A Neuron-Specific Histone Demethylase Complex Fine Tunes Neurodevelopment

Abstract

Cell-type specific transcriptional programs are thought to be defined by cell-type specific transcription factors (TFs) that bind to specific DNA sequences. In contrast, chromatin regulators are ubiquitously expressed and exert the same intrinsic activity but rely on cell-type specific TFs to engage specific genomic loci. However, mutations in chromatin regulators have emerged as a major driver of neurodevelopmental disorders, such as autism spectrum disorder and intellectual disability, challenging the ubiquitous action of chromatin regulators. Why does the disruption of widely-expressed chromatin factors lead to cognitive dysfunction?

This thesis work addresses this question through the investigation of neuron-specific isoforms of chromatin regulators. By surveying literature and publically available data, we identified 76 chromatin regulators that undergo neuron-specific microexon splicing, which led us to hypothesize that neuron-specific chromatin regulators themselves can exert unique intrinsic activity to establish the neuron chromatin landscape. We characterized one specific human disease example through RNA-Seq and follow-up validation how haploinsufficiency of PHF21A in Potocki Shaffer Syndrome patient-derived cells leads to a deficiency in the transcriptional response to stimulus.

To test this hypothesis, we investigated a neuron-specific histone demethylase complex where the histone H3K4 demethylase LSD1 and the accompanying histone reader PHF21A both undergo neuron-specific microexon inclusion. Interestingly, neuronal microexon inclusion in LSD1 and PHF21A alters enzymatic and nucleosome-binding domains, respectively. We expressed and reconstituted neuronal and canonical LSD1/PHF21A/CoREST complexes and found that the neuronal complex exhibits reduced binding to nucleosomes and reduced H3K4me demethylation capability. To test the impact of PHF21A in neurodevelopment, we performed RNA- and ChIP-Seq as well as confocal microscopy to evaluate synapse formation in Phf21a knockout and rescue mouse models. Phf21a-KO mice have a defect in synapse number and accompanying transcriptomic changes in neuronal genes. However, animals with aberrant expression of the canonical-PHF21A isoform in neurons show an abnormally elevated synapse number. Our results demonstrate that microexon inclusion in the LSD1/PHF21A complex leads to a biochemical dampening of complex function impacting the neuronal histone methylome and thereby fine-tunes gene expression for proper synapse formation. This work illuminates how chromatin regulators can have neuron-specific forms with distinct activity to shape the neuronal chromatin landscape.