Ultrastructural analysis of proteins involved in synaptic development and function (PhD Dissertation Seminar)

Abstract: The nervous system of animals underlies our ability to sense and respond to our environment and to learn from these experiences. Neurons within this system process and transmit information to their target cells through the release of chemical neurotransmitters at highly specialized endings known as synapses. These synaptic terminals contain an active zone that consists of an electron dense structure, known as the dense projection. At the active zone, a family of conserved proteins help convert incoming electrical signals to outgoing chemical signals. Electron microscopy shows that the dense projection is surrounded by synaptic vesicles that we know contain the neurotransmitters to be released. Decades of research have shed light on essential machinery and processes that underlie synaptic development and transmission, yet these components and their mechanisms of action are not completely understood. Unlike higher organisms, many active zone proteins are represented by a single homolog in the soil nematode Caenorhabditis elegans, whose well-defined nervous system and robust genetic capabilities provide an ideal model system. The neuromuscular junction of C. elegans is particularly well-suited for examination by high-pressure freeze and freeze substitution electron microscopy. Using this technique, I characterized the ultrastructural phenotypes of mutations in genes critical for organizing and regulating synapses. Two of these genes encode novel active zone proteins, which we have named clarinet and sentryn. Clarinet regulates synapse number and vesicle release, helps tether synaptic vesicles to the dense projection and may be an integral component of this structure. Sentryn operates with a specialized class of active zone enriched proteins, including SAD Kinase and SYD-2/Liprin-a, to capture synaptic and dense core vesicles at synapses. Sentryn and SAD Kinase also influence the size of the dense projection. Finally, we have made inroads into the function of a transcription factor UNC-55, DEG/ENaC channel UNC-8, and Iroquois homeodomain protein IRX-1 in the selective removal of active zone proteins during synaptic disassembly in a developmental remodeling program.