The PDZ domain protein Mcc is a novel effector of non-canonical Wnt signaling during convergence and extension in zebrafish.

During vertebrate gastrulation, a complex set of mass cellular rearrangements shapes the embryonic body plan and appropriately positions the organ primordia. In zebrafish and Xenopus, convergence and extension (CE) movements simultaneously narrow the body axis mediolaterally and elongate it from head to tail. This process is governed by polarized cell behaviors that are coordinated by components of the non-canonical, ß-catenin-independent Wnt signaling pathway, including Wnt5b and the transmembrane planar cell polarity (PCP) protein Vangl2. However, the intracellular events downstream of Wnt/PCP signals are not fully understood. Here, we show that zebrafish mutated in colorectal cancer (mcc), which encodes an evolutionarily conserved PDZ domain-containing putative tumor suppressor, is required for Wnt5b/Vangl2 signaling during gastrulation. Knockdown of mcc results in CE phenotypes similar to loss of vangl2 and wnt5b, whereas overexpression of mcc robustly rescues the depletion of wnt5b, vangl2 and the Wnt5b tyrosine kinase receptor ror2. Biochemical experiments establish a direct physical interaction between Mcc and the Vangl2 cytoplasmic tail. Lastly, CE defects in mcc morphants are suppressed by downstream activation of RhoA and JNK. Taken together, our results identify Mcc as a novel intracellular effector of non-canonical Wnt5b/Vangl2/Ror2 signaling during vertebrate gastrulation.

Cdx2 contributes to the expansion of the early primordial germ cell population in the mouse.

Cdx gene products regulate the extent of axial elongation from the posterior growth zone. These transcription factors sustain the emergence of trunk and tail tissues by providing a suitable niche in the axial progenitor zone, via regulation of Wnt signaling. Cdx genes are expressed in and along the complete primitive streak including its posterior part wherefrom the extraembryonic mesoderm of the allantois emerges. Cdx genes are required for the full development of the allantois and its derivatives in the placental labyrinth. The mouse germ cell lineage also originates from the proximo-posterior epiblast of the primitive streak, and is established within the extraembryonic mesoderm that generates the allantois. We asked whether the expression of Cdx genes around the newly specified PGCs is necessary for the maintenance and expansion of this population, as it is for the allantois and axial progenitors. We observed a significantly lower number of PGCs in Cdx2(null) embryos than in controls. We found that Wnt3a loss of function decreases the PGC population to the same extent as Cdx2 inactivation. Moreover, exogenous Wnt3a corrects the lower PGC number in Cdx2(null) posterior embryonic tissues cultured in vitro. Cdx2 is not expressed in PGCs themselves, and we propose that the expression of Cdx2 in posterior extraembryonic tissues contributes to the proper niche of the germ cell progenitors by stimulating canonical Wnt signaling. Since PGC residence within the posterior growth zone is a mouse-specific feature, our data suggest that mouse PGCs opportunistically became dependent on the axial progenitor niche.

Mutated in colorectal cancer (Mcc), a candidate tumor suppressor, is dynamically expressed during mouse embryogenesis.

Mutated in Colorectal Cancer (MCC) encodes a multiple PSD-95/Dlg/ZO-1 (PDZ) domain-containing protein implicated, as its name suggests, in the pathogenesis of human colon cancer. To date, however, what role, if any, MCC plays in normal tissue homeostasis and development remains unclear. In an effort to expand our understanding of MCC function and distribution, we examined the expression of the evolutionarily conserved mouse Mcc homolog between embryonic days (E) 6.5 and 12.5 using conventional whole-mount in situ hybridization and two independent Mcc reporter alleles. Mcc is expressed in the posterior primitive streak during gastrulation and in diverse tissues of both mesodermal and endodermal origin. In addition, Mcc transcripts localize to the posterior neural tube and identify discrete neuronal subtypes and ganglia within the developing central nervous system. Genetically, however, Mcc is entirely dispensable, as mice homozygous for the Mcc(Gt(D062B07)) gene trap allele, which generates a loss-of-function mutation, are viable and fertile, with no ostensible phenotype.

Concerted involvement of Cdx/Hox genes and Wnt signaling in morphogenesis of the caudal neural tube and cloacal derivatives from the posterior growth zone.

