Major psychiatric disorders, such as schizophrenia (SZ) and major depression, involve abnormal social behaviors and onset typically occurs after puberty. The genetic risk factors for SZ are frequently heterozygous in affected individuals and the clinical phenotypes can be highly variable. However, the factors that contribute to phenotypic variance and the timing of the onset of these disorders are poorly understood. Genomic imprinting is an epigenetic mechanism that causes preferential expression of the maternal or paternal allele for some genes. Imprinting causes parent-of-origin effects that influence the phenotypic effects of inherited mutations. Canonical imprinting involves complete silencing of one parent's allele and impacts a small number of genes. However, in a recent publication, the author's lab discovered that hundreds of genes in the mouse that exhibit a maternal or paternal allele expression bias. This phenomenon is called noncanonical imprinting. Noncanonical imprinting is highly enriched in the brain compared to the liver or muscle. Using a novel allele- specific in situ hybridization approach, the author found that noncanonical imprinted genes exhibit monoallelic expression in subpopulations of cells in the brain, suggesting a highly cell-type specific form of imprinting in the brain. Currently, it is not known whether noncanonical imprinting is a cell-type specific form of imprinting in the brain. Further, the field does not know the identity of the genes that are imprinted in specific cell-types of the brain, whether noncanonical imprinting can change at puberty or whether social behaviors are impacted. The author found that tyrosine hydroxylase (Th) and dopa decarboxylase (Ddc) are noncanonical imprinted genes with a maternal allele bias in specific regions of the brain. TH and DDC synthesize monoamine neurotransmitters, which have important roles in social behavior and mental illness. Here, the author will test the hypothesis that noncanonical imprinting effects in monoaminergic circuits are a highly cell-type specific form of imprinting that can change in response to pubertal development and impact social behaviors with roles in mental illness.
Aim 1 (K99) will determine whether noncanonical imprinting is a cell- type specific form of imprinting in the monoamine system and uncover the identity of the genes that are imprinted in molecularly-defined subtypes of monoaminergic neurons in the brain. These studies involve generating maternal and paternal allele-specific reporter mice to resolve imprinting at the cellular level for Ddc and new methods to profile imprinting at the cellular level in isolated monoaminergic neurons from different brain regions using single-cell RNASeq.
Aim 2 (R00) will determine whether noncanonical imprinting in the preoptic area (POA) and arcuate nucleus (ARN) changes at puberty by profiling imprinting before and after puberty using RNASeq methods developed in our lab. The POA and ARN play important roles in puberty and social behaviors and preliminary data shows that noncanonical imprinting can change in these regions in response to hormonal signaling.
In Aim 3 (R00), Th and Ddc single and compound mutant mice will be used to determine whether noncanonical imprinting effects impacting genes in the same pathway function synergistically to amplify parental influences on offspring social behaviors. Social behaviors that are impacted in mental illness will be tested, including aggression, social preference, social recognition, social reward and reproductive behaviors. These experiments will provide essential new training in mouse genome engineering and genomics (K99) and establish the foundations for an independent research program (R00) focused on the roles of noncanonical imprinted genes and hormone signaling in social behaviors and mental health. The K99 portion of this proposal will provide the principle investigator (PI) with research training in state-of-the art CRISPR-Cas9 mediated mouse genome engineering, programming for bioinformatics and genomic analysis, and single cell transcriptomics. Career development to independence will consist of regular faculty mentor, co-mentor, and faculty committee meetings to ensure that research and career benchmarks are being met in a timely and appropriate fashion. The mentorship team will aid in the application and negotiation process for faculty positions, the preparation of an independent research proposal and job talks, and putting together the Aims and Research Strategy for the PI's first R01 grant application before leaving the University of Utah. The job talks and R01 proposals will also be presented to the ?Faculty Think Tank? to get critical feedback from the whole department. The principle investigator will also improve communication skills by presenting research in progress talks to the department and relevant journal clubs, attending scientific meetings where the PI will have an opportunity to give oral presentations, and attending a grantsmanship course offered at the University of Utah. Combined with the foundation of the independent research program, the career development activities will ensure that the PI is well prepared to enter an independent faculty position at a major research oriented university and emerge as a leader in the fields of imprinting, social behavior and mental health.
This training program and study will advance our understanding of the expression of maternally and paternally inherited alleles in the brain and in neurons with important roles in social behavior and mental health. The proposed research is relevant to public health because it could lead to improvements in our ability to evaluate genetic risk factors for mental illness according to whether mutations are maternally or paternally derived.
|Jacobi, Ashley M; Rettig, Garrett R; Turk, Rolf et al. (2017) Simplified CRISPR tools for efficient genome editing and streamlined protocols for their delivery into mammalian cells and mouse zygotes. Methods 121-122:16-28|