasGAP locally or globally through optogenetic regulation affected Ras signaling, feedback loops, cytoskeletal changes, and morphological changes in migration^11,12. In another study on Ras GAPs, it was shown that GPCR-mediated activation of Ras activated negative feedback mechanisms¹³. Specifically, the negative regulator, C2GAP1, was activated by Ras at the leading edge of chemotaxing cells, where it was retained at the membrane to inhibit sustained Ras signaling.

Does Ras regulate spontaneous motility?

Cells have an intrinsic ability to produce random cell migration or basal motility to explore their environment when chemoattractants are not present⁴. Early studies on Dictyostelium cells lacking Gβ did not activate downstream PI3K signaling events in response to chemoattractants; however, they still produced random motility in the absence of heterotrimeric G proteins¹⁹, implying that there may be internal signals that sufficiently control this basal motility. Firtel and colleagues identified the internal signal responsible for basal motility as the Ras-PI3K-F-Actin signaling circuit²⁰. The Devreotes group revealed that Ras and PI3K are the key elements of this excitable network that produces chemotaxis-independent movement in basal motility²¹. This was supported by Fukushima et al., who showed that an asymmetric distribution of Ras triggered symmetry breaking to produce polarization and directed movement in basal motility²². Furthermore, another study showed that switching between an amoeba-like migratory mode (repeated extending and retracting of pseudopods) to a keratocyte-like gliding (single broad anterior protrusion) could be achieved by altering Ras/Rap-related activities, further implicating Ras as a symmetry-breaking mechanism that regulates migratory mode determination in basal motility²³. A current study utilized optogenetic tools to specifically control Ras activity through recruitment of GEFs and GAPs to the front or rear of human neutrophils and showed that this controlled activation of Ras was sufficient to control random motility²⁴. Importantly, this Ras-dependent slow, excitable signaling network does not require the cytoskeleton for activation, but is capable of coupling with cytoskeletal networks to control migration²¹. Accordingly, recent data suggest that there are complementary feedback loops between this Ras excitable network and increased branch actin polymerization that control front- and back-states of the cell to alter both cell polarity and migration²⁵. Cytoskeleton offer highly sensitive Ras activation and GEF assays to determine the GTP-bound levels of this protein in cell and tissue extracts (see below for more details).

Summary and future insights

These findings collectively point towards a fundamental role for Ras in both chemotactic migration and cell-directed motility. Remarkably, these Ras-dependent signaling events can be activated through external chemoattractants and chemorepellents, or can occur through excitable events, which suggests that the core Ras circuit is essential for cell movement, but must be specifically controlled spatiotemporally through distinct GEFs and GAPs to effectively control these unique types of motile events. For example, what happens to a cell exposed to both a chemoattractant and chemorepellent agent simultaneously? While the current body of work provides key insights towards our understanding of Ras in directed cell movement, there are still more pieces to uncover to fully understand how these different migratory processes converge on Ras to regulate migration.

References

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