Dorsal dermis and epaxial muscle have been shown to arise from the central dermomyotome in the chick. En1 is a homeobox transcription factor gene expressed in the central dermomyotome. We show by genetic fate mapping in the mouse that En1-expressing cells of the central dermomyotome give rise to dorsal dermis and epaxial muscle and, unexpectedly, to interscapular brown fat. Thus, the En1-expressing central dermomyotome normally gives rise to three distinct fates in mice. Wnt signals are important in early stages of dermomyotome development, but the signal that acts to specify the dermal fate has not been identified. Using a reporter transgene for Wnt signal transduction, we show that the En1-expressing cells directly underneath the surface ectoderm transduce Wnt signals. When the essential Wnt transducer beta-catenin is mutated in En1 cells, it results in the loss of Dermo1-expressing dorsal dermal progenitors and dermis. Conversely, when beta-catenin was activated in En1 cells, it induces Dermo1 expression in all cells of the En1 domain and disrupts muscle gene expression. Our results indicate that the mouse central dermomyotome gives rise to dermis, muscle, and brown fat, and that Wnt signalling normally instructs cells to select the dorsal dermal fate.

The otic placode, the anlagen of the inner ear, develops from an ectodermal field characterized by expression of the transcription factor Pax2. Previous fate mapping studies suggest that these Pax2 + cells will give rise to both otic placode tissue and epidermis, but the signals that divide the Pax2 + field into placodal and epidermal territories are unknown. We report that Wnt signaling is normally activated in a subset of Pax2 + cells, and that conditional inactivation of ␤-catenin in these cells causes an expansion of epidermal markers at the expense of the otic placode. Conversely, conditional activation of ␤-catenin in Pax2 + cells causes an expansion of the otic placode at the expense of epidermis, and the resulting otic tissue expresses exclusively dorsal otocyst markers. Together, these results suggest that Wnt signaling acts instructively to direct Pax2 + cells to an otic placodal, rather than an epidermal, fate and promotes dorsal cell identities in the otocyst.

The neuroepithelial layer of the developing eyecup contains multipotential precursor cells that give rise to all of the neurons and the one glial cell type present in the adult retina. Patterning within the retinal neuroepithelium is regulated by cell intrinsic as well as cell extrinsic mechanisms. Although the identity of some of the signaling molecules that regulate retinal development is known, the function of many others, especially members of the Wnt family, has yet to be characterized in the context of retinal development. We undertook a comprehensive in situ hybridization analysis to examine the expression of Wnt pathway components in the developing and adult mouse neural retina. Our findings confirm and extend previous expression studies in mice and other vertebrates, as we show that Wnt-3, -5a, -5b, and -7b are expressed in the neural retina and that there is a dynamic pattern of Wnt receptor (Mouse frizzled

␤-Catenin signaling has been shown to be involved in triggering axis formation in several organisms, including Xenopus and zebrafish. Genetic analysis has demonstrated that the Wnt/␤-catenin signaling pathway is also involved in axis formation in the mouse, since a targeted deletion of ␤-catenin results in embryos that have a block in anterior-posterior axis formation, fail to initiate gastrulation, and do not form mesoderm. However, because ␤-catenin is ubiquitously expressed, the precise time and cell types in which this signaling pathway is active during early embryonic development remain unknown. Thus, to better understand the role of the Wnt/␤-catenin signaling pathway in axis formation and mesoderm specification, we have examined both the distribution and signaling activity of ␤-catenin during early embryonic development in the mouse. We show that the N-terminally nonphosphorylated form of ␤-catenin as well as ␤-catenin signaling is first detectable in the extraembryonic visceral endoderm in day 5.5 embryos. Before the initiation of gastrulation at day 6.0, ␤-catenin signaling is asymmetrically distributed within the epiblast and is localized to a small group of cells adjacent to the embryonic-extraembryonic junction. At day 6.5 and onward, ␤-catenin signaling was detected in the primitive streak and mature node. Thus, ␤-catenin signaling precedes primitive streak formation and is present in epiblast cells that will go on to form the primitive streak. These results support a critical role for the Wnt/␤-catenin pathway in specifying cells to form the primitive streak and node in the mammalian embryo as well as identify a novel domain of Wnt/␤-catenin signaling activity during early embryogenesis. Developmental Dynamics 231:416 -424, 2004.

Extracellular signal-regulated protein kinase (ERK, or mitogen-activated protein kinase [MAPK]) regulatory cascades in fungi turn on transcription factors that control developmental processes, stress responses, and cell wall integrity. CEK1 encodes aCandida albicans MAPK homolog (Cek1p), isolated by its ability to interfere with the Saccharomyces cerevisiae MAPK mating pathway. C. albicans cells with a deletion of theCEK1 gene are defective in shifting from a unicellular budding colonial growth mode to an agar-invasive hyphal growth mode when nutrients become limiting on solid medium with mannitol as a carbon source or on glucose when nitrogen is severely limited. The same phenotype is seen in C. albicans mutants in which the homologs (CST20, HST7, and CPH1) of the S. cerevisiae STE20, STE7, andSTE12 genes are disrupted. In S. cerevisiae, the products of these genes function as part of a MAPK cascade required for mating and invasiveness of haploid cells and for pseudohyphal development of diploid cells. Epistasis studies revealed that theC. albicans CST20, HST7, CEK1, andCPH1 gene products lie in an equivalent, canonical, MAPK cascade. While Cek1p acts as part of the MAPK cascade involved in starvation-specific hyphal development, it may also play independent roles in C. albicans. In contrast to disruptions of theHST7 and CPH1 genes, disruption of theCEK1 gene adversely affects the growth of serum-induced mycelial colonies and attenuates virulence in a mouse model for systemic candidiasis.

