The Experts below are selected from a list of 309 Experts worldwide ranked by ideXlab platform
Ayse Dosemeci - One of the best experts on this subject based on the ideXlab platform.
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FAM81A protein, a novel component of the Postsynaptic Density in adult brain.
Neuroscience letters, 2019Co-Authors: Ayse Dosemeci, Thomas S. Reese, Hannah Loo, Dana Toy, Christine A. Winters, Jung-hwa Tao-chengAbstract:Abstract Analysis of affinity-purified PSD-95 complexes had previously identified a ‘hypothetical protein’, product of the gene FAM81A [ 1 ]. The present study examined the tissue and subcellular distribution of FAM81A protein and its expression levels during development. Comparison of different organs indicates selective expression of FAM81A protein in brain. FAM81A is expressed late in development, with a post-natal gradual increase in brain levels that parallels the expression of PSD-95. Comparison of subcellular fractions from adult brain shows that the distribution of FAM81A protein is similar to that of PSD-95, with a drastic enrichment in the Postsynaptic Density fraction. Immuno-electron microscopy of adult brain tissue reveals specific immunogold labeling for FAM81A protein at Postsynaptic densities in the forebrain. The label for FAM81A protein is concentrated at the cytoplasmic edge of the electron-dense core of the Postsynaptic Density, with a mean distance of ∼33 nm from the Postsynaptic membrane. These observations firmly establish FAM81A protein as a component of the Postsynaptic Density in the adult brain, suggesting a role in synaptic function.
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irsp53 accumulates at the Postsynaptic Density under excitatory conditions
PLOS ONE, 2017Co-Authors: Ayse Dosemeci, Hannah Loo, Dana Toy, Junghwa Taocheng, Amelia BurchAbstract:IRSp53 (BAIAP2) is an abundant protein at the Postsynaptic Density (PSD) that binds to major PSD scaffolds, PSD-95 and Shanks, as well as to F-actin. The distribution of IRSp53 at the PSD in cultured hippocampal neurons was examined under basal and excitatory conditions by immuno-electron microscopy. Under basal conditions, label for IRSp53 is concentrated at the PSD. Upon depolarization by application of a medium containing 90 mM K+, the intensity of IRSp53 label at the PSD increased by 36±7%. Application of NMDA (50 μM) yielded 53±1% increase in the intensity of IRSp53 label at the PSD compared to controls treated with APV, an NMDA antagonist. The accumulation of IRSp53 label upon application of high K+ or NMDA was prominent at the deeper region of the PSD (the PSD pallium, lying 40–120 nm from the Postsynaptic plasma membrane). IRSp53 molecules that accumulate at the distal region of the PSD pallium under excitatory conditions are too far from the plasma membrane to fulfill the generally recognized role of the protein as an effector of membrane-bound small GTPases. Instead, these IRSp53 molecules may have a structural role organizing the Shank scaffold and/or linking the PSD to the actin cytoskeleton.
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The Postsynaptic Density: There Is More than Meets the Eye.
Frontiers in synaptic neuroscience, 2016Co-Authors: Ayse Dosemeci, Thomas S. Reese, Jung-hwa Tao-cheng, Richard J WeinbergAbstract:The Postsynaptic Density (PSD), apparent in electron micrographs as a dense lamina just beneath the Postsynaptic membrane, includes a deeper layer, the “pallium,” containing a scaffold of Shank and Homer proteins. Though poorly defined in traditionally prepared thin-section electron micrographs, the pallium becomes denser and more conspicuous during intense synaptic activity, due to the reversible addition of CaMKII and other proteins. In this Perspective we review the significance of CaMKII-mediated recruitment of proteins to the pallium with respect to both the trafficking of receptors and the remodeling of spine shape that follow synaptic stimulation. We suggest that the level and duration of CaMKII translocation and activation in the pallium will shape activity-induced changes in the spine.
