WNT Protein-independent Constitutive Nuclear Localization of β-Catenin Protein and Its Low Degradation Rate in Thalamic Neurons

Abstract

Nuclear localization of ␤-catenin is a hallmark of canonical Wnt signaling, a pathway that plays a crucial role in brain development and the neurogenesis of the adult brain. We recently showed that ␤-catenin accumulates specifically in mature thalamic neurons, where it regulates the expression of the Ca v 3.1 voltage-gated calcium channel gene. Here, we investigated the mechanisms underlying ␤-catenin accumulation in thalamic neurons. We report that a lack of soluble factors produced either by glia or cortical neurons does not impair nuclear ␤-catenin accumulation in thalamic neurons. We next found that the number of thalamic neurons with ␤-catenin nuclear localization did not change when the Wnt/Dishevelled signaling pathway was inhibited by Dickkopf1 or a dominant negative mutant of Dishevelled3. These results suggest a WNT-independent cellautonomous mechanism. We found that the protein levels of APC, AXIN1, and GSK3␤, components of the ␤-catenin degradation complex, were lower in the thalamus than in the cortex of the adult rat brain. Reduced levels of these proteins were also observed in cultured thalamic neurons compared with cortical cultures. Finally, pulse-chase experiments confirmed that cytoplasmic ␤-catenin turnover was slower in thalamic neurons than in cortical neurons. Altogether, our data indicate that the nuclear localization of ␤-catenin in thalamic neurons is their cell-intrinsic feature, which was WNT-independent but associated with low levels of proteins involved in ␤-catenin labeling for ubiquitination and subsequent degradation.

Novel β-catenin target genes identified in thalamic neurons encode modulators of neuronal excitability

“…The cultures contained both neurons and glia (approximately 1:1), which is vital for the survival of thalamic neurons [38]. …”

“…Thalamic neurons cultured in vitro maintain the nuclear localization of β-catenin [38]. To decrease its level, the cultures were treated with an adenovirus that carried Axin2 , the product of which is a component of the β-catenin destruction complex and as such should reduce its cytoplasmic and nuclear pool.…”

“…Dissociated primary thalamic cells were obtained from the brains of embryonic day 19 Wistar rat embryos as described previously [38]. The cells were plated on coverslips coated with poly-D-lysine (30 μg/ml; Sigma) at a density of 2.5 × 10 5 cells per well of a 24-well plate in Minimal Essential Medium (MEM; Gibco) supplemented with 10% fetal bovine serum and 0.2 mM glutamine (Sigma).…”

“…These functions resemble the role of Wnt/β-catenin signaling during neuronal development. Nonetheless, nuclear β-catenin has also been shown to accumulate in mature neurons - in hippocampal cells upon NMDA (N-Methyl-D-aspartate) receptor activation [35-37] and constitutively in thalamic cells [38]. The identification of β-catenin target genes in neurons may provide insights into its role in these cells and the adult brain.…”

The expression and activity of β-catenin in the thalamus and its projections to the cerebral cortex in the mouse embryo

“…While attention has focussed on the importance of Wnt/β-catenin signalling in neural progenitors in the developing thalamus and elsewhere in the brain, the function of β-catenin activity in post-mitotic projecting neurons remains relatively unexplored [9,21,36,37,43]. The developing embryonic thalamus is a site of high levels of TCF/LEF transcription mediated by nuclear β-catenin [20,57] & present study.…”