Successful implantation relies on precisely orchestrated and reciprocal signaling between the implanting blastocyst and the receptive uterus. We have examined the role of the Wnt͞␤-catenin signaling pathway during the process of implantation and demonstrate that this pathway is activated during two distinct stages. Wnt͞␤-catenin signaling is first transiently activated in circular smooth muscle forming a banding pattern of activity within the uterus on early day 4. Subsequently, activation is restricted to the luminal epithelium at the prospective site of implantation. Activation at both sites requires the presence of the blastocyst. Furthermore, inhibition of Wnt͞␤-catenin signaling interferes with the process of implantation. Our results demonstrate that the Wnt͞␤-catenin signaling pathway plays a central role in coordinating uterus-embryo interactions required for implantation.blastocyst ͉ uterus A crucial event during mammalian embryonic development is the process of implantation, during which the free-living blastocyst attaches to the uterine endometrium. Successful implantation depends on precisely orchestrated and reciprocal signaling between the implanting blastocyst and the receptive uterus (1). For instance, blastocysts can only implant once they have been activated by uterine factor(s) that are regulated by ovarian steroid hormones (2). Conversely, expression of several uterine genes are regulated by a signal(s) emanating from activated blastocysts (3, 4). Multiple signaling pathways have been shown to participate in the implantation process, and several cytokines and growth factors have been shown to be expressed in the uterus at the time of implantation and to play important roles in this process (5). However, there is increasing evidence that members of other families of growth factors implicated in embryogenesis may also participate in the implantation process. It has been demonstrated that members of the hedgehog, bone morphogenetic, and Wnt proteins are expressed in the uterus at the time of implantation (6). However, the precise role of these different growth factor signaling pathways in the implantation process has not been determined. Furthermore, the cell types within the uterus that respond to these growth factors are not known. In this study, we set out to determine the role of the canonical Wnt͞␤-catenin signaling pathway in the implantation process. We demonstrate that this pathway is activated during two distinct stages before implantation. Signaling is transiently detected in circular smooth muscle forming a banding pattern of activity. Subsequently, activation is restricted to the luminal epithelium at the prospective site of implantation. Activation at both sites requires the presence of the blastocyst. Furthermore, we show that inhibition of Wnt͞␤-catenin signaling interferes with the process of implantation. Materials and MethodsMating and Experimental Manipulation of Transgenic Animals. The generation and characterization of the TCF͞Lef-LacZ transgenic mice have been described in ...

The canonical WNT signaling pathway plays a crucial role in patterning of the embryo during development, but little is known about the specific developmental events which are under WNT control. To understand more about how the WNT pathway orchestrates mammalian organogenesis, we studied the canonical ␤-catenin-mediated WNT signaling pathway in kidneys of mice bearing a ␤-catenin-responsive TCF/␤Gal reporter transgene. In metanephric kidney, intense canonical WNT signaling was evident in epithelia of the branching ureteric bud and in nephrogenic mesenchyme during its transition into renal tubules. WNT signaling activity is rapidly downregulated in maturing nephrons and becomes undetectable in postnatal kidney. Sites of TCF/␤Gal activity are in proximity to the known sites of renal WNT2b and WNT4 expression, and these WNTs stimulate TCF reporter activity in kidney cell lines derived from ureteric bud and metanephric mesenchyme lineages. When fetal kidney explants from HoxB7/GFP mice were exposed to the canonical WNT signaling pathway inhibitor, Dickkopf-1, arborization of the ureteric bud was significantly reduced. We conclude that restricted zones of intense canonical WNT signaling drive branching nephrogenesis in fetal kidney. nephrogenesis; ␤-catenin; branching morphogenesis THE WNT FAMILY is comprised of 19 secreted glycoproteins which act as short-range intercellular signaling molecules, recognizing one of the 10 frizzled receptors expressed at the surface of nearby target cells. The canonical signaling pathway is activated by WNTs which bind to cognate frizzled receptors heterodimerized with LRP5 or LRP6 coreceptors (2). Activated receptors recruit dishevelled protein (Dvl) and inhibit degradation of cytoplasmic ␤-catenin via the GSK3␤-axin-APC complex (11). When its degradation is blocked, cytoplasmic ␤-catenin is available to translocate to the nucleus, dimerize with partners belonging to the T-cell factor (TCF) family, and activate target genes. TCF recognition motifs have been well-studied, allowing design of vectors (e.g., TOPFlash) which drive transcription of reporter genes in response to canonical WNT signaling activity (32). In general, canonical ␤-catenin/TCF signaling is thought to activate gene targets (e.g., c-myc) involved in cell proliferation (3, 27).More than 35 years ago, Unsworth and Grobstein (33) reported that tissue from spinal cord could induce formation of renal tubules when cocultured with isolated metanephric mesenchyme. In 1994, Herzlinger et al. (12) found that WNT1-expressing NIH3T3 cells were also able to induce tubule formation in the coculture assay, suggesting that the canonical (␤-catenin-mediated) WNT signaling pathway is essential for mammalian nephrogenesis. However, the precise function of canonical WNT signaling in renal development is unknown. Surprisingly, WNT1 is not present in the developing kidney, but numerous other WNTs are transiently expressed in specific cell lineages (34). Several of these are able to activate the canonical signaling pathway in other context...

The TCF/Lef-LacZ transgene is a faithful reporter of canonical Wnt signaling in the retina. The pattern of TCF/Lef-LacZ reporter gene activation and of TCF/Lef transcription factor expression suggests that activation of the canonical Wnt pathway is developmental-stage dependent and is spatially modulated. Our findings also imply the involvement of this pathway in the specification and/or generation of ciliary epithelium, cellular differentiation, axon guidance, and connectivity to targets in the central nervous system and in the maintenance or function of specific retinal neurons in the adult.