Ric-8A, a GEF for heterotrimeric G-proteins, controls cranial neural crest cell polarity during migration

Abstract

The neural crest (NC) is a transient embryonic cell population that migrates extensively during development. Ric-8A, a guanine nucleotide exchange factor (GEF) for different Gα subunits regulates cranial NC (CNC) cell migration in Xenopus through a mechanism that still remains to be elucidated. To properly migrate, CNC cells establish an axis of polarization and undergo morphological changes to generate protrusions at the leading edge and retraction of the cell rear. Here, we aim to study the role of Ric-8A in cell polarity during CNC cell migration by examining whether its signaling affects the localization of GTPase activity in Xenopus CNC using GTPase-based probes in live cells and aPKC and Par3 as polarity markers. We show that the levels of Ric-8A are critical during migration and affect the localization of polarity markers and the subcellular localization of GTPase activity, suggesting that Ric-8A, probably through heterotrimeric G-protein signaling, regulates cell polarity during CNC migration.

Gαi2 Interaction with EB1 Controls Microtubule Dynamics and Rac1 Activity inXenopusNeural Crest Cell Migration

“…These findings closely align with the results obtained in this investigation, demonstrating that Gαi2 loss of function reduces microtubule catastrophes and promotes tubulin stabilization, resulting in increased localization of active Rac1 at the leading edge and cell-cell contacts. This strikingly contrasts with the normal cranial NC migration phenotype, where Rac1 is suppressed at cell-cell contacts during CIL, leading to a shift in polarity towards the cell-free edge to sustain directed migration (Theveneau et al, 2010;Shoval and Kalcheim, 2012;Leal et al, 2018). In addition, our findings here, where we observed a partial rescue at the Gαi2 morphant migratory behavior using a low concentration of Rac1 inhibitor (NSC23766) strongly support this possible mechanism.…”

“…and Lifeact-GFP (150 pg X.t ., 300 pg X.l .). In the case of mRNA from the GTPase-based probes pGBD-GFP or rGBD-mCherry, injections were carried out the same way as described before (Leal et al, 2018).…”

“…Numerous studies have demonstrated that heterotrimeric G-protein-mediated signals are involved in cell migration, including in NC cells (Cotton and Claing, 2009; Nobes and Hall, 1995; Kjoller and Hall, 1999; Sah et al, 2000; Rohde and Heisenberg, 2007; Fuentealba et al, 2013; Toro-Tapia et al, 2018; Leal et al, 2018). Recent studies have demonstrated that Ric-8A, a Guanine Nucleotide Exchange Factor (GEF) for Gα subunits of heterotrimeric G proteins, plays a regulatory role in NC cell migration by influencing polarity properties (Leal et al, 2018) and facilitating focal adhesion formation through Gα13 subunit (Fuentealba et al, 2013; Toro-Tapia et al, 2018). Nevertheless, all the Gα family subunits from heterotrimeric G proteins have been implicated in cell migration (Cotton and Claing, 2009), and their expression has been observed in the NC region of Xenopus embryos (Fuentealba et al, 2016).…”

“…Gαi2 regulates the subcellular distribution of active Rac1 in cranial NC explants under Gαi2 loss-of-function conditions, considering Rac1 pivotal roles in cranial NC migration and contact inhibition of locomotion (CIL)(Carmona-Fontaine et al, 2011;Moore et al, 2013;Leal et al, 2018). Hence, we employed mRNA encoding Rac1 small GTPase-based probe, enabling specific visualization of the GTP-bound states of this protein.…”

The origin and evolution of vertebrate neural crest cells

“…Another key feature of migratory neural crest is the dynamic regulation of the cellular cytoskeleton accompanied by breakdown of the basal lamina of the neural tube by proteases such as ADAMs and matrix metalloproteases (MMPs) [66][67][68][69][70][71][72][73][74]. During this period, neural crest cells alter their cell polarity and generate a leading edge for long-range and directed migration by genes such as the Rho family of small GTPases [56,57,[75][76][77][78][79][80][81].…”