Thesis (Ph.D)


Deciphering molecular interactions in presynaptic dense projection formation in C. elegans

Abstract

Neurons are unique in their capacity to build complex systems of information transfer, which comes as a product of their ability to communicate with one another. Neuronal communication occurs at biological structures known as synapses the majority of which are chemical. Chemical synapses comprise a presynaptic neuron, which releases molecular messengers such as neurotransmitters, and a postsynaptic cell, which receives these messages and respond appropriately. To perform its specific role the presynapse utilises a distinct set of proteins at a site known as the active zone, which dictates the location and timing of neurotransmitter release. These active zone functions are thought to revolve around a central protein scaffold, which is visible under electron microscopy as a structure known as the dense projection. While several of the proteins localised to these structures are known, the removal of individual components typically has limited effect on the formation of the presynaptic active zone scaffold/ dense projection. This has made it difficult to determine the function of the structure in neurotransmission. One of the only proteins which notably affects active zone scaffold formation is Liprin-α/SYD-2. In the nematode worm Caenorhabditis elegans the loss of SYD-2 causes fewer dense projections to form along motor neurons; the remaining structures are also smaller and have reduced ultrastructural complexity. As dense projections continue to form in the absence of SYD 2, however; there must be some degree of functional redundancy with other active zone proteins. This thesis explores the combined contribution of the known major active zone organiser SYD-2 (Liprin α), and the more recently characterised active zone proteins HLB-1 (Liprin-β) and CLA-1, in the formation of a functional presynaptic active zone scaffold in Caenorhabditis elegans. Both double and triple mutant strains carrying a syd-2 null mutation combined with hlb-1 and cla-1 mutations were generated and investigated alongside single mutants using a combination of approaches. These included confocal imaging of active zone components, locomotor behaviour assays, pharmacological assessment of neurotransmitter release and electron microscopy of the presynaptic ultrastructure. My results indicate that CLA-1 acts downstream of SYD-2 in the formation of the presynaptic active zone scaffold, whereas HLB-1 is not involved in the formation of the structure. Both CLA-1 and HLB-1 do have roles in managing the subsynaptic localisation of synaptic vesicles, however. CLA-1 maintains a subset of synaptic vesicles proximal to the dense projection and supports their docking at the presynaptic plasma membrane. HLB-1 meanwhile appears to act as a negative regulator of SYD 2 in synaptic vesicle docking. Loss of these proteins and their respective functions also modulates crawling locomotion. Therefore, while HLB 1 and CLA-1 can be eliminated as proteins which either work independently of SYD-2 or compensate for SYD-2 loss in active zone scaffold/ dense projection formation, this thesis provides new evidence for the respective roles of HLB-1 and CLA-1 at the presynaptic active zone.

DOI (Digital Object Identifier)

Permanent link to this resource: https://doi.org/10.24384/zsqx-ts15

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  • Type: PDF Document Filename: Cockram2023MolecularInteractions.pdf Size: 5.74 MB Views (since Sept 2022): 33

Authors

Cockram, Lewis A

Contributors

Supervisors: Kittelmann, Maike ; Bermudez, Isabel

Oxford Brookes departments

Department of Biological and Medical Sciences
Faculty of Health and Life Sciences

Dates

Year submitted for examination: 2023
RADAR publication date: 2024-01-24


© Cockram, Lewis A.
Published by Oxford Brookes University
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Details

  • Owner: Lewis Cockram (removed)
  • Collection: eTheses
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  • Views (since Sept 2022): 303