Learning impairments and molecular changes in the brain caused by β-catenin loss

Abstract

Intellectual disability (ID), defined as IQ<70, occurs in 2.5% of individuals. Elucidating the underlying molecular mechanisms is essential for developing therapeutic strategies. Several of the identified genes that link to ID in humans are predicted to cause malfunction of β-catenin pathways, including mutations in CTNNB1 (β-catenin) itself. To identify pathological changes caused by β-catenin loss in the brain, we have generated a new β-catenin conditional knockout mouse (β-cat cKO) with targeted depletion of β-catenin in forebrain neurons during the period of major synaptogenesis, a critical window for brain development and function. Compared with control littermates, β-cat cKO mice display severe cognitive impairments. We tested for changes in two β-catenin pathways essential for normal brain function, cadherin-based synaptic adhesion complexes and canonical Wnt (Wingless-related integration site) signal transduction. Relative to control littermates, β-cat cKOs exhibit reduced levels of key synaptic adhesion and scaffold binding partners of β-catenin, including N-cadherin, α-N-catenin, p120ctn and S-SCAM/Magi2. Unexpectedly, the expression levels of several canonical Wnt target genes were not altered in β-cat cKOs. This lack of change led us to find that β-catenin loss leads to upregulation of γ-catenin (plakoglobin), a partial functional homolog, whose neural-specific role is poorly defined. We show that γ-catenin interacts with several β-catenin binding partners in neurons but is not able to fully substitute for β-catenin loss, likely due to differences in the N-and C-termini between the catenins. Our findings identify severe learning impairments, upregulation of γ-catenin and reductions in synaptic adhesion and scaffold proteins as major consequences of β-catenin loss.

Excessive β-Catenin in Excitatory Neurons Results in Reduced Social and Increased Repetitive Behaviors and Altered Expression of Multiple Genes Linked to Human Autism

“…In contrast, APC/βcat cKOs demonstrated normal social interest (Figures 2A,B) suggesting that the aberrant social behavior of APC cKOs is averted by preventing elevated β-cat in neurons predisposed to excessive β-cat by APC loss. Similarly, β-cat cKOs (low βcat, normal APC levels) show that β-cat down-regulation in the excitatory neurons, using the same CamKIIα-Cre driver did not affect their social behavior, relative to control littermates (Wickham et al, 2019).…”

“…β-cat cOEs bury significantly fewer marbles than controls, phenotype seen in APC cKOs. Similarly, mice with β-cat cKO alone displayed normal behavior in the marble-burying assay (Wickham et al, 2019). Studies of post-mortem brains from autistic patients show that dendritic complexity and spine density are commonly altered in the disease (Hutsler and Zhang, 2010;Tang et al, 2014;Weir et al, 2018).…”

“…APC cKO (APC fl/fl ) mice were generated as previously described (Mohn et al, 2014). β-cat cOE (ctnnb1 fl(ex3)/+ ; Harada et al, 1999), β-cat cKO (ctnnb1 fl/fl ; Wickham et al, 2019), and APC/βcat cKOs (APC fl/fl /ctnnb1 fl/fl ) mice were generated with the identical CamKIIα-Cre-93 recombinase carrying line (Rios et al, 2001). For all experiments, 2-3 month-old mice of both sexes were used.…”

“…We have utilized the CamKIIα Cre driver that is predominantly expressed in forebrain excitatory neurons and fully activated during the first three postnatal weeks in mice (Rios et al, 2001;Pirone et al, 2017) equivalent to the developmental age when glutamatergic neurons exhibit convergent expression of several ASD linked genes in both the human and mouse cortex (Parikshak et al, 2013;Willsey et al, 2013). We have used this CamKIIα-Cre driver to target the same cell types at the same developmental age in all of our mouse lines, including the APC cKO and β-cat cKO lines (Mohn et al, 2014;Wickham et al, 2019).…”

“…(A) Schematic showing the floxed ctnnb1 and adenomatous polyposis coli (APC) genes used to alter β-catenin levels in mice carrying CamKIIα-Cre recombinase. For excessive levels: β-cat cOE-overexpression of stable N-terminally truncated β-cat by deleting the degradation domain, and APC conditional knockout (cKO; Mohn et al, 2014)-deletion of the major negative regulator of β-cat; for reduced levels: APC/β-cat cKO with unstable, rapidly degraded protein products from both genes, and β-cat cKO (Wickham et al, 2019). (B) Immunoblot and quantification of β-cat prefrontal cortex (PFC) levels.…”