The autism susceptibility candidate 2 (AUTS2) gene is associated with multiple neurological diseases, including autism, and has been implicated as an important gene in human-specific evolution. In this thesis, I begin (chapter 1) with an introduction reviewing the literature regarding AUTS2, including its discovery, expression, association with autism and other neurological and non-neurological traits, implication in human evolution, function, regulation, and genetic pathways. Part of the research included in this introduction was performed by me and co-authors, and described in detail in the following chapters.
In chapter 2, I investigate the expression, function, and regulation of auts2. auts2 is expressed primarily in the zebrafish brain. Knockdown of this gene in zebrafish leads to a smaller head size, neuronal reduction and decreased mobility. I identified twenty-three functional zebrafish enhancers, ten of which are active in the brain. Mouse enhancer assays characterized three brain enhancers that overlap an ASD-associated deletion and four enhancers that reside in regions implicated in human evolution, two of which are active in the brain. Combined, I show that AUTS2 is important for neurodevelopment and expose candidate enhancer sequences in which nucleotide variation could lead to neurological disease and human-specific traits.
In chapter 3, I investigate the regulatory role and targets of Auts2. Using ChIP-seq and RNA-seq on mouse embryonic day 16.5 forebrains, we I elucidated the gene regulatory networks of Auts2. It was found that the majority of promoters bound by Auts2 belong to genes highly expressed in the developing forebrain, suggesting that Auts2 is involved in transcriptional activation. Auts2 non-promoter bound regions significantly overlap developing brain-associated enhancer marks and are located near genes involved in neurodevelopment. Auts2 marked sequences are enriched for binding site motifs of neurodevelopmental transcription factors, including Pitx3 and TCF3. I characterized ten non-coding Auts2 marked sites near critical ASD-related genes for enhancer activity in zebrafish, four of which showed positive enhancer activity. Additionally, I characterized two of the positive brain enhancers near NRXN1 and ATP2B2 in mice. The results implicate Auts2 as an active regulator of important neurodevelopmental genes and pathways and identify novel genomic regions which could be associated with ASD and other neurodevelopmental diseases. In summary, this thesis investigates the function of AUTS2. I conclude that this gene is critical for the proper development of neurons, and may act as a cofactor to positively regulate genes expressed in the forebrain involved in neurodevelopment.