DEVELOPMENT 865 RESEARCH ARTICLE INTRODUCTION One of the central questions in developmental biology is how a homogeneous field of cells undergoes patterning to give distinct territories or cell types. Embryos use a variety of strategies to establish such inductive interactions. In some cases, a field of cells responds to a gradient of inducing signals, with different cell types differentiating according to their position relative to the morphogen gradient (Briscoe et al., 1999; De Robertis and Kuroda, 2004; Lander et al., 2002; Mizutani et al., 2005; Stamataki et al., 2005). In other cases, inducing signals act over very short distances to produce an essentially all-or-none response (Bier, 2000; Shilo, 2003). Fine- grained patterning at the cellular level can also emerge in a cell field by cell-cell interactions such as lateral inhibition (Artavanis- Tsakonas et al., 1999; Lai, 2004) or gap junctional communication (Levin, 2002). Despite the great progress in understanding these different signaling strategies, much still remains to be learned about how they are orchestrated in the induction and development of particular organs. The inner ear is an increasingly well-characterized example of embryonic induction. It derives from a simple patch of thickened ectoderm, the otic placode, that arises next to the posterior hindbrain (Barald and Kelley, 2004; Brown et al., 2003; Groves, 2005; Kiernan et al., 2002; Riley and Phillips, 2003; Torres and Giraldez, 1998). Studies in different species have suggested roles for both the hindbrain and cranial paraxial mesoderm in otic placode induction, and whereas the relative contribution of these two tissues to the induction process in different species is still uncertain, it is clear that members of the fibroblast growth factor (Fgf) family play a central and crucial role in this induction in fish, amphibians, birds and mammals (Ladher et al., 2000; Ladher et al., 2005; Leger and Brand, 2002; Liu et al., 2003; Lombardo et al., 1998; Maroon et al., 2002; Phillips et al., 2001; Phillips et al., 2004; Wright and Mansour, 2003). Fgf signaling induces the expression of a variety of molecular markers (such as the transcription factors Pax2 and Pax8) in presumptive placodal ectoderm before the placode becomes morphologically distinct (Alvarez et al., 2003; Ladher et al., 2005; Wright and Mansour, 2003). The induction of Pax2 in cranial ectoderm is commonly thought to be synonymous with the induction of the otic placode. However, several lines of evidence suggest that many Pax2 + ectodermal cells that are initially induced by Fgf signaling will not contribute to the otic placode or the inner ear. Fate-mapping studies in chicken show that cells lying within the Pax2 domain can give rise to structures other than the otocyst, such as the epidermis or epibranchial placodes (Streit, 2002). Genetic fate mapping of Pax2 + ectoderm using Pax2-Cre mice also shows that many Pax2 + cells in the presumptive placodal region ultimately give rise to epidermis and possibly to epibranchial placodes (Ohyama and Groves, 2004b). In light of these observations, what are the mechanisms by which a field of Pax2 + precursor cells is sub-divided into otic placode and epidermis? Activation of the Wnt signaling pathway by Wnt8 family members has been proposed to participate in otic placode induction (Ladher et al., 2000). However, more recent studies in zebrafish suggest that otic placode induction can proceed in the absence of Wnt8 expression in the hindbrain, although the otocysts of such embryos were usually of reduced size (Phillips et al., 2004). It is possible that Wnt signaling is not necessary for the induction of Pax2 + precursor cells, but instead determines the size of the otic placode by instructing these precursor cells to differentiate into placodal tissue, rather than epidermis. In the present study, we show that the canonical Wnt signaling pathway is activated in a subset of Pax2 + cells during early development. Disruption of the canonical Wnt signaling pathway in Pax2 + cells by conditional deletion of the -catenin gene leads to an expansion of cranial epidermis at the expense of the otic placode and otocyst. Conversely, constitutive activation of the canonical Wnt signaling pathway by stabilization of -catenin in Pax2 + cells causes an expansion of the otic placode at the expense of epidermis. Our results suggest that Wnt signaling mediates a placode-epidermis fate decision by acting instructively on a field of Pax2 + precursors to direct them to an otic placode fate. Wnt signals mediate a fate decision between otic placode and epidermis Takahiro Ohyama 1 , Othman A. Mohamed 2 , Makoto M. Taketo 3 , Daniel Dufort 2 and Andrew K. Groves 1, * 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. KEY WORDS: Mouse, Otic Placode, Wnt Development 133, 865-875 doi:10.1242/dev.02271 1 Gonda Department of Cell and Molecular Biology, House Ear Institute, 2100 West 3rd Street, Los Angeles CA 90057, USA. 2 Department of Obstetrics and Gynecology, McGill University, Montreal, Canada. 3 Department of Pharmacology, Graduate School of Medicine, Kyoto University, Kyoto, Japan. *Author for correspondence (e-mail: agroves@hei.org) Accepted 3 January 2006