This project will focus on the investigation of “microtubule” genes. First, we will focus on Pcm1, Dpysl2, and Kctd13 (a key pathological driver of the phenotypes associated with the human chromosome 16p11.2 duplication) in mouse models.
The initial validation of these genetically engineered mice will be performed at the anatomical/histological levels in early development for further characterization in Projects 2 and 3. In addition, mouse genetics experiments will be conducted to further address the role of Kctd13, and examine the temporal requirement of these genes in adult phenotypes relevant to schizophrenia (SZ).
Second, human cell models [glutamatergic neurons induced from human induced pluripotent stem (iPS) cells] will be used to test how genetic variations in “microtubule” genes (KCTD13, DPYSL2, SDCCAG8, and CKAP5) associated with SZ affect cellular signaling. In addition, we will test how additional stressors to cells exacerbate the changes. RNA-seq will be used to obtain overall expression pattern changes, and the data will be compared with those from the frontal cortex of the mice with corresponding genetic mutations.
The RNA-seq data will also be compared with those from human postmortem brains.
We expect that genes and pathways found to be altered in cultured induced neurons in response to modifications in the “microtubule” genes, as well as to dexamethasone treatment, will include those that are active specifically during development and/or change during adolescence.
Lastly, using human genetics and zebrafish biology, we will identify additional “microtubule” genes that are associated with SZ, in particular treatment-refractory
cases that may have stronger associations with neurodevelopmental deficits.
This project will include two complementary components (genetic models from Project 1, and human biospecimens, including blood from two independent cohorts). First, by using mouse models, we will examine whether microtubule deficits precipitate stress-related molecular and brain alterations in adolescence, which lead to cognitive impairment relevant to SZ in young adulthood. This candidate target approach will be followed by assessment of unbiased molecular expression using RNA-seq. In particular, we will study how adolescent stressors exacerbate (or make detectable from a sub-threshold level) the abnormalities conferred by the genetic risk factors (disturbance of “microtubule” genes) during early development. These data will form the basis for the
functional study in Project 3.
The RNA-seq data will also be compared to those from postmortem adolescent brains, brain sets from patients and controls, and human cell models in collaboration with
Project 1, and Cores A and B.
Through these analyses, we will examine how developmentally regulated transcripts during adolescence are differentially expressed in the mouse models; will study how differentially expressed molecules in the models are represented in different disease conditions; and will determine the molecular pathways impacted by cell autonomous vs. cell non-autonomous effects of the genetic variants (in comparison with the cell data obtained by Project 1).
Lastly, this project will also study the complement system and other stress-associated molecules in blood from two independent prospective human cohorts, and compare
them to those in the mouse models.
This project will use mouse models that show SZ-relevant molecular and neurocircuitry changes, as well as behavioral deficits in adulthood, either by themselves or in combination with social isolation, following characterization in Projects 1 and 2. We will determine “critical period” when stress-associated molecules are activated, and when neurocircuitry changes become prominent. The neurocircuitry changes in the cortex will be validated mainly by slice electrophysiology.
We hypothesize that E-I imbalance in the cortex emerges during adolescence due to activation of stress-associated molecules in mice that have mild developmental brain deficits from microtubule-associated genetic risks, and are exacerbated by adolescent stressors.
Lastly, we will address the causality of these stress pathways in neural and behavioral phenotypes via general or brain region-specific genetic manipulations as well as pharmacological interventions. Effects of these interventions will be tested with Core C, by using working memory tasks corresponding to medial PFC (mPFC), and behavioral
flexibility tests corresponding to orbitofrontal cortex (OFC).
This Core will serve as the central administration and scientific and ethical leadership, organizing all the human tissue and genetic resources, and integrating data for the entire database. This core will also be responsible for education and public outreach activities of this Center.
This Core leads the analysis of RNA-seq using human cells and animal models. This core will compare these data with RNA-seq data from postmortem adolescent brains (Lieber institute), as well as from established brain sets from patients and controls (Stanley Division), and will lead the discussion on data interpretation with Projects 1-3.
This Core leads the studies of animal behavior. This core will help the project investigators (Projects 1-3) to conduct basic behavioral analyses as an initial screening. This core will mainly conduct hypothesis-driven, circuitry-oriented behavioral assays that are associated with PFC, such as behavioral flexibility and working memory.