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Title: Understanding the role of Caspr2 in the developing cerebellum
Author: Argent, Liam
ISNI:       0000 0004 7654 5533
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 2019
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Despite receiving renewed attention due to the revelation it is involved in higher cognitive functions, we still know little about how the cerebellum develops and operates. Studies suggest Contactin-associated protein-like 2 (Caspr2), a member of the Neurexin superfamily, has a cerebellar function. However, the cellular and molecular bases for this remain unknown. An initial assessment of the spatio-temporal expression pattern of Caspr2, using immunohistochemistry and biochemical fractionation, suggested that Caspr2 is expressed at specific synapses in the developing cerebellum. Little is known about the function of Caspr2 at synapses and so the Caspr2 domain exposed to the synaptic cleft, the Caspr2 extra-cellular domain (ECD), was focused on. The addition of recombinant Caspr2 ECD to organotypic cerebellar slice cultures prepared from Cntnap2-/- mice rescued a dendritic morphology defect observed in untreated cultures, suggesting Caspr2 affects cellular morphology during cerebellar development. To understand the molecular basis for this effect, a pull down assay was used to identify the synaptic binding partners of Caspr2. Unexpectedly, mass spectrometry analysis of this experiment revealed that the Ca2+ channel Inositol 3,4,5- trisphosphate receptor type I (IP3R1) is a cognate partner of Caspr2 at synapses in the developing cerebellum. Further biochemical assays revealed both that the Caspr2-IP3R1 interaction underlies the regulation of cellular morphology by Caspr2 and that Caspr2 may modulate cytoplasmic Ca2+ levels. Combined with data from co-immunoprecipitation assays suggesting that Caspr2 interacts with the IP3R1 poreforming domain, these data are consistent with Caspr2 acting via IP3R1 to regulate cellular morphology during cerebellar development, most likely by directly affecting the IP3R1 pore region environment. Together, these experiments therefore reveal novel cellular and molecular roles for Caspr2 in the developing cerebellum. As well as improving our understanding of the nervous system, knowledge such as this will likely be increasingly clinically relevant, given the previously overlooked links between the cerebellum and cognition.
Supervisor: Becker, Esther ; Seiradake, Elena Sponsor: Wellcome Trust
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Cellular and molecular neuroscience ; Protein biochemistry