Synapse Development and Function in C. elegans
The human brain consists of over 100 billion individual neurons. A single Hippocampal neuron in the brain can make 10,000 synaptic connections and utilize many different neurotransmitter receptors. How are specific receptors targeted to the correct synapse? Research in the lab is focused on understanding how synaptic proteins and neurotransmitter receptors, like the glutamate receptor, are specifically targeted and regulated at synapses in the genetic model organism, C. elegans. The worm C. elegans has a simple, compact nervous system with exactly 302 neurons and 7,000 chemical synapses. We use a combination of genetics, biochemistry, quantitative fluorescence microscopy, and behavior to study the molecular mechanisms involved in the targeting and turnover of synaptic proteins.
Neurons communicate with each other through specialized junctions called synapses. Synapses consist of a pre- and postsynaptic element. The presynaptic element is designed for the regulated release of neurotransmitter filled synaptic vesicles. The postsynaptic element consists of neurotransmitter receptors, ion channels, scaffolding molecules and signaling molecules, and is specifically designed to respond to neurotransmitter that has been released into the synaptic cleft. Understanding the genes and molecular mechanisms involved in localizing and regulating synaptic proteins will reveal how synapses are built during development (synaptogenesis), and regulated in the mature nervous system during learning and memory. Deregulated synaptic transmission has been implicated in many diseases of the nervous system. Aberrant glutamate receptor signaling mediates excitotoxicity-induced neurodegeneration in response to stroke and ischemia (reduced blood flow). Studies revealing the function of proteins at normal synapses will provide the foundation for understanding what has gone awry in various neurological diseases.
Figure 1. Regulation of the abundance of glutamate receptors (shown in red) in the postsynaptic membrane is highly dynamic and plays a major role in regulating synaptic strength during learning and memory. Ubiquitination of GLR-1 (Ub-GLR-1, right panel) results in the endocytosis and downregulation of the glutamate receptor.
Glutamate Receptor Trafficking
Glutamate is the major excitatory neurotransmitter in the mammalian brain and the abundance of postsynaptic glutamate receptors regulates the strength of synaptic transmission. We are interested in identifying genes and mechanisms involved in regulating glutamate receptor levels at the synapse. We have discovered two novel regulators of synaptic glutamate receptors in C. elegans, the ubiquitin ligase APC and the kinase CDK5.
Figure 2. We use fluorescence microscopy to quantitate the abundance and subcellular localization of synaptic proteins in vivo. The right panel illustrates the pan-neuronal expression pattern of the p35 gene.
Ubiquitin and the Anaphase Promoting Complex as Synapses
The Anaphase Promoting Complex (APC) is a multisubunit ubiquitin ligase known for its role in regulating cell cycle progression. Like phosphorylation, ubiquitin modification of proteins has emerged as a widely used post-translational regulatory signal with diverse cell biological roles. Recent evidence implicates ubiquitination as an important signaling mechanism that regulates synaptic transmission. We have discovered a novel role for the APC, outside the cell cycle, in postmitotic neurons. We showed that the APC regulates the abundance of the glutamate receptor GLR-1 at synapses in C. elegans. Little is known about how the APC functions in neurons. Current projects in the lab are focused on identifying upstream regulators and downstream targets of the APC in neurons.
The Role of CDK5 at Synapses
CDK5 is a cyclin-dependent kinase which plays a role in many cellular processes such as cell migration, axon outgrowth and neurodegeneration. CDK5 activity requires association with the cyclin-like regulatory molecule, p35. As in mammals, CDK5 is more broadly expressed, whereas p35 is expressed almost exclusively in the nervous system. Recent work suggests that CDK5 can also function at synapses. We have found that CDK5 positively regulates glutamate receptors at synapses in C. elegans. This effect may be mediated, in part, through the scaffolding protein LIN-10/Mint1, since CDK5 can directly phosphorylate LIN-10/Mint1 in vitro and negatively regulate LIN-10 levels in vivo. Current projects are focused on understanding how CDK5 regulates glutamate receptors at synapses, and the role of CDK5 at the neuromuscular junction.