Local protein synthesis allows for the rapid expression of proteins on-site and on-demand, as opposed to transporting pre-existing proteins from elsewhere. In neurons, the site-specific translation of mRNAs provides a means for restricting gene expression to individual structures that can be far from the cell body, such as synapses and growth cones. Dysregulated protein synthesis is implicated in a variety of neurodevelopmental and cognitive disorders. However, despite the importance of local translation for normal neuronal function, the most widely used methods for manipulating protein synthesis suffer from low spatial or temporal resolution. In this proposal we will generate new optogenetic methods for regulating the local translation of individual mRNA molecules. These methods will be broadly applicable to different mRNAs for use in a wide array of organisms and cell types.
The first aim develops a new loss-of-function approach to inhibit local translation, while the second aim generates a complementary gain-of-function method to acutely stimulate local protein synthesis. As a proof of principle, we will use these methods to determine both the requirement and sufficiency of local ?- actin translation for synaptic plasticity using primary hippocampal neurons. In response to neuronal activity, ?- actin mRNA is rapidly transported within dendrites to active synapses where it is translated, and the nascent ?- actin protein is specifically retained within the active dendritic spine. In this proposal, we will develop new tools that will provide researchers with exquisite control over post-transcriptional gene regulation in specific subcellular compartments.
The local synthesis of new proteins at synapses is critical for learning and memory. Subsequently, dysregulated protein synthesis is linked to a wide variety of human disorders, including Fragile X Syndrome, autism spectrum disorders, and Spinal Muscular Atrophy. The goal of this project is to develop new methods for manipulating local protein synthesis which can be applied to the study of neuronal development and function, and improve our understanding of these neurologic disorders.
