Molecular and Cellular Phenotypes of AUTS2 Deficiency in the Developing Mouse Cortex and Hippocampus

Thesis (Ph.D.)--University of Washington, 2018Variants in the gene Autism Susceptibility Candidate 2 (AUTS2) are a rare cause of intellectual disability, developmental delay and autistic features in patients. However, the mechanism by which loss of AUTS2 causes specific neurological symptoms, the underlying neuroanatomical abnormalities, and the syndrome’s pathological progression are not well established. The core molecular function of this intellectual disability syndrome gene also remains uncertain. Previous studies have proposed two independent mechanisms of AUTS2 function. One found that Auts2 modifies the Polycomb repressive complex 1 (PRC1) to activate transcription of target neurodevelopmental genes by recruiting the histone acetyltransferase EP300. A second study found that cytoplasmic AUTS2 regulates cytoskeletal assembly through PREX1 to promote neurite outgrowth and migration. To further investigate Auts2 functions, we screened for AUTS2 interactors in the cerebral cortex and profiled changes of transcript expression in the developing cortex of Auts2 conditional mutant mice. AUTS2 immunoprecipitation experiments identified interactions with proteins involved in pre-mRNA processing including the multifunctional RNA-processing regulator NONO. RNA-immunoprecipitation experiments identified enrichment of specific transcripts associated with AUTS2 including mRNAs which encode the previously identified effectors of Auts2 function EP300 and PREX1. Additional preliminary experiments identified a network of transcripts encoding a Notch signaling related protein-protein interaction network which were co-bound by AUTS2 and NONO. We propose that AUTS2 interacts with an RNA-binding complex that directly targets transcripts encoding Auts2 effectors. This mechanism provides a unifying model that accounts for previously disparate hypotheses of AUTS2 functions. Prior work developed mouse models of Auts2 deficiency, however, these models did not fully account for additional, functional, isoforms expressed from the c-terminus of the locus. Our work developed a mouse model which is expected to ablate all c-terminal containing AUTS2 isoforms under the control of CRE-recombinase (Auts2 cKO). RNA-sequencing of frontal cortex and hippocampus from P0.5 Auts2 cKO mice identified dysregulation of RNAs which encode proteins previously identified to be effectors of AUTS2 and which were targeted by the AUTS2 complex. We additionally identified dysregulated pathways encoding developmental process including cytoskeletal organization, cell migration, adhesion, core transcriptional and metabolic processes. These pathways provide a molecular link between AUTS2 and NONO deficiency and overlap with pathways dysregulated in a large study of human neurological disorder transcriptomes. Histological examination of the AUTS2 deficient brain identified severe abnormalities in early post-natal hippocampal development. Dentate gyrus volume was reduced and early colonization of the adult neurogenic niche by TBR2+ intermediate neuronal progenitors was dramatically impaired. These hippocampi also exhibited near total absence of hilar mossy neurons, a population which regulates hippocampal interconnectivity and excitability. EEG analysis of recordings from these animals identified significant abnormalities consistent with disinhibition identified in both autism and epilepsy. The work presented in this dissertation significantly expands on our understanding of the neurobiology regulated by AUTS2. We also identify functional and molecular connections with ASD and epilepsy pathology. These experiments identify mechanisms by which the pathogenesis of the AUTS2 syndrome can be further investigated