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The molecular context of short stop during synaptic compartment formation in Drosophila

Reference	BB/C515998/1
Principal Investigator / Supervisor	Professor Andreas Prokop
Co-Investigators / Co-Supervisors
Institution	The University of Manchester
Department	Life Sciences
Funding type	Research
Value (£)	237,099
Status	Completed
Type	Research Grant
Start date	01/10/2005
End date	30/09/2008
Duration	36 months

Abstract

In contrast to mechanisms underlying the plasticity of synaptic cell junctions, very little is known about the machinery required for their de novo formation, ie. in contexts where neuronal circuits develop a new (eg. embryogenesis, postnatal period in mammals, metamorphosis on holometabolous insects). Synaptic cell junctions are composed of closely adhering compartments of pre- and postsynaptic cells (in the following simply termed synaptic compartments), and synapses are formed at these sites. The questions that we are interested in are how these compartments acquire their characteristic positions, sizes and shapes, and how structural and functional components of synapses become localised and assembled at these compartments. To address these questions and uncover the relevant molecular mechanisms we study embryos of the fruitfly Drosophila. Previously, we have introduced the cytoskeletal interacting spectraplakin protein Drosophila short stop (shot) as a key factor during the formation of two distinct synaptic compartments of motorneurons ¿ their postsynaptic dendrites and presynaptic neuromuscular terminals. Shot is predicted to execute its function via multiple interactions with other proteins. In a recent yeast two-hybrid screen with stringent selection procedures we have already uncovered 13 potential shot interactors. Further investigations are now required to capitalise on the potential of these genes. For one of them, Drosophila paxillin (Dpxn) we have already obtained independent proof that it interacts with shot directly and down-regulates growth-promoting activity of shot at NMJs. Experiments will be carried out to test our additional findings that the interaction of Dpxn with shot seems dependent on phosphorylation through phosphokinase A (PKA). Of the remaining 12 candidate genes, 5 are of completely unknown function showing no obvious homologies, 3 contain domains with known homologies (SH3, SAM/pointed, RNP1-like RNA-binding region), and 4 genes havebeen cloned previously during work in other biological contexts (two to them in the nervous system). We will use phenotypic analyses in combination with genetic deficiencies, existing mutations or RNA interference strategies to select those candidate genes required for synaptic compartment formation. We will determine the spatial and temporal expression profiles of these genes. Finally, we will study their functions via in silico searches of data bases, detailed phenotypic analyses and biochemical and genetic experiments testing their interactions with shot. Analyses of the candidate genes have great potential to advance the sparse molecular insights into the mechanisms underlying the formation of synaptic compartments or cell junctions. They will enhance our understanding of shot function at the molecular level. Moreover, shot is closely related to a mammalian homologue, MACF1/ACF7, and we believe that our work on flies will open up important avenues of research for understanding how synaptic cell junctions develop and are maintained in higher organisms.

Summary

unavailable

Committee	Closed Committee - Genes & Developmental Biology (GDB)
Research Topics	Neuroscience and Behaviour, The 3 Rs (Replacement, Reduction and Refinement of animals in research)
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

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