UC Davis

Healthy brain development requires the proper establishment of synaptic connections between neurons. Many synaptogenic molecules have been identified in the mammalian brain, but few synapse-limiting molecules have been discovered. Of the latter, many have canonical functions in the immune system. One such family of molecules, called major histocompatibility class I (MHCI), plays a key role in the innate and adaptive immune systems, where it serves as a ligand for cytotoxic effector cells. MHCI is also found on multiple cell types in the central nervous system. Neuronal MHCI has been found to regulate synapse density, visual system plasticity and synapse removal. Despite the many functions mediated by MHCI, it remains unclear whether MHCI has different functions in regulating the synapses formed onto dendrite or formed by axons of individual neurons, or what binding partners MHCI signals through to enact these functions. In this thesis, I consider the role of MHCI in regulating the number of synapses that a neuron makes onto its targets, and develop a methodological pipeline for identifying binding patterns of MHCI in young cortical neurons.In Chapter 1, I review the current literature of synapse formation and the roles of MHCI in the CNS. In Chapter 2, I describe experiments aimed at testing the sufficiency of H2-Kb and H2-Db to regulate the number of synapses that an axon forms onto dendrites

of neighboring neurons. I find that neither MHCI molecule affects the number of synapses that axons form, but both decrease the synapse density on dendrites of overexpressing neurons. I also report the novel finding that synapse density is increased on dendrites of young cortical neurons in the absence of H2-Kb and H2-Db. In Chapter 3, I describe a proximity biotinylation based approach coupled with tandem mass spectrometry for finding binding partners of MHCI in young cortical neurons, including novel TurboID fusion constructs, functional validation and a protocol for scaled up protein collection from cultured neurons. Chapter 4 describes the implications of my findings from Chapter 2 and alternative ways to study MHCI in the intact brain. I also propose follow-up experiments for the proteomics results in Chapter 3 and their connection to presynaptic roles of MHCI. Finally, I discuss the challenges and exciting advances in the future of MHCI biology in brain development.