Decrease in Cdx dosage in an allelic series of mouse Cdx mutants leads to progressively more severe posterior vertebral defects. These defects are corrected by posterior gain of function of the Wnt effector Lef1. Precocious expression of Hox paralogous 13 genes also induces vertebral axis truncation by antagonizing Cdx function. We report here that the phenotypic similarity also applies to patterning of the caudal neural tube and uro-rectal tracts in Cdx and Wnt3a mutants, and in embryos precociously expressing Hox13 genes. Cdx2 inactivation after placentation leads to posterior defects, including incomplete uro-rectal septation. Compound mutants carrying one active Cdx2 allele in the Cdx4-null background (Cdx2/4), transgenic embryos precociously expressing Hox13 genes and a novel Wnt3a hypomorph mutant all manifest a comparable phenotype with similar uro-rectal defects. Phenotype and transcriptome analysis in early Cdx mutants, genetic rescue experiments and gene expression studies lead us to propose that Cdx transcription factors act via Wnt signaling during the laying down of uro-rectal mesoderm, and that they are operative in an early phase of these events, at the site of tissue progenitors in the posterior growth zone of the embryo. Cdx and Wnt mutations and premature Hox13 expression also cause similar neural dysmorphology, including ectopic neural structures that sometimes lead to neural tube splitting at caudal axial levels. These findings involve the Cdx genes, canonical Wnt signaling and the temporal control of posterior Hox gene expression in posterior morphogenesis in the different embryonic germ layers. They shed a new light on the etiology of the caudal dysplasia or caudal regression range of human congenital defects.

Cdx and Hox genes differentially regulate posterior axial growth in mammalian embryos.

Hox and Cdx transcription factors regulate embryonic positional identities. Cdx mutant mice display posterior body truncations of the axial skeleton, neuraxis, and caudal urorectal structures. We show that trunk Hox genes stimulate axial extension, as they can largely rescue these Cdx mutant phenotypes. Conversely, posterior (paralog group 13) Hox genes can prematurely arrest posterior axial growth when precociously expressed. Our data suggest that the transition from trunk to tail Hox gene expression successively regulates the construction and termination of axial structures in the mouse embryo. Thus, Hox genes seem to differentially orchestrate posterior expansion of embryonic tissues during axial morphogenesis as an integral part of their function in specifying head-to-tail identity. In addition, we present evidence that Cdx and Hox transcription factors exert these effects by controlling Wnt signaling. Concomitant regulation of Cyp26a1 expression, restraining retinoic acid signaling away from the posterior growth zone, may likewise play a role in timing the trunk-tail transition.

Hox, Cdx, and anteroposterior patterning in the mouse embryo.

Cdx and Hox gene families descend from the same ProtoHox cluster, already present in the common ancestors of bilaterians and cnidarians, and thought to act by providing anteroposterior (A-P) positional identity to axial tissues in all bilaterians. Mouse Cdx and Hox genes still exhibit common features in their early expression and function. The initiation and early shaping of Hox and Cdx transcriptional domains in mouse embryos are very similar, in keeping with their common involvement in conveying A-P information to the nascent tissues during embryonic axial elongation. Considerations of the impact on axial patterning of the early expression phase of these genes that correlates with the temporally collinear expression of 3'-5'Hox genes suggest that it is concerned with the acquisition of A-P information by the three germ layers as the axis extends. This early A-P information acquired by all cells emerging from the primitive streak or tailbud and their neighbors in the caudal neural plate gets further modulated by the second phase of gene expression occurring later as the tissues mature and differentiate along the growing axis. We discuss the possibility that regulatory phase 1, common to all Cdx and Hox genes, is inherent to the concerted mechanism sequentially turning on 3'-5'Hox genes at early stages, and keeping expression of the initiated genes subsequently in the new materials added posteriorly at the axis extends. The posterior Hox gene expression domain would be subsequently complemented by Hox regulatory phase 2, consisting in a variety of gene-specific, region-specific, and/or tissue-specific gene expression controls. We also touch on the unanswered question whether vertebrate Cdx gene expression delivers A-P positional information in its own right, as Caudal does in Drosophila, or whether it does so exclusively by upregulating Hox genes.