Functional Analysis Of Autism-associated Gene Shank In Drosophila Synapse

Various recent studies showed that postsynaptic Shank scaffold proteins involved in autism, schizophrenia and Alzheimer’s disease. Shank was firstly identified in 1998, and its family consists of three members:Shankl, Shank2 and Shank3. All of them expressed broadly in the brain, including areas of higher cognitive function like cortex and hippocampus. Its homologous gene exists in a wide spectrum of species from nematode to mammals, and its functional domain is highly conserved. Shank functions as major scaffolding molecules at postsynaptic density (PSD) within the central nervous system. Predominantly localized beneath the postsynaptic membrane and harboring multiple protein-protein interaction domains, they cross-link receptor complexes and cytoskeleton elements, thereby forming an indispensable framework for the assembly of the PSD. The disturbance of shank related signaling pathway causes the loss of synaptic stability, the failure of developmental mature and the breakage of balance between excitement and inhibition of neural circuit. However, its pathogenesis remains unknown, thus further study is necessary, which could offer a number of scientific theories about targeted therapy of related diseases. Drosophila melanogaster genome has only one shank gene and Drosophila is an ideal model to study function of nervous system. Therefore, this project employed Drosophila model and different kinds of experimental technique, such as molecule biology, biochemistry, genetics, development, electrophysiology, super-resolution microscopy and so on to reveal the Shank function and pathogenesis in nervous system.To investigate the role of Shank in Drosophila melanogaster, we established shank mutants and rabbit anti-Shank polyclonal antibody with good specificity. By using these tools, we described the distribution of Shank in nervous system of Drosophila. Null mutant results in behave obstacle, however, shank frame shift mutant results in abnormal NMJ structure. Shank is important in Drosophila neurodevelopment. Null mutant does not show abnormal NMJ structure as observed in frame-shift mutant, thus we supposed that shank frame shift mutant is gain-of-function allele, which is consistant with the results Durand founds in human disease model. Furthermore, IHC and electron microscope shows that PSD size of at NMJ in shank mutants increase compared with wild type significantly, and the development of postsynaptic reticular tissue is abnormal. These results suggest Shank plays important roles in the development of synapse.