Rizzardi, L. | Search bioRxiv

bioRxiv2026

Decoupled calcium homeostasis and signaling associated with cytoskeletal instability in YWHAG R132C induced pluripotent stem cell-derived cortical neurons

YWHAG Syndrome (Developmental and Epileptic Encephalopathy 56, DEE56) is an ultra- rare childhood epilepsy associated with neurodevelopmental delays, with no therapeutic intervention available. Multiple de novo mutations in the YWHAG gene, encoding for the 14-3-3{gamma} protein, have been identified as causative for YWHAG Syndrome. 14-3-3{gamma} interacts with various targets, including major neurodevelopmental signaling proteins such as components of the ROCK pathway. Despite substantial evidence of the essential role of 14-3-3{gamma} in neurite outgrowth, cytoskeletal rearrangements, and neuronal migration during cortical development, little is known regarding the molecular consequences of YWHAG mutations and their effect on neuronal function and survival. Here, we characterized an isogenic, pluripotent stem cell (iPSC) model of YWHAGR132C/+ cortical neurons. The YWHAGR132C/+iPSC-derived neurons exhibited early cytoskeletal phenotypes, coupled with an elevated calcium baseline, lower frequency of calcium spikes, and reduced network activity. The widespread alterations in the transcriptome of mutant neurons revealed a biphasic dysregulation in the core genes and modulators associated with the ROCK pathway that resulted in maturation-dependent changes to cytoskeletal protein stability and calcium phenotypes. Direct inhibition of ROCK with Y27632 further increased the calcium baseline compared to the isogenic control. Exposure of YWHAGR132C/+ neurons to Trypsin-EDTA revealed underlying cytoskeletal instability, which was partially reversed by lovastatin treatment. Further, lovastatin partially rescued the elevated calcium baseline, but not the frequency or amplitude of calcium spikes. Together, these results suggest decoupling of calcium homeostasis and calcium signaling associated with cytoskeletal instability in YWHAGR132C/+ neurons. These findings lay the groundwork for future mechanistic studies of YWHAG function and molecular therapeutic targets for YWHAG Syndrome and YWHAG-associated conditions.

neuroscience↗

bioRxiv2017

Neuronal brain region-specific DNA methylation andchromatin accessibility are associated with neuropsychiatric disease heritability

Epigenetic modifications confer stable transcriptional patterns in the brain, and both normal and abnormal brain function involve specialized brain regions, yet little is known about brain region-specific epigenetic differences. Here, we compared prefrontal cortex, anterior cingulate gyrus, hippocampus and nucleus accumbens from 6 individuals, performing whole genome bisulfite sequencing for DNA methylation. In addition, we have performed ATAC-seq for chromatin accessibility, and RNA-seq for gene expression in the nucleus accumbens and prefrontal cortex from 6 additional individuals. We found substantial neuron- and brain region-specific differences in both DNA methylation and chromatin accessibility which were largely non-overlapping, and were greatest between nucleus accumbens and the other regions. In contrast, glial methylation and chromatin were relatively homogeneous across brain regions, although neuron/glia ratios varied greatly, demonstrating the necessity for cellular fractionation. Gene expression was also largely the same across glia from different brain regions and substantially different for neurons. Expression was correlated with methylation and accessibility across promoters and known enhancers. Several classes of transcription factor binding sites were enriched at regions of differential methylation and accessibility, including many that respond to synaptic activity. Finally, both regions of differential methylation and those of differential accessibility showed a surprising >10-fold enrichment of explained heritability associated with addictive behavior, as well as schizophrenia- and neuroticism-associated regions, suggesting that common psychiatric illness is mediated through brain region-specific epigenetic marks.

neuroscience↗