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D-Serine is a D-amino acid that occurs at high levels in the mammalian brain and is an endogenous ligand of the "glycine site" of N-methyl D-aspartate (NMDA) 1 receptors (1-4). NMDA receptors play key roles in excitatory synaptic transmission, plasticity, and learning and memory (5). Overactivation of the NMDA receptor and the resultant influx of calcium into cells is a major culprit in the cell death that occurs following stroke and neurodegenerative diseases. Blockers of the "glycine site" of the receptor are neuroprotective in animal models of stroke (5). Endogenous D-serine is required for NMDA receptor activation, and its removal markedly decreases NMDA receptor activity (3). In the vertebrate retina, endogenous D-serine may also mediate the light-dependent increase in neuronal activity by activating NMDA receptors (6). More recently, D-serine was suggested to play a role in the long term potentiation of synaptic transmission in the hippocampus, indicating a role of endogenous D-serine in long term synaptic plasticity (7).D-Serine is synthesized by serine racemase, a pyridoxal phosphate (PLP)-dependent enzyme enriched in the mammalian brain (8, 9). Serine racemase has high sequence homology with the fold-type II group of PLP enzymes, such as serine/threonine dehydratase and D-serine dehydratase (10, 11). In addition to converting L-to D-serine, serine racemase catalyzes the ␣,␤-elimination of water from L-serine to form pyruvate and ammonia (12). The initial rates of racemization and ␣,␤-elimination of L-serine by serine racemase are strongly stimulated by magnesium and ATP, indicating that the complex Mg⅐ATP is a physiological ligand of the enzyme (12).In accordance with accepted mechanisms of PLP-catalyzed reactions (13-16), a mechanism for racemization and ␣,␤-elimination catalyzed by serine racemase is depicted in Scheme 1. PLP, bound to the enzyme through an internal aldimine with The termination of signaling by a neurotransmitter in the brain normally requires its re-uptake and metabolism. D-Serine signaling is thought to involve its release from cells to

Mammalian serine racemase is a brain-enriched enzyme that converts L-into D-serine in the nervous system. D-Serine is an endogenous co-agonist at the "glycine site" of N-methyl D-aspartate (NMDA) receptors that is required for the receptor/ channel opening. Factors regulating the synthesis of D-serine have implications for the NMDA receptor transmission, but little is known on the signals and events affecting serine racemase levels. We found that serine racemase interacts with the Golgin subfamily A member 3 (Golga3) protein in yeast two-hybrid screening. The interaction was confirmed in vitro with the recombinant proteins in co-transfected HEK293 cells and in vivo by co-immunoprecipitation studies from brain homogenates. Golga3 and serine racemase co-localized at the cytosol, perinuclear Golgi region, and neuronal and glial cell processes in primary cultures. Golga3 significantly increased serine racemase steady-state levels in co-transfected HEK293 cells and primary astrocyte cultures. This observation led us to investigate mechanisms regulating serine racemase levels. We found that serine racemase is degraded through the ubiquitin-proteasomal system in a Golga3-modulated manner. Golga3 decreased the ubiquitylation of serine racemase both in vitro and in vivo and significantly increased the protein half-life in pulse-chase experiments. Our results suggest that the ubiquitin system is a main regulator of serine racemase and D-serine levels. Modulation of serine racemase degradation, such as that promoted by Golga3, provides a new mechanism for regulating brain D-serine levels and NMDA receptor activity.2 type of glutamate receptors play key roles in excitatory synaptic transmission and are involved in many physiological processes including learning and memory (1). NMDA receptor activity is tightly regulated, as its overactivation contributes to pathologic conditions such as stroke and neurodegenerative diseases (2). An interesting feature of NMDA receptors is the requirement of simultaneous binding of two agonists for channel opening, i.e. the NMDA channel only operates when both a glutamate site and a coagonist site are occupied (1). It has been shown that binding of glycine to the co-agonist site is an obligatory requirement for NMDA receptor/channel operation (3, 4). Subsequent studies have shown that brain D-serine is an endogenous ligand of the glycine site of NMDA receptors (5-9).Regulation of NMDA receptor activity by the co-agonist D-serine plays critical roles. Removal of endogenous D-serine decreases NMDA receptor responses (8) and blocks NMDAdependent migration of immature granule cells in the cerebellum (9). D-Serine is the dominant endogenous co-agonist for NMDA neurotoxicity, as removal of D-serine abolishes NMDA receptor-elicited cell death in hippocampal slices (6). In the vertebrate retina, endogenous D-serine mediates the light-dependent increase in neuronal activity by activating NMDA receptors (10). Furthermore, endogenous D-serine is required for the long term potentiation of the synaptic t...

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