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camkii mediated displacement of aida 1 out of the Postsynaptic Density core
FEBS Letters, 2016Co-Authors: Ayse Dosemeci, Dana Toy, Junghwa Taocheng, Amelia Burch, Ulrich K BayerAbstract:Ankyrin repeat and sterile alpha motif domain-containing protein 1B (ANKS1B, also known as AIDA-1) is a major component of the Postsynaptic Density (PSD) in excitatory neurons where it concentrates at the electron-dense core under basal conditions and moves out during activity. This study investigates the molecular mechanism underlying activity-induced displacement of AIDA-1. Experiments with PSD fractions from brain indicate phosphorylation of AIDA-1 upon activation of endogenous CaMKII. Immuno-electron microscopy studies show that treatment of hippocampal neurons with NMDA results in an ~ 30 nm shift in the median distance of the AIDA-1 label from the Postsynaptic membrane, an effect that is blocked by the CaMKII inhibitor tatCN21. CaMKII-mediated redistribution of AIDA-1 is similar to that observed for SynGAP. CaMKII-mediated removal of two abundant PSD-95-binding proteins from the PSD core during activity is expected to initiate a molecular reorganization at the PSD.
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zinc stabilizes shank3 at the Postsynaptic Density of hippocampal synapses
PLOS ONE, 2016Co-Authors: Junghwa Taocheng, Thomas S. Reese, Dana Toy, Christine A. Winters, Ayse DosemeciAbstract:Shank3 is a Postsynaptic Density (PSD) scaffold protein of the Shank family. Here we use pre-embedding immunogold electron microscopy to investigate factors influencing the distribution of Shank3 at the PSD. In dissociated rat hippocampal cultures under basal conditions, label for Shank3 was concentrated in a broad layer of the PSD, ~20–80 nm from the Postsynaptic membrane. Upon depolarization with high K+ (90 mM, 2 min), or application of NMDA (50 μM, 2 min), both the labeling intensity at the PSD and the median distance of label from the Postsynaptic membrane increased significantly, indicating that Shank3 molecules are preferentially recruited to the distal layer of the PSD. Incubation in medium supplemented with zinc (50 μM ZnCl2, 1 hr) also significantly increased labeling intensity for Shank3 at the PSD, but this addition of Shank3 was not preferential to the distal layer. When cells were incubated with zinc and then treated with NMDA, labeling intensity of Shank3 became higher than with either treatment alone and manifested a preference for the distal layer of the PSD. Without zinc supplementation, NMDA-induced accumulation of Shank3 at the PSD was transient, reversing within 30 min after return to control medium. However, when zinc was included in culture media throughout the experiment, the NMDA-induced accumulation of Shank3 was largely retained, including Shank3 molecules recruited to the distal layer of the PSD. These results demonstrate that activity induces accumulation of Shank3 at the PSD and that zinc stabilizes PSD-associated Shank3, possibly through strengthening of Shank-Shank association.
Mary B Kennedy - One of the best experts on this subject based on the ideXlab platform.
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A sex difference in the response of the rodent Postsynaptic Density to synGAP haploinsufficiency
eLife, 2020Co-Authors: Tara L Mastro, Anthony Preza, Shinjini Basu, Sumantra Chattarji, Sally M. Till, Peter C. Kind, Mary B KennedyAbstract:SynGAP is a Postsynaptic Density (PSD) protein that binds to PDZ domains of the scaffold protein PSD-95. We previously reported that heterozygous deletion of Syngap1 in mice is correlated with increased steady-state levels of other key PSD proteins that bind PSD-95, although the level of PSD-95 remains constant (Walkup et al., 2016). For example, the ratio to PSD-95 of Transmembrane AMPA-Receptor-associated Proteins (TARPs), which mediate binding of AMPA-type glutamate receptors to PSD-95, was increased in young Syngap1+/-mice. Here we show that only females and not males show a highly significant correlation between an increase in TARP and a decrease in synGAP in the PSDs of Syngap1+/-rodents. The data reveal a sex difference in the adaptation of the PSD scaffold to synGAP haploinsufficiency.
