he assembly of functional neuronal circuits during development of the central nervous system requires an array of selective cell-cell interactions. These interactions direct cell migration, targeted growth and branching of axonal and dendritic processes, recognition of appropriate target cells, differentiation of pre- and postsynaptic structures, and recruitment of synapse-specific release machinery and neurotransmitter receptors. The aim of our studies is to understand the molecular mechanisms underlying the formation of specific connections between neurons in the central nervous system. In particular, we are examining the trans-synaptic signals that coordinate the choice of synaptic partners, assembly of synaptic junctions and stabilization of appropriate contacts. To address these questions, we are using a combination of functional in vitro assays that facilitate the identification of new signaling mechanisms and in vivo analysis of neuronal circuits in the mouse cerebellum. The cerebellum is an excellent model system for the analysis of synaptic specificity due to it’s highly organized structure. Moreover, we have established genetic labeling and marking techniques that facilitate dissection of synaptic specificity in vivo system.
Over the past years, we characterized the function of a family of neuronal cell adhesion molecules, called neuroligins and neurexins that form a heterophilic cell adhesion complex and that have potent synapse-organizing activities. These activities are regulated through extensive alternative splicing in sequences encoding the extracellular domains of these proteins. In our ongoing projects we are applying cell biological, biochemical, genetic and anatomical approaches to explore the molecular mechanisms underlying the choice of specific isoforms and isoform-specific functions. In the long-term, we aim to test the hypothesis that alternative splicing of neuroligin and neurexin isoforms underlies cell type- and synapse-specific interactions or properties in neuronal circuits. In a second set of studies we will examine how trans-synaptic cell surface interactions may be interpreted to guide the assembly of presynaptic structures. In particular, we are focusing on a novel protein called Synapse-defective-1 which has emerged as a key regulator of presynaptic assembly in invertebrates. The following aims will be the main focus of the research in my laboratory for the coming years and are described in detail in the Research Plan.
(1) Signaling Mechanisms Downstream of Synaptic Adhesion Molecules: In the vertebrate CNS multiple trans-synaptic adhesion systems have been identified that organize presynaptic terminals. However, it is unknown how adhesion complexes reorganize the axonal cytoskeleton and drive recruitment of active zone components and synaptic vesicles. We will examine the function of mammalian orthologues of Synapse-defective 1 (SYD1), a key regulator of synapse formation in invertebrates.
(2) Testing the Neurexin Code in Neuronal Networks: With over 3,000 isoforms, the Neurexins represent one of the molecularly most diverse families of neuronal cell surface proteins in vertebrates. Our previous studies suggest that neurexin splice isoform diversity underlies a synapse-specific adhesive code. In this project we will combine single cell analysis of genetically identified cerebellar neurons and cell type specific ablation of specific neurexin splice variants to examine the contribution of neurexins to neuronal identity and the selective wiring of cerebellar circuits.
(3) Activity-dependent Alternative Splicing Regulation of Synaptic Receptors: In this project we are analyzing the molecular machinery that regulates neurexin splice isoform choice. We have identified an alternative splicing factor that associates with neurexin-1 mRNA and is essential for the regulation of neurexin alternative splicing in vivo. Specifically, this splicing factor regulates the selection of exon 20, an alternative exon that plays a fundamental role in synapse-specific trans-synaptic signaling through select ligand interactions.