Genetics of New Synaptic Components and Their Functions

This proposal focuses on two types of change in synapse morphology:expansion of postsynaptic membranes and formation of presynaptic boutons. Anatomicalspecializations are a hallmark of synapses. On the postsynaptic side, anatomical specializationsinclude dendritic spines and membrane folds. On the presynaptic side, an enlargement of the neuriteto form a rounded presynaptic bouton or en passant swelling is a nearly universal feature of synapses.Aim 1 of this proposal tests a specific hypothesis for how activity can regulate anatomical changes atthe postsynapse in the fly neuromuscular junction and in mammalian dendritic spines. In particular, itseeks to elucidate the role of Ral as a mediator of activity-dependent anatomical plasticity. We haveuncovered a novel pathway for synaptic plasticity in which the small GTPase Ral, by activating theexocyst complex, serves a central role in transducing Ca2+ influx from glutamate receptors intoenhanced transport of membrane to the postsynaptic region. In consequence, the membrane area ofthe postsynaptic specialization at the fly neuromuscular junction (NMJ) expands in an activitydependentmanner. This proposal asks what patterns of synaptic activity are necessary to recruit theexocyst and how Ral comes to be localized to postsynaptic membranes. It goes on to investigate thesignificance of Ral for the formation of dendritic spines in the mammalian CNS. Aim 2 uses Drosophilaas a model system in which to uncover the machinery that allows synaptic boutons to form. Thematuration of presynaptic terminals from growth cones to synaptic boutons is a critical late step insynaptogenesis but poorly understood. Neither the cytoskeletal elements that underlie the shapechange nor signaling molecules that trigger it are known. In a previous mutant screen in Drosophila wediscovered that mutations in α2δ-3 arrest synaptogenesis after initial synapse formation and preventthe formation of synaptic boutons at the fly NMJ. We propose to pursue the process of boutonformation in greater detail by a combination of biochemical and genetic approaches to uncoveradditional players in the formation of this poorly understood aspect of synaptic anatomy. Therefore, inAim 2A we will use biochemical methods to identify binding partners for α2δ-3 and determine if they arerequired for bouton formation. In Aim 2B we will uncover additional players in bouton formation through a mutant screen of the Drosophila genome at the embryonic NMJ.