Final Report Summary - MASDUHID (Molecular Analysis of Synaptic Dysfunctions Underlying Human Intellectual Disabilities) Synapses of the central nervous system are the cellular structures mediating communication between neurons. Mouse mutations studies of synaptic proteins have shown how they ultimately modulate animal cognition and behaviour. Furthermore, there is a growing body of data suggesting a central role of the synapse in neurodegenerative, mental and behavioural disorders. This project aims at understanding the synaptic molecular dysfunctions associated with intellectual disorders, particularly non-syndromic intellectual disability (NSID). By doing so it also aspires at learning on the molecular mechanisms behind cognition. The current proposal also attempts at identifying proteins that can become pharmacological targets to treat intellectual disability and to study to what extend mental disorders can be reverted after birth. The overarching goal of this research is to study the synapse as a whole biological system and to characterize the changes it undergoes due to disease. That is why proteomics and bioinformatics will be used to quantitatively analyse the dynamics of the synaptic proteome in control and model animals. With the disease-related molecular dysfunctions characterized it might be possible to identify pharmacological targets and to test drugs in the animal models to look for phenotypic recovery. All this objectives will only be achieved if we can improve the anatomical resolution at which we perform proteomics. This has also become a major scientific goal in the project. We have focussed on developing new methods that shall allow us to look at the synaptic proteome at a much lower scale. Proteomics has always lagged behind other neuroscience technologies, such as imaging or electrophysiology technologies, in the resolution of its analysis. While we can record synaptic activity or look at protein localization in one neuron, proteomics typically require large brain regions. This is a major drawback for the advancement of brain molecular studies, as brain is a highly heterogeneous tissue in the human body. To overcome this we have developed a micro-proteomics protocol based on the combined use of i) Laser capture microdissection ii) biochemical enrichment in synapses of microdissected tissue and iii) mass spectrometry proteome profiling. Using this we have characterised the postsynaptic proteome of glutamatergic synapses from CA1, CA3 and dentate gyrus in the mouse hippocampus, identifying those proteins specific to the synapses of these regions, which confer them unique electrophysiological characteristics. Furthermore we have also investigated postsynaptic proteome dynamics in CA1 after LTP induction, characterising the main changes in the synaptic molecular composition after LTP induction. Regarding the molecular analysis of NSID we have focused our attention on studying the model of the autosomal dominant intellectual disability caused by SYNGAP1 haploinsufficiency. We have been analysing how does the hippocampal synaptic proteome respond to reducing SynGAP protein by half and recovering it back to normal. This can be achieved by the combined use of two Syngap1 conditional knock-out (KO) mouse lines, a KO inducible line and a rescue line. The Career Integration Grant (CIG) has become fundamental for the establishment of the laboratory directed by the fellow (www.moleuclar-synapse.org) importantly contributing to his establishment as an independent investigator and fostering the successful application to other funding opportunities.