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Liquid Phase Transition in the Postsynaptic Density
Trends in biochemical sciences, 2016Co-Authors: Mary B Kennedy, Tara L MastroAbstract:An assembly of scaffold proteins termed the Postsynaptic Density (PSD) is attached to the Postsynaptic membrane of excitatory glutamatergic synapses [1]. The scaffold serves to immobilize glutamate receptors in the membrane directly across from the position where glutamate is released from the presynaptic terminal. It also houses and organizes biochemical machinery whose job is to respond to particular patterns of electrical activity by increasing the strength of the synapse [2]. Synaptic strengthening helps to form new circuits that represent our experience [3]. These circuits are our memories. Biochemical machinery in the PSD strengthens the synapse by increasing the size of the PSD scaffold, the number of anchored receptors, and the size of the presynaptic active zone. A larger scaffold, more release sites, and more receptors means a stronger synapse, and vice versa.
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Organizing signal transduction in the Postsynaptic Density
The FASEB Journal, 2014Co-Authors: Mary B KennedyAbstract:Excitatory neurons in the CNS receive thousands of synaptic contacts from other excitatory neurons. An increase in the strength of a relatively small number of these synapses can store a memory by binding connected neurons into a circuit in which all of the neurons fire each time the specific memory is evoked. An increase in synaptic strength involves addition of new glutamate receptors to the synaptic membrane, and re-arrangement of the cytoskeleton to support a larger synapse. Biochemical signaling enzymes that control this process, which is called “activity-dependent synaptic plasticity,” are organized near the Postsynaptic membrane. The enzymes include several protein kinases, adenylyl cyclase, and regulators of Ras and Rap. Many of them are attached to a multiprotein scaffold called the “Postsynaptic Density.” At least five classes of scaffold proteins contribute to the precise orchestration of changes in synaptic strength: PSD-95 and other MAGUKS, SHANKs, Homer, AKAPs, and Densin. What we have learned thus far about the dynamic properties of the Postsynaptic Density reveals new ways to rapidly regulate the number and sensitivity of membrane receptors.
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The Role of the Postsynaptic Density and the SpineCytoskeleton in Synaptic Plasticity
Learning and Memory: A Comprehensive Reference, 2008Co-Authors: Edoardo Marcora, Holly J. Carlisle, Mary B KennedyAbstract:Glutamatergic Postsynaptic spines have a specialized cytoskeletal structure consisting of an elaborate submembrane scaffold attached to synaptic receptors, termed the Postsynaptic Density (PSD), and an actin-based cytoskeleton that maintains their mushroom-like shape. The PSD comprises a layered arrangement of scaffold proteins that organize signaling enzymes to respond to activation of N-methyl-d-aspartate-type and metabotropic glutamate receptors. At every level (receptors, proximal scaffolds, and distal scaffolds), effector proteins, regulatory enzymes, and adaptors are recruited into signal transduction modules that enable specialized adaptive functions. Signaling machinery in the PSD and cytoskeletal regulatory proteins work together to alter the structure of the spine to produce LTP or LTD.
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The Postsynaptic Density at glutamatergic synapses.
Trends in neurosciences, 1997Co-Authors: Mary B KennedyAbstract:The Postsynaptic Density (PSD) is a tiny, amorphous structure located beneath the Postsynaptic membrane of synapses in the CNS. Until recently, the molecular composition and function of the PSD were mostly matters of speculation. With the advent of powerful new microchemical tools and molecular-genetic methods, three new classes of proteins have been identified in the PSD at glutamatergic synapses: the PSD-95 family, the NR2B subunit of the NMDA-type glutamate receptor, and densin-180. The PSD-95 family is involved in clustering of NMDA receptors. NR2B is phosphorylated by Ca^(2+)-calmodulin-dependent protein kinase type II, a prominent constituent of the PSD. Densin-180 might represent a new class of synaptic adhesion molecule. Study of these molecules is beginning to reveal the functional significance of the PSD.
Thomas S. Reese - One of the best experts on this subject based on the ideXlab platform.
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FAM81A protein, a novel component of the Postsynaptic Density in adult brain.
Neuroscience letters, 2019Co-Authors: Ayse Dosemeci, Thomas S. Reese, Hannah Loo, Dana Toy, Christine A. Winters, Jung-hwa Tao-chengAbstract:Abstract Analysis of affinity-purified PSD-95 complexes had previously identified a ‘hypothetical protein’, product of the gene FAM81A [ 1 ]. The present study examined the tissue and subcellular distribution of FAM81A protein and its expression levels during development. Comparison of different organs indicates selective expression of FAM81A protein in brain. FAM81A is expressed late in development, with a post-natal gradual increase in brain levels that parallels the expression of PSD-95. Comparison of subcellular fractions from adult brain shows that the distribution of FAM81A protein is similar to that of PSD-95, with a drastic enrichment in the Postsynaptic Density fraction. Immuno-electron microscopy of adult brain tissue reveals specific immunogold labeling for FAM81A protein at Postsynaptic densities in the forebrain. The label for FAM81A protein is concentrated at the cytoplasmic edge of the electron-dense core of the Postsynaptic Density, with a mean distance of ∼33 nm from the Postsynaptic membrane. These observations firmly establish FAM81A protein as a component of the Postsynaptic Density in the adult brain, suggesting a role in synaptic function.
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The Postsynaptic Density: There Is More than Meets the Eye.
Frontiers in synaptic neuroscience, 2016Co-Authors: Ayse Dosemeci, Thomas S. Reese, Jung-hwa Tao-cheng, Richard J WeinbergAbstract:The Postsynaptic Density (PSD), apparent in electron micrographs as a dense lamina just beneath the Postsynaptic membrane, includes a deeper layer, the “pallium,” containing a scaffold of Shank and Homer proteins. Though poorly defined in traditionally prepared thin-section electron micrographs, the pallium becomes denser and more conspicuous during intense synaptic activity, due to the reversible addition of CaMKII and other proteins. In this Perspective we review the significance of CaMKII-mediated recruitment of proteins to the pallium with respect to both the trafficking of receptors and the remodeling of spine shape that follow synaptic stimulation. We suggest that the level and duration of CaMKII translocation and activation in the pallium will shape activity-induced changes in the spine.
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zinc stabilizes shank3 at the Postsynaptic Density of hippocampal synapses
PLOS ONE, 2016Co-Authors: Junghwa Taocheng, Thomas S. Reese, Dana Toy, Christine A. Winters, Ayse DosemeciAbstract:Shank3 is a Postsynaptic Density (PSD) scaffold protein of the Shank family. Here we use pre-embedding immunogold electron microscopy to investigate factors influencing the distribution of Shank3 at the PSD. In dissociated rat hippocampal cultures under basal conditions, label for Shank3 was concentrated in a broad layer of the PSD, ~20–80 nm from the Postsynaptic membrane. Upon depolarization with high K+ (90 mM, 2 min), or application of NMDA (50 μM, 2 min), both the labeling intensity at the PSD and the median distance of label from the Postsynaptic membrane increased significantly, indicating that Shank3 molecules are preferentially recruited to the distal layer of the PSD. Incubation in medium supplemented with zinc (50 μM ZnCl2, 1 hr) also significantly increased labeling intensity for Shank3 at the PSD, but this addition of Shank3 was not preferential to the distal layer. When cells were incubated with zinc and then treated with NMDA, labeling intensity of Shank3 became higher than with either treatment alone and manifested a preference for the distal layer of the PSD. Without zinc supplementation, NMDA-induced accumulation of Shank3 at the PSD was transient, reversing within 30 min after return to control medium. However, when zinc was included in culture media throughout the experiment, the NMDA-induced accumulation of Shank3 was largely retained, including Shank3 molecules recruited to the distal layer of the PSD. These results demonstrate that activity induces accumulation of Shank3 at the PSD and that zinc stabilizes PSD-associated Shank3, possibly through strengthening of Shank-Shank association.
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differential distribution of shank and gkap at the Postsynaptic Density
PLOS ONE, 2015Co-Authors: Junghwa Taocheng, Yijung Yang, Thomas S. Reese, Ayse DosemeciAbstract:Shank and GKAP are scaffold proteins and binding partners at the Postsynaptic Density (PSD). The distribution and dynamics of Shank and GKAP were studied in dissociated hippocampal cultures by pre-embedding immunogold electron microscopy. Antibodies against epitopes containing their respective mutual binding sites were used to verify the expected juxtapositioning of Shank and GKAP. If all Shank and GKAP molecules at the PSD were bound to each other, the distribution of label for the two proteins should coincide. However, labels for the mutual binding sites showed significant differences in distribution, with a narrow distribution for GKAP located close to the Postsynaptic membrane, and a wider distribution for Shank extending deeper into the cytoplasm. Upon depolarization with high K+, neither the intensity nor distribution of label for GKAP changed, but labeling intensity for Shank at the PSD increased to ~150% of controls while the median distance of label from Postsynaptic membrane increased by 7.5 nm. These results indicate a preferential recruitment of Shank to more distal parts of the PSD complex. Conversely, upon incubation in Ca2+-free medium containing EGTA, the labeling intensity of Shank at the PSD decreased to ~70% of controls and the median distance of label from Postsynaptic membrane decreased by 9 nm, indicating a preferential loss of Shank molecules in more distal parts of the PSD complex. These observations identify two pools of Shank at the PSD complex, one relatively stable pool, closer to the Postsynaptic membrane that can bind to GKAP, and another more dynamic pool at a location too far away to bind to GKAP.
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camkii mediates recruitment and activation of the deubiquitinase cyld at the Postsynaptic Density
PLOS ONE, 2014Co-Authors: Soe Thein, Thomas S. Reese, Junghwa Taocheng, Ulrich K Bayer, Ayse DosemeciAbstract:NMDA treatment of cultured hippocampal neurons causes recruitment of CYLD, as well as CaMKII, to the Postsynaptic Density (PSD), as shown by immunoelectron microscopy. Recruitment of CYLD, a deubiquitinase specific for K63-linked polyubiquitins, is blocked by pre-treatment with tatCN21, a CaMKII inhibitor, at a concentration that inhibits the translocation of CaMKII to the PSD. Furthermore, CaMKII co-immunoprecipitates with CYLD from solubilized PSD fractions, indicating an association between the proteins. Purified CaMKII phosphorylates CYLD on at least three residues (S-362, S-418, and S-772 on the human CYLD protein Q9NQC7-1) and promotes its deubiquitinase activity. Activation of CaMKII in isolated PSDs promotes phosphorylation of CYLD on the same residues and also enhances endogenous deubiquitinase activity specific for K63-linked polyubiquitins. Since K63-linked polyubiquitin conjugation to proteins inhibits their interaction with proteasomes, CaMKII-mediated recruitment and upregulation of CYLD is expected to remove K63-linked polyubiquitins and facilitate proteasomal degradation at the PSD.
Tobias M. Boeckers - One of the best experts on this subject based on the ideXlab platform.
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in depth protein profiling of the Postsynaptic Density from mouse hippocampus using data independent acquisition proteomics
Proteomics, 2014Co-Authors: Ute Distler, Tobias M. Boeckers, Michael J. Schmeisser, Assunta Pelosi, Dominik Reim, Jorg Kuharev, Roland Weiczner, Jan Baumgart, Robert Nitsch, Johannes VogtAbstract:Located at neuronal terminals, the Postsynaptic Density (PSD) is a highly complex network of cytoskeletal scaffolding and signaling proteins responsible for the transduction and modulation of glutamatergic signaling between neurons. Using ion-