β-Catenin Up-regulates Atoh1 Expression in Neural Progenitor Cells by Interaction with an Atoh1 3′ Enhancer

Abstract

Atoh1, a basic helix-loop-helix transcription factor, plays a critical role in the differentiation of several epithelial and neural cell types. We found that ␤-catenin, the key mediator of the canonical Wnt pathway, increased expression of Atoh1 in mouse neuroblastoma cells and neural progenitor cells, and baseline Atoh1 expression was decreased by siRNA directed at ␤-catenin. The up-regulation of Atoh1 was caused by an interaction of ␤-catenin with the Atoh1 enhancer that could be demonstrated by chromatin immunoprecipitation. We found that two putative Tcf-Lef sites in the 3 enhancer of the Atoh1 gene displayed an affinity for ␤-catenin and were critical for the activation of Atoh1 transcription because mutation of either site decreased expression of a reporter gene downstream of the enhancer. Tcf-Lef co-activators were found in the complex that bound to these sites in the DNA together with ␤-catenin. Inhibition of Notch signaling, which has previously been shown to induce bHLH transcription factor expression, increased ␤-catenin expression in progenitor cells of the nervous system. Because this could be a mechanism for up-regulation of Atoh1 after inhibition of Notch, we tested whether siRNA to ␤-catenin prevented the increase in Atoh1 and found that ␤-catenin expression was required for increased expression of Atoh1 after Notch inhibition.Progenitor cells in several tissues require the basic helixloop-helix (bHLH) 3 transcription factor, Atoh1, for their development into mature neurons or epithelial cells (1, 2). Upstream regulators of Atoh1 are likely to have an important role in the regulation of development in the central and peripheral nervous systems and in the intestinal epithelium, all of which rely on Atoh1 for differentiation. This finding was clear from the analysis of an Atoh1-null mouse, which lacks many of the cell types of the intestinal epithelium, and has incomplete development of cerebellar and spinal neurons and a complete lack of inner ear hair cells (1). The expression of bHLH transcription factors is partly regulated by components of the Notch pathway (3-5), but these may be only a part of the complex regulatory circuits governing the timing and amount of bHLH transcription factor expression as well as the tissue specificity of expression. The Wnt pathway plays a key role in early development of several of these tissues, including the intestinal epithelium and the inner ear (6 -11), and is thus a potential candidate for upstream signaling leading to Atoh1 expression. Indeed, disruption of Wnt signaling prevents intestinal epithelial differentiation to mature cell types and is accompanied by decreased expression of Atoh1 (8).In a search for genes that affected Atoh1 expression, a number of genes were tested for their effect on Atoh1 expression by screening of an adenoviral library that allowed us to express the genes in various cell types. One such gene was ␤-catenin, the intracellular mediator of the canonical Wnt pathway. Its overexpression in neural progenitor cell types increased acti...

Dynamic Expression of Lgr5, a Wnt Target Gene, in the Developing and Mature Mouse Cochlea

“…Interestingly, Lgr5-EGFP expression rapidly disappears in hair cells and a subset of supporting cells. The notion that Wnt signaling plays a role in cell fate determination in the inner ear is supported by two previous studies: overexpression of ß-catenin, the central player of the canonical Wnt pathway, upregulates Atoh1 (Shi et al 2010) and induces ectopic sensory patches (Stevens et al 2003). While our data indicate that Lgr5 is not required for cochlear development, its expression likely reflects the pattern of Wnt-activated cells within the cochlear ductal epithelium.…”

Distinct capacity for differentiation to inner ear cell types by progenitor cells of the cochlea and vestibular organs

“…Notchmediated lateral inhibition (Daudet and Lewis, 2005;Lanford et al, 1999) and innate Wnt signaling are both crucial for the development of HCs (Shi et al, 2013. The Wnt pathway interacts with the Notch pathway in development (Collu et al, 2014;Shi et al, 2010) and this interaction could regulate the level of expression of Atoh1 and other genes. Bypassing key components of developmental pathways via viral Atoh1 transduction might disrupt mechanisms that control levels of Wnt and Notch factors and thus create 'primordial' HCs.…”

Atoh1, an Essential Transcription Factor in Neurogenesis and Intestinal and Inner Ear Development: Function, Regulation, and Context Dependency

“…Several proteins have been identified that directly interact with an Atoh1 enhancer as demonstrated by chromatin immunoprecipitation, electrophoretic mobility shift assays, or luciferase activity, for example, Zic1, cdx2, Hfn1a, β-catenin, and Hic1 (Akazawa et al 1995;Ebert et al 2003;Mutoh et al 2006;Briggs et al 2008;D'Angelo et al 2010;Shi et al 2010). Atoh1, Sox2, β-catenin, Hfn1a, and Cdx2 upregulate Atoh1 expression (Helms et al 2000;Ebert et al 2003;Gazit et al 2004;Mutoh et al 2006;D'Angelo et al 2010;Shi et al 2010;Neves et al 2011). Hes1, Hes5, Sox2, Hic1, Ngn1, NeuroD1, and Zic1 repress Atoh1 at the level of transcription (Akazawa et al 1995;Gowan et al 2001;Ebert et al 2003;Qian et al 2006;Briggs et al 2008;Dabdoub et al 2008;Pan et al 2009;Jahan et al 2010).…”