The development of cortico-cerebellar circuits in a genetic form of intellectual disability

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PROJECT SUMMARY Intellectual disability (ID) is a prevalent neurodevelopmental disorder with no effective pharmacological treatment. ID is often co-morbid with motor problems, and the nature of shared risk between these clinical manifestations is not fully understood. Amongst the ID high-risk genes with significant motor involvement is the X-linked gene DDX3X. Mutations in DDX3X affect almost exclusively females. DDX3X encodes an RNA helicase regulating mRNA translation, and its role in neurodevelopment is just beginning to emerge. The circuit-level changes induced by DDX3X mutations are unknown. There is a critical need to fill these gaps because understanding the role of DDX3X in the formation of brain circuits might offer a new key to decipher ID and its co-morbidity with motor problems. To address this unmet need, a mouse modeling DDX3X loss-of- function mutations (Ddx3x+/-) was generated and characterized in our laboratory. The long-term goal is to map the circuit-level drivers of ID and assess them as therapeutic targets. The overall objective is to examine the role of DDX3X in shaping cortico-cerebellar circuits contributing to cognition and motor control. The central hypothesis is that DDX3X regulates the development of cortico-cerebellar circuits subserving cognitive and motor function. The rationale is that, once we understand the circuit-level drivers of ID, mechanism-based precision therapeutics can be developed. The central hypothesis will be tested by pursuing two Specific Aims: 1) Identify the neural populations altered in the cerebellum of Ddx3x mutant mice; and, 2) Test the role of cortico-cerebellar communication in behavioral deficits of Ddx3x mutant mice. Under Aim 1, the alterations in cerebellar populations of Ddx3x+/- mice will be captured using single-cell RNA sequencing. The architecture of the cerebellar cortex in Ddx3x+/- mice will be analyzed using immunostaining of cerebellar populations with authenticated cell-specific markers during embryonic and postnatal life. The morphogenesis and synaptogenesis of Ddx3x+/- Purkinje cells will be examined using single-embryo primary cultures. Under Aim 2, the cortico-ponto-cerebellar pathway will be analyzed in Ddx3x+/- mice by simultaneously tracing cortico-pons and ponto-cerebellar connections. Purkinje-specific Ddx3x+/- conditional mice will be generated and tested for cognition and motor behavior using well-established behavioral paradigms. This proposal is innovative because it will define the neurobiology of a largely unknown ID gene by mapping the cerebellar cells and circuits affected by DDX3X mutations, and their relationship with behavior. It is also innovative because it bridges developmental biology, cellular biology, behavioral neuroscience, and single-cell transcriptomics. The application is significant because it will advance our understanding of ID and co-morbid motor deficits, and look specifically at females, which have been neglected. It is also significant because it will shed new light on cerebellar development, a fundamental process for brain function. These results are expected to have a positive impact because they will pave the way for better methods for prevention and treatment of ID.