Omics Approaches for the Analysis of Shank1/3-associated Autism Spectrum Disorders and the Lessel-Kreienkamp syndrome ; Omics-Ansätze zur Analyse der Shank1/3-assoziierten Autismus-Spektrum-Störungen und des Lessel-Kreienkamp-Syndroms

Neurodevelopmental disorders arise due to malfunctions in brain development and often are genetically caused. The disease-associated genes encode three major classes of proteins: transcriptional regulators, synaptic proteins, and RNA-binding proteins. It was proposed that the pathomechanisms may converge in shared molecular pathways. I functionally analyzed and compared the pathomechanisms of insufficiency for SHANK1 and SHANK3, two genes associated with autism spectrum disorders, with a variant of the AGO2 gene, encoding the RNA-binding protein Argonaute-2 and causing Lessel-Kreienkamp syndrome. The SHANK genes encode Shank proteins, scaffolds of excitatory, glutamatergic synapses that indirectly connect postsynaptic glutamate receptors to F-actin via a protein network. The N-terminus of Shank3 interacts with Ras family G-proteins, connecting Shank to the MAPK pathway, which is involved in translational regulation. Altered translation has been implicated in the pathology of autism spectrum disorders. The hypothesis of this project was that loss of Shank may alter the regulation of translation. Omics were applied to test this hypothesis. The translatome and proteome of mouse models for Shank-associated autism spectrum disorders were investigated. Actively translated mRNAs were purified from hippocampal neurons of Shank3αβ knockout mice via RNA affinity purification and analyzed by RNA sequencing. Absence of Shank3αβ resulted in subtly altered translation of a subset of neuronal transcripts. Proteomics were performed with biochemically purified postsynaptic density fractions from hippocampi of Shank1 and Shank3αβ knockout mice. The loss of Shank1 and Shank3αβ severely altered the postsynaptic proteome. The abundance of active, phosphorylated CaMKIIα was increased in Shank knockout mice, which may contribute to misregulated neuronal signaling. Argonaute-2 functions in RNA interference. To execute translational silencing, Argonaute-2 associates with microRNA and forms the RNA induced silencing complex. It was ...