Synapses are continuously modified by experience- and usage-dependent manner. This plastic nature of synapses forms the basis of information storage in the brain. The long-term goal of my research program is to understand molecular and cellular mechanisms of synaptic plasticity. Unraveling the cell biological mechanisms of how neurons control precisely the targeting, clustering, and removal of synaptic proteins into and from synapses, and how synaptic activity modulates these processes, is a fundamental goal in my lab. These protein trafficking events are all intimately related to the proper development of neurons and the efficacy of synaptic transmission, and further learning and memory. In particular, we are interested in the dynamic reorganization and movement of postsynaptic proteins in excitatory synapses during experience. We employ multi-disciplinary approaches ranging from biochemistry and molecular biology to cell biology, imaging, and mouse genetics to address these questions.
Activity-dependent Remodeling of PSD Proteins in Excitatory Synapses
A hallmark of excitatory synapses in the central nervous system is electron-dense structure underlying the postsynaptic membrane called 'postsynaptic density' (PSD). The PSD consists of neurotransmitter receptors, scaffolds and cytoskeletal proteins, and signaling molecules (see diagram). These proteins put together a signaling microdomain, translating presynaptic inputs to postsynaptic responses. Two types, AMPA- and NMDA-type, of glutamate receptors mediate most of the fast excitatory transmission. AMPA receptors provide the depolarization of postsynaptic membrane, which in turn helps open NMDA receptors to drive Ca2+ influx that triggers signaling cascades.
Interestingly, protein components in the PSDs are highly regulated by synaptic activity; NMDA receptor activation leads to either removal or accumulation of different set of proteins from or in synapses. Ubiquitin-dependent protein degradation mechanism seems important for the removal of synaptic proteins.
We aim to dissect out the detailed molecular mechanisms underlying the activity-dependent remodeling of PSD proteins. Among many synaptic proteins, current research focus on a scaffold protein called GKAP (Guanylate Kinase Associated Protein, a.k.a. SAPAP or DAP), which connects NMDA receptors/PSD-95 family of proteins to Shank/Homer complex. This protein exhibits one of the most dramatic changes by activity modulation. The specific questions we're trying to address are: what is the molecular queue causing the removal of this protein from synapses? Is this process dependent on the protein phosphorylation? What protein kinase might be responsible? How does the phosphorylation event relate to ubiquitination? What ubiquitin ligase is involved? What functional relationship does the removal or accumulation of GKAP have with the regulation of glutamate receptors and other synaptic proteins? In addition to answering these questions, ongoing efforts are underway to visualize the movement of GKAP in and out of synapses in real time by live-imaging. If successful, GKAP may serve as a vital marker for synaptic activity.
Protein Trafficking and Synaptic Plasticity
AMPA receptor trafficking is the most extensively studied model in synaptic plasticity. Increased incorporation of AMPA receptors to synapses correlates with the long term potentiation (LTP) of synaptic efficacy. Conversely, the removal of synaptic AMPA receptor provides mechanism for long term depression (LTD). Since synaptic activity seems to regulate not only the trafficking of glutamate receptors but also a variety of synaptic proteins, it becomes increasingly important to analyze the trafficking mechanisms of those proteins as well. Specifically, the research in my lab is centered on the identification of specific motor proteins that are involved in the transport of postsynaptic proteins to and from synapses. Recent data suggest that an actin-based motor protein MyoV is important for the transport of at least one postsynaptic protein, GKAP, to synapses via dynein light chain (DLC) interaction. Further characterization of GKAP-DLC-MyoV interaction is ongoing in my laboratory. As an independent and non-biased way to screen for specific motor proteins, we are also developing reagents based on siRNA knockdown strategy.
Evers, D.M., Matta, J.A., Hoe, H.-S., Zarkowsky, D., Lee, S.H., Issac, J.T., and Pak, D.T.S. (2010) Plk2 attachment to NSF induces homeostatic removal of GluA2 during chronic overexcitation. Nat. Neurosci. 13, 1199-1207.
Danielson, E., Zhang, N., Metallo, J., Kaleka, K., Shin, S.M., Gerges, N., and Lee, S.H. (2012) S-SCAM is an essential synaptic scaffolding molecule for the GluA2-containing maintenance pool of AMPA receptors. J. Neurosci. 32, 6967-6980.
Danielson, E., Metallo, J., and Lee, S.H. (2012) Role of TARP interaction in S-SCAM-mediated regulation of AMPA receptors. Channels 6, 393-397.
Shin, S.M., Zhang, N., Hansen, J., Gerges, N.Z., Pak, D.T.S., Sheng, M., and Lee, S.H. (2012) GKAP orchestrates activity-dependent postsynaptic protein remodeling and homeostatic scaling. Nat. Neurosci. 15, 1655–1666.
Okamoto, K.-I., Narayanan, R., Lee, S.H., Murata, K., and Hayashi, Y. (2007) The role of CaMKII as an F-actin-bundling protein crucial for maintenance of dendritic spine structure. Proc. Natl. Acad. Sci. U.S.A. 104, 6418-6423.
Kastning, K., Kukhtina, V., Kittler, J.T., Chen, G., Yan, Z., Enders, S., Pechstein, A., Lee, S.H., Sheng, M., and Haucke, V. (2007) Molecular determinants for the interaction of AMPA receptors and the clathrin adaptor complex AP-2. Proc. Natl. Acad. Sci. U.S.A. 104, 2991-2996.
Fu, Z., Lee, S.H., Simonetta, A., Hansen, J., Sheng, M. and Pak, D.T.S. (2007) Differential roles of Rap1 and Rap2 small GTPases in neurite retraction and synapse elimination in hippocampal spiny neurons. J. Neurochem. 100, 118-131.
Lee, S.H., Simonetta, A., Sheng, M. (2004) Subunit rules governing the sorting of internalized AMPA receptors in hippocampal neurons. Neuron 43, 221-236.
Ahmadian, G., Ju, W., Liu, L., Wyszynski, M., Lee, S.H., Dunah, A.W., Taghibiglou, C., Wang, C., Liu, J., Wong, T.P., Sheng, M., Wang Y.T. (2004) Tyrosine phosphorylation of GluR2 is required for insulin-stimulated AMPA receptor endocytosis and LTD. EMBO J. 23, 1040-1050.
Sheng, M., Lee, S.H. (2003) AMPA receptor trafficking and synaptic plasticity: major unanswered questions. Neurosci. Res. 46, 127-134.
Lee, S.H., Liu, L., Wang, Y.T., Sheng, M. (2002) Clathrin adaptor AP2 and NSF interact with overlapping site of GluR2 and play distinct roles in AMPA receptor trafficking and hippocampal LTD. Neuron 36, 661-674.
Lee, S.H., Valtschanoff, J.G., Kharazia, V.K., Weinberg, R., Sheng, M. (2001) Biochemical and morphological characterization of an intracellular membrane compartment containing AMPA receptors. Neuropharmacology 41, 680-692.
Sheng, M., Lee, S.H. (2001) AMPA receptor trafficking and the control of synaptic transmission. Cell 105, 825-828.
Lin, J.W., Ju, W., Foster, K., Lee, S.H., Ahmadian, G., Wyzynski, M.W., Wang, Y.T., Sheng, M. (2000) Distinct molecular mechanisms and divergent endocytotic pathways of AMPA receptor internalization. Nat. Neurosci. 3, 1282-1290.
Sheng, M., Lee, S.H. (2000) Growth of the NMDA receptor industrial complex. Nat. Neurosci. 3, 633-635.
Lee, S.H., Sheng, M. (2000) Development of Neuron-Neuron Synapses. Curr. Opin. Neurobiol. 10, 125-131.