The Small GTPase RhoA Is Required to Maintain Spinal Cord Neuroepithelium Organization and the Neural Stem Cell Pool

Abstract

The regulation of adherens junctions (AJs) is critical for multiple events during CNS development, including the formation and maintenance of the neuroepithelium. We have addressed the role of the small GTPase RhoA in the developing mouse nervous system using tissue-specific conditional gene ablation. We show that, in the spinal cord neuroepithelium, RhoA is essential to localize N-cadherin and ␤-catenin to AJs and maintain apical-basal polarity of neural progenitor cells. Ablation of RhoA caused the loss of AJs and severe abnormalities in the organization of cells within the neuroepithelium, including decreased neuroepithelial cell proliferation and premature cell-cycle exit, reduction of the neural stem cell pool size, and the infiltration of neuroepithelial cells into the lumen of the ventricle. We also show that, in the absence of RhoA, its effector, mammalian diaphanous-related formin1 (mDia1), does not localize to apical AJs in which it likely stabilizes intracellular adhesion by promoting local actin polymerization and microtubule organization. Furthermore, expressing a dominant-negative form of mDia1 in neural stem/progenitor cells results in a similar phenotype compared with that of the RhoA conditional knock-out, namely the loss of AJs and apical polarity. Together, our data show that RhoA signaling is necessary for AJ regulation and for the maintenance of mammalian neuroepithelium organization preventing precocious cell-cycle exit and differentiation.

Cell Type-specific Signaling Function of RhoA GTPase: Lessons from Mouse Gene Targeting

“…Deletion of rhoA results in dysplasia and disrupts the distribution of adherens junction components such as N-cadherin, ␣E-catenin, ␤-catenin, and F-actin (37-39). In rhoA knock-outs, rosette-like structures, seen previously in N-cadherin null mutants (40), were observed in the brain and spinal cord, and cells were found to invade the lumen of the neural tube (38), suggesting a disturbance of adherens junctions and a disruption of cell-cell contact in the neuroepithelium.…”

“…Axons of rhoA-deficient corticospinal neurons and interneurons display a similar midline-crossing phenotype, as well as a loss of ephrin B3 expression at the spinal cord midline, causing a deficiency in ephrin B3-EphA4 signaling (Fig. 2D) (38). In contrast to corticospinal neurons or interneurons, the central and peripheral projections of rhoA null (driven by Wnt1-Cre) dorsal root ganglion (DRG) neurons appear normal (46).…”

“…(driven by Brn4-Cre) reduced neuron proliferation and caused precocious cell cycle exit, perhaps as a result of premature differentiation of neural progenitor cells (38). Interestingly, phosphohistone 3-labeled mitotic cells dispersed throughout the neuroepithelium in rhoA-deficient brain and spinal cord, instead of the tight luminal localization in the wild-type neural tube (37,38), suggesting an alteration of the neural progenitor niche.…”

“…Not surprisingly, effector knock-out and rhoA knock-out mouse models have been reported to show both consistent and discordant phenotypes. For example, neuroepithelial integrity was impaired in both rhoA and mDia knock-outs (37,38,70), suggesting that the RhoA-mDia axis might be critical in regulating adherens junctions in the developing central nervous system. In contrast, although the rhoA-deleted central nervous system showed no obvious cytokinesis defects, knocking out citron kinase in mice caused a severe cytokinesis blockage (71).…”

Rho GTPases in embryonic development

“…51 Moreover, the tissue-specific deletion of RhoA in the neuroepithelium of the developing spinal cord has also highlighted the critical role of RhoA in the maintenance of apical adherens junction integrity, organization of the neuroepithelium in the developing spinal cord and left-right locomotor behavior. 52,53 Three independent groups have investigated the effect of a forebrain-specific deletion of Rac1 beginning at E9.5, 54-56 after neural tube closure. This conditional Rac1 deletion led to late embryonic lethality.…”