Struktur-Funktions Analyse der aktiven Zone in Drosophila
Final Report Abstract
Here, new functionalities of the BRP cytomatrix for the synaptic exo-/endocycle were uncovered by systematically analyzing BRP isoforms using and further adapting knock-in like genetic techniques in the Drosophila system. In our system the Drosophila NMJ, it has become clearer that the scaffold protein BRP might function as a large docking station that brings many crucial AZ components like Ca2+ channels, SVs and proteins with different functionalities together, either via direct interaction or via establishing a protein meshwork. Generating isoform-specific mutants, the functions of the major 170 and 190kDa BRP isoforms were investigated. With the help of isoform-specific genomic constructs and alleles, both isoforms were found to alternate in a circular array within individual active zones of the larval neuromuscular junction. Most importantly, lack of both isoforms reduced active zone cytomatrix size, and likely as a direct result the synapses suffered from a reduction in the size of the readily-releasable pool of SVs (RRP). Correspondingly, a reduction in the number of SVs docked close to the remaining cytomatrix could be observed. Hence, a novel role for the membrane-close active zone cytomatrix was revealed in maintaining or setting the proper size of the RRP, which was previously masked by the strong vesicular release probability defect of brp null mutants. Furthermore, specific roles of the BRP-isoforms were analyzed in the Drosophila brain. Both brp isoforms revealed a clearly distinct expression pattern within specific brain regions. Loss of BRP-190 – but not so much BRP-170 – had dramatic consequences on adult vitality and locomotion. For BRP-170 mutants, a specific deficit in anesthesia resistant olfactory memory could be demonstrated. This work here is a starting point for systematic attempts to unravel principles of active zone diversification in the Drosophila CNS and beyond. We thus were able to characterize different synapse types of the Drosophila PNS and CNS with regard to a type-specific spectrum of BRP isoforms. We here exemplarily tested the hypothesis that differences in the composition of the AZ-associated cytomatrix contribute to functional diversity between synapse types.
Publications
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Naked dense bodies provoke depression. J Neurosci. 2010 Oct 27;30(43):14340-5
Hallermann S, Kittel RJ, Wichmann C, Weyhersmüller A, Fouquet W, Mertel S, Owald D, Eimer S, Depner H, Schwärzel M, Sigrist SJ, Heckmann M
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The active zone T-bar -a plasticity module? J Neurogenet. 2010 Sep;24(3):133-45
Wichmann C, Sigrist SJ
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(2013): „ Bruchpilot and RIM-binding protein determine presynaptic release“
Elena Knoche
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(2013): „ Bruchpilot isoforms regulate the ready-releasable pool of synaptic vesicles “
Tanja Matkovic
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(2013):” Structure, Function and Transport of Components of the Cytomatrix at the Active Zone in Drosophila”
Matthias Siebert
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The Bruchpilot cytomatrix determines the size of the readily releasable pool of synaptic vesicles. J Cell Biol. 2013 Aug 19;202(4):667-83
Matkovic T, Siebert M, Knoche E, Depner H, Mertel S, Owald D, Schmidt M, Thomas U, Sickmann A, Kamin D, Hell SW, Bürger J, Hollmann C, Mielke T, Wichmann C, Sigrist SJ
