July 15, 2003;
Distinct functions of Rho and Rac are required for convergent extension during Xenopus gastrulation.
We have undertaken the first detailed analysis of Rho
GTPase function during vertebrate development by analyzing how RhoA
control convergent extension of axial mesoderm
during Xenopus gastrulation. Monitoring of a number of parameters in time-lapse recordings of mesoderm
explants revealed that Rac
have both distinct and overlapping roles in regulating the motility of axial mesoderm
cells. The cell behaviors revealed by activated or inhibitory versions of these GTPases in native tissue
were clearly distinct from those previously documented in cultured fibroblasts. The dynamic properties and polarity of protrusive activity, along with lamellipodia formation, were controlled by the two GTPases operating in a partially redundant manner, while Rho
contributed separately to cell shape and filopodia formation. We propose that Rho
operate in distinct signaling pathways that are integrated to control cell motility during convergent extension.
[+] show captions
Fig. 1. Mesoderm induction occurs normally in Rho GTPase-injected embryos but differentiated mesoderm is disorganized. mRNA (2 ng) encoding a dominant negative or constitutively active Rho GTPase was microinjected into the presumptive dorsoanterior mesoderm at the two to four-cell stage. The embryos were cultured until stage 10.5 for in situ hybridization (A–J) or stage 26 for whole-mount immunohistochemistry (K–P). In situ hybridization using cRNA probes for Xbra (Xenopus brachyury; A–E) and Gsc (goosecoid; F–J). The normal expression pattern of Xbra (A) and gsc (F) was unaffected by overexpression of dominant negative (B, D, G, I) or constitutively active (C, E, H, J) Rho GTPase. Vegetal views of embryos are shown (A–J). Whole-mount immunohistochemistry of embryos double stained with notochord-specific Tor70 (black) and somite-specific 12/101 (brown) antibodies revealed that the pattern of differentiated mesoderm was disorganized in Rho GTPase-injected embryos (L, M, O, P) compared with controls (K, N). Two views of control embryos are shown (K, lateral view; N, dorsal view). Severe examples of Rho GTPase-injected embryos are shown in (L; dorsal view) dnRho, (M; dorsal views) caRho, (O; lateral view) dnRac, and (P; lateral view) caRac. Rho GTPase-injected embryos developed with spina bifida and reduced head structures. Whereas differentiated somites and notochord were present in these embryos, both sets of structures were shorter than normal and disorganized.
Notochord Ab1 (Tor 70), in X laevis at NF stage , 26, laterl view, dorsal up, anterior right, showing expression in notochord.