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Molecular and behavioral consequences of Ube3a gene overdosage in mice
A. Mattijs Punt, … , Ype Elgersma, Benjamin D. Philpot
A. Mattijs Punt, … , Ype Elgersma, Benjamin D. Philpot
Published September 22, 2022
Citation Information: JCI Insight. 2022;7(18):e158953. https://doi.org/10.1172/jci.insight.158953.
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Research Article Neuroscience

Molecular and behavioral consequences of Ube3a gene overdosage in mice

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Abstract

Chromosome 15q11.2–q13.1 duplication syndrome (Dup15q syndrome) is a severe neurodevelopmental disorder characterized by intellectual disability, impaired motor coordination, and autism spectrum disorder. Chromosomal multiplication of the UBE3A gene is presumed to be the primary driver of Dup15q pathophysiology, given that UBE3A exhibits maternal monoallelic expression in neurons and that maternal duplications typically yield far more severe neurodevelopmental outcomes than paternal duplications. However, studies into the pathogenic effects of UBE3A overexpression in mice have yielded conflicting results. Here, we investigated the neurodevelopmental impact of Ube3a gene overdosage using bacterial artificial chromosome–based transgenic mouse models (Ube3aOE) that recapitulate the increases in Ube3a copy number most often observed in Dup15q. In contrast to previously published Ube3a overexpression models, Ube3aOE mice were indistinguishable from wild-type controls on a number of molecular and behavioral measures, despite suffering increased mortality when challenged with seizures, a phenotype reminiscent of sudden unexpected death in epilepsy. Collectively, our data support a model wherein pathogenic synergy between UBE3A and other overexpressed 15q11.2–q13.1 genes is required for full penetrance of Dup15q syndrome phenotypes.

Authors

A. Mattijs Punt, Matthew C. Judson, Michael S. Sidorov, Brittany N. Williams, Naomi S. Johnson, Sabine Belder, Dion den Hertog, Courtney R. Davis, Maximillian S. Feygin, Patrick F. Lang, Mehrnoush Aghadavoud Jolfaei, Patrick J. Curran, Wilfred F.J. van IJcken, Ype Elgersma, Benjamin D. Philpot

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Figure 1

Generation and validation of Ube3aOE mice with dose-dependent overexpression of Ube3a transcript and UBE3A protein.

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Generation and validation of Ube3aOE mice with dose-dependent overexpres...
(A) Schematic of BAC transgenic strategy to generate conditional Ube3a overexpression mice. Exon/intron numbering is relative to Ube3a isoform 2. (B) Droplet digital PCR (ddPCR) analysis of total and transgenic Ube3a genomic copy number in Ube3a+2 overexpression mice from line E. ddPCR assays specific to the Ube3a intron 3/exon 3 boundary were used to assess total (i.e., endogenous plus transgenic) Ube3a copy number (left); ddPCR amplification of loxP sequences enabled specific detection of transgenic Ube3a copies (right). Homozygous floxed Ube3a-knockin mice (Ube3afl/fl) served as controls. (C) Representative Western blot (WB) depicting immunofluorescent detection of bands corresponding to total whole-brain UBE3A and the loading control protein, GAPDH. Bands at left (red) correspond to the molecular weight marker. (D) Mean ± SEM UBE3A WB immunofluorescence intensities as determined from WT (n = 7), AS (n = 7), and Ube3a+1 (n = 14), Ube3a+2 (n = 14), and Ube3a+4 (n = 14) whole-brain lysates. Welch’s ANOVA, Dunnett’s post hoc. (E) Mean ± SEM whole-brain Ube3a transcript levels as determined from ddPCR experiments (2.5 ng cDNA input). Samples were prepared from cerebral hemispheres opposite those used to prepare the protein lysates assayed in D. One-way ANOVA, Tukey’s post hoc. (F) Box plots of ratios of whole-brain Ube3a transcript and UBE3A protein levels for corresponding samples from E and D, respectively. Whiskers represent 5% to 95% confidence intervals. One-way ANOVA. (G) Mean ± SEM ratios of Ube3a isoform 3 to Ube3a isoform 2 transcript levels, broken out by specific Ube3aOE lines, as determined from ddPCR experiments. One-way ANOVA. **P < 0.01, ****P < 0.0001.

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