Characterization of schizophrenia liability genes in models of human microglial synaptic pruning

点击翻译按钮获取中文摘要

Schizophrenia is a chronic, disabling, and strongly heritable illness. Existing treatments do not restore function for many patients, nor do they modify the disease process, and development of novel therapeutics is hindered by a lack of biological targets. Postmortem studies demonstrate reduced cortical dendritic spine density among individuals with schizophrenia, consistent with clinical structural neuroimaging studies. Convergent lines of evidence from rodent models and human genomics suggest that these abnormalities may arise from microgliamediated pruning dysfunction. The investigators have developed, validated and published patient-specific models of microglia-mediated pruning by generating reprogrammed microglia-like cells from patients and healthy controls, and assaying them with isolated and highly purified synapses (synaptosomes) from induced pluripotent stem cell (iPSC)-derived neuronal cultures. In their published studies, they have demonstrated robust evidence of dysfunction in synaptic engulfment attributable to abnormalities in both microglia and neurons from individuals with schizophrenia. To date, efforts to link these abnormalities to disease risk loci have largely been limited to the complement C4 locus, and focused almost entirely on neurons. The proposed investigation will draw on the patient-derived cellular biobank created by the investigators to apply genetic engineering methods to address the role of microglia in synaptic pruning directly. Specifically, it will investigate the contributions on synaptic engulfment of both microglial expressed schizophrenia liability genes and putative upstream regulators of neuronal complement synaptic expression through engineered induced microglia and neuronal cultures, respectively. For Aim 1, the investigators propose to perform CRISPR screening applied to induced microglia-like cells, applying high scale in vitro synaptic pruning assays to discover novel disease-associated gene functions implicated in schizophrenia. In Aim 2, the investigators will further validate loci from the screen in Aim 1 in CRISPR-engineered iPSC-derived microglia models to confirm roles in synaptic engulfment and determine other effects on microglia development and function. To complement these aims, in Aim 3, the investigators will generate engineered iPSC-derived neuronal cultures manipulating the expression of human complement C4 alleles and modulatory genes. They will then use synaptosomes from these lines for functional in-vitro microglialmediated engulfment assays to assess effects of manipulation of these genes in neurons. Taken together, these aims will characterize the effects of schizophrenia liability loci on human synaptic pruning, in terms of microglia function as well as neuronal complement expression, using validated, scalable patient-derived model systems. They will prioritize targets for treatment development and inform the biology of schizophrenia risk.