The long-term goal of this project is to understand the molecular mechanisms of neurotransmitter signaling through heterotrimeric G proteins. A set of >100 G protein coupled neurotransmitter and neuropeptide receptors signal in the brain by activating a small set of heterotrimeric G proteins. By far the most abundant neural G protein is G?o, which inhibits neural function by a mechanism that remains poorly understood. Our genetic work in C. elegans suggests that signaling by G?o in given neuron is not predominantly activated by any one receptor, but rather may result from the additive effects of many different receptors.
Our first aim i s to understand how many different receptors present in a single neuron can all activate one G protein to do something that makes biological sense. We will create a GPCR expression atlas and use it to identify all the receptors expressed in a model C. elegans neuron, knock them out in combinations, and analyze the resulting effects on G?o signaling.
Our second aim i s to characterize how acetylation of G?o is used to regulate neural signaling. We recently used mass spectrometry to show that G?o in both worms and mouse brain is post- translationally modified by acetylation on multiple lysine residues. Knocking out a lysine acetyltransferase enzyme that creates these modifications results in specific defects in serotonin signaling. We will determine how and why acetylation is used to regulate G?o signaling.
Our third aim i s to identify the long-sought effectors through which G?o signals. A mystery in G?o signaling is that no downstream ?effector? protein activated by directly binding to G?o has yet been identified. Genetic studies in C. elegans suggest that such an effector should exist, yet a variety of approaches have failed to identify it. In the course of our mass spectrometry analysis of G?o, we found that specific neural signaling proteins can co-purify with G?o at sub-stoichiometric levels. We will test the hypothesis that these proteins are G?o effectors.
Aim 1. We will identify every cell in C. elegans expressing its 26 G protein coupled small molecule neurotransmitter receptors, and document and partially identify the cells expressing >100 neuropeptide receptors. We will demonstrate the utility of this GPCR atlas by identifying the receptors expressed in a specific pair of neurons that control egg laying, and then characterize knockouts for these receptors to determine how the receptors function together.
Aim 2. We will determine how acetylation of G?o by the lysine acetyltransferase ?elongator' affects serotonin signaling, and how this acetylation is regulated.
Aim 3. We will determine if signaling proteins that co-purify with G?o function as its long-sought effectors.
This proposal investigates the basic mechanisms by which neurons in the brain communicate with each other using chemical neurotransmitters, focusing in particular on the neurotransmitter serotonin. Defects in signaling by serotonin and other neurotransmitters appear to underlie depression, schizophrenia and other disorders of mental health, but the precise details of these defects are in most cases unknown. Understanding in detail how neurotransmitters act should make it possible to precisely define the defects that cause mental disorders and to thus develop effective treatments for them.
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