Exosomal protein deficiencies: how abnormal RNA metabolism results in childhood-onset neurological diseases

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All genes are copied into short-lived RNA molecules, which are then translated to protein, forming the building box of the cells in the body. The transcription of DNA, the processing of pre-mRNA into mature mRNA (splicing), the degradation of mRNA and the translation into proteins are all tightly regulated. Both too much and too little of a certain RNA species could be dangerous for a cell, but our understanding of the mechanisms to fine-tune the RNA amounts is still limited. The regulation of gene expression (RNA metabolism) is of utmost importance for normal cell function, nerve cells, however, seem less able to cope with errors. This is illustrated by an increasing number of severe inherited neurological diseases of infancy or childhood caused by a defect in the RNA production machinery; these disorders are characterized by abnormalities of the development and/or the structure of the brain and the central nervous system. The aim of this proposal is to study and characterize a novel form of RNA related neurological disorder due to dysfunction of a multi-protein complex called the human exosome. The role of the exosome consists of degradation and maturation of RNA. Mutations in the exosome subunit gene EXOSC3 were reported in patients with infantile-onset degeneration of the brainstem and cerebellum and muscle weakness due to spinal motor neuron dysfunction. We have recently identified mutations in EXOSC8, encoding another component of the human exosome, in infants with a similar severe neurological disease. In addition, these children also lose myelin in the brain and spinal cord, which normally coats and insulates nerves and serves to speed up signalling. We also identified a homozygous mutation in a novel gene RBM7 in a child with severe spinal motor neuron dysfunction. RBM7 is an interacting partner of the human exosome. The severe and isolated neurological symptoms in these children raise the possibility that the exosome is particularly important in neurons; however, there are still many open questions. The aim of this proposal is to further characterize the role of the exosome in nerve cells and will address the following aspects:1. Neuronal cells from patients: We will analyse RNA levels in patient derived skin cells and will transform them into neuronal cells with recently published methods. We will determine which RNA species are modified by changes in the exosome in the different cell types. 2. Genetically modified zebrafish: In parallel, we will create zebrafish models of exosomal protein deficiencies. By artificially removing single exosome components and by introducing mutations in zebrafish embryos we will explore its effect on the development and function of the brain and different types of neurons. In addition, we will also study the effect of modifying the exosome on myelin formation in zebrafish. Our results will identify the most important roles of the exosome in regulating gene expression and why is it more damaging for neurons. Combining the data in human cells and zebrafish will enable us to better define the important genes and interacting partners of the exosome. By modifying exosome components or some of the here identified interacting factors we may discover potential pathways to alter RNA degradation or processing in neurons, which can be further developed as a therapeutic approach in exosomal diseases or in other types of neurodegenerative diseases caused by abnormal RNA function.