June 1, 2005;
Distinct PAR-1 proteins function in different branches of Wnt signaling during vertebrate development.
The kinase PAR-1 plays conserved roles in cell polarity. PAR-1 has also been implicated in axis establishment in C. elegans and Drosophila and in Wnt signaling, but its role in vertebrate development is unclear. Here we report that PAR-1 has two distinct and essential roles in axial development in Xenopus mediated by different PAR-1 isoforms. Depletion of PAR-1A or PAR-1BX causes dorsoanterior deficits, reduced Spemann organizer
gene expression, and inhibition of canonical Wnt-beta-catenin signaling. By contrast, PAR-1BY depletion inhibits cell movements and localization of Dishevelled
protein to the cell cortex, processes associated with noncanonical Wnt signaling. PAR-1 phosphorylation sites in Dishevelled
are required for this translocation, but not for canonical Wnt signaling. We conclude that PAR-1BY is required in the PCP branch and mediates Dsh
membrane localization while PAR-1A and PAR-1BX are essential for canonical signaling to beta-catenin, possibly via targets other than Dishevelled
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Figure 3. PAR-1A and PAR-1BX but Not PAR-1BY Depletion Reduces Expression of Organizer Genes(A) PAR-1A and PAR-1BX MOs inhibit organizer-specific expression of Xnr3 and chordin while Control MO and PAR-1BY MO do not. Embryos were injected with the amount of MO per embryo given in panel (C) at the 4-cell stage in both dorsal blastomeres. Arrows indicate the approximate injection site; light blue stain visible in some panels is due to staining for coinjected of βGal RNA (50 pg) used as a lineage tracer.(B) Control MO, PAR-1A MO, and PAR-1BY MO have little effect on expression of ventrolateral marginal zone marker gene Xvent-2 or pan-mesodermal marker Xbra. Embryos are shown in vegetal view, dorsal at the top at stage 10.5–11, except for PAR-1BY MO-injected embryos in (B), which are stage 12.(C) Penetrance of the effects of PAR-1 MOs on organizer genes assayed in situ, including goosecoid and Otx-2 as well as those shown in (A). Doses of PAR-1 MOs are shown at the bottom of the histogram are per embryo. Control MO does not noticeably affect development of these markers. Numbers of injected embryos are shown above the bars.
Figure 4. Rescue of the Inhibition of Organizer Gene Expression in PAR-1-Depleted Embryos by PAR-1 mRNA(A) Organizer gene inhibition by PAR-1 MOs rescued by corresponding RNA injection. Embryos were injected with the indicated MOs as in Figure 5 with or without 350 pg per blastomere of the indicated MO-resistant myc-tagged RNAs. Stage 10.5–11 embryos are shown in vegetal view, dorsal at the top. PAR-1 RNA injection results in the modest expansion of their expression domain (right) relative to controls.(B) Quantitation of rescue of PAR-1 MO-induced inhibition of organizer marker genes by PAR-1 mRNA. Expression of gsc and chordin was analyzed in embryos injected with PAR-1A MO (40 ng per blastomere), and expression of Xnr3 was analyzed in PAR-1BX MO-injected samples (10 ng per blastomere). The corresponding PAR-1 mRNA was injected at 350 pg per blastomere.