β-Catenin signaling is required for neural differentiation of embryonic stem cells

Culture of embryonic stem (ES) cells at high density inhibits bothβ-catenin signaling and neural differentiation. ES cell density does not influence β-catenin expression, but a greater proportion ofβ-catenin is targeted for degradation in high-density cultures. Moreover,in high-density cultures, β-catenin is preferentially localized to the membrane further reducing β-catenin signaling. Increasing β-catenin signaling by treatment with Wnt3a-conditioned medium, by overexpression ofβ-catenin, or by overexpression of a dominant-negative form of E-cadherin promotes neurogenesis. Furthermore, β-catenin signaling is sufficient to induce neurogenesis in high-density cultures even in the absence of retinoic acid (RA), although RA potentiates the effects of β-catenin. By contrast,RA does not induce neurogenesis in high-density cultures in the absence ofβ-catenin signaling. Truncation of the armadillo domain ofβ-catenin, but not the C terminus or the N terminus, eliminates its proneural effects. The proneural effects of β-catenin reflect enhanced lineage commitment rather than proliferation of neural progenitor cells. Neurons induced by β-catenin overexpression either alone or in association with RA express the caudal neuronal marker Hoxc4. However, RA treatment inhibits the β-catenin-mediated generation of tyrosine hydroxylase-positive neurons, suggesting that not all of the effects of RA are dependent upon β-catenin signaling. These observations suggest thatβ-catenin signaling promotes neural lineage commitment by ES cells, and that β-catenin signaling may be a necessary co-factor for RA-mediated neuronal differentiation. Further, enhancement of β-catenin signaling with RA treatment significantly increases the numbers of neurons generated from ES cells, thus suggesting a method for obtaining large numbers of neural species for possible use in for ES cell transplantation.