Gene Dosage- and Age-Dependent Differential Transcriptomic Changes in the Prefrontal Cortex of Shank2-Mutant Mice

Abstract

Shank2 is an abundant postsynaptic scaffolding protein that is known to regulate excitatory synapse assembly and synaptic transmission and has been implicated in various neurodevelopmental disorders, including autism spectrum disorders (ASD). Previous studies on Shank2-mutant mice provided mechanistic insights into their autistic-like phenotypes, but it remains unclear how transcriptomic patterns are changed in brain regions of the mutant mice in age- and gene dosage-dependent manners. To this end, we performed RNA-Seq analyses of the transcripts from the prefrontal cortex (PFC) of heterozygous and homozygous Shank2-mutant mice lacking exons 6 and 7 at juvenile (week 3) and adult (week 12) stages. Juvenile heterozygous Shank2-mutant mice showed upregulation of glutamate synapse-related genes, downregulation of ribosomal and mitochondrial genes, and transcriptomic changes that are opposite to those observed in ASD (anti-ASD) such as upregulation of ASD_down (downregulated in ASD), GABA neuron-related, and oligodendrocyte-related genes. Juvenile homozygous Shank2 mice showed upregulation of chromatin-related genes and transcriptomic changes that are in line with those occurring in ASD (pro-ASD) such as downregulation of ASD_down, GABA neuron-related, and oligodendrocyte-related genes. Adult heterozygous and homozygous Shank2-mutant mice both exhibited downregulation of ribosomal and mitochondrial genes and pro-ASD transcriptomic changes. Therefore, the gene dosage- and age-dependent effects of Shank2 deletions in mice include differential transcriptomic changes across distinct functional contexts, including synapses, chromatin, ribosomes, mitochondria, GABA neurons, and oligodendrocytes.

Age, brain region, and gene dosage-differential transcriptomic changes in Shank3-mutant mice

“…These results suggest that these mice undergo a strong agedependent transcriptomic change from a reverse-ASD pattern to an ASD-like pattern through the alteration of synaptic gene expression. A similar age-dependent change from a reverse-ASD to ASD-like pattern was previously reported in the mPFC region of Shank2-HM mice (exons 6-7), although the altered biological functions and ASD-related/risk genes were different (Lee et al, 2021). Despite this difference, the results from Shank2 and Shank3 mice collectively indicate at the minimum age-dependent transcriptomic inversion with respect to ASD-related/risk gene expressions.…”

“…In addition, we analyzed transcriptomic patterns in the prefrontal cortex (termed cortex hereafter), hippocampus, and striatum regions of adult (∼postnatal day 90 or P90) Shank3 heterozygous (HT)- and homozygous (HM)-mutant mice lacking exons 14–16 (Shank3-HT/HM mice). We also compared these results with those previously reported from Shank2 -mutant mice ( Lee et al, 2021 ; Yoo et al, 2022 ). Our findings collectively indicate that there are age, brain region, and gene dosage-differential transcriptomes within and between Shank2 - and Shank3 -mutant mice, which may provide insight into altered biological functions and ASD-related/risk gene expression patterns.…”

Brain region and gene dosage-differential transcriptomic changes in Shank2-mutant mice

“…In addition, we previously performed transcriptomic analyses of the medial prefrontal brain regions from heterozygous and homozygous Shank2-mutant mice lacking exons 6 and 7. We observed stronger reverse-ASD-like transcriptomic changes in heterozygous mutant mice than in homozygous mutant mice, which were more prominent at juvenile stages relative to adult stages (Lee et al, 2021b). This led us to question whether such changes are conserved across a larger prefrontal area and/or in other brain regions.…”

“…When the current GSEA results for Shank2-HT and Shank2-HM cortical/prefrontal transcripts, covering wider areas of the prefrontal cortex including more caudal regions, are compared with the previous results for Shank2-HT and Shank2-HM mPFC transcripts (Lee et al, 2021b) for ASD-related/risk patterns, the reverse-ASD pattern of the current Shank2-HT (but not Shank2-HM showing ASD-like pattern) prefrontal transcripts are more similar to the previous results (reverse-ASD for both Shank2-HT and Shank2-HM mPFC transcripts). It might be possible that the mPFC region may be more resilient in inducing compensatory and reverse-ASD transcriptomic changes even in the presence of a strong HM Shank2 deletion.…”

“…Shank2 has been implicated in various brain disorders, including ASD, intellectual disability, developmental delay, attention deficit/hyperactivity disorder, schizophrenia, epilepsy, and obsessive compulsive disorder ( Berkel et al, 2010 , 2011 ; Pinto et al, 2010 ; Leblond et al, 2012 , 2014 ; Rauch et al, 2012 ; Sanders et al, 2012 ; Chilian et al, 2013 ; Liu et al, 2013 ; Guilmatre et al, 2014 ; Costas, 2015 ; Peykov et al, 2015a , b ; Homann et al, 2016 ; Bowling et al, 2017 ; Mossa et al, 2017 ; Yuen et al, 2017 ; Bai et al, 2018 ; Guo et al, 2018 ; Lu et al, 2018 ; Callaghan et al, 2019 ; Satterstrom et al, 2020 ; Trost et al, 2020 ; Wang et al, 2020 ; Ma et al, 2021 ; Ohashi et al, 2021 ; Chenbhanich and So, 2022 ). Previous studies in mouse/rat models of ASD and human neurons deficient of Shank2 have suggested various molecular, synaptic, neuronal, and circuit mechanisms underlying Shank2-related phenotypes ( Schmeisser et al, 2012 ; Won et al, 2012 ; Ey et al, 2013 , 2018 ; Lee et al, 2015b , 2018 , 2020 , 2021a , b ; Ferhat et al, 2016 ; Ha et al, 2016 ; Ko et al, 2016 ; Peter et al, 2016 ; Lim et al, 2017 ; Pappas et al, 2017 ; Sato et al, 2017 , 2020 ; Yoon et al, 2017 ; Heise et al, 2018 ; Kim et al, 2018 ; Modi et al, 2018 ; Wegener et al, 2018 ; Chung et al, 2019 ; de Chaumont et al, 2019 ; Zaslavsky et al, 2019 ; Chen et al, 2020 ; Han et al, 2020 ; Daini et al, 2021 ; Gra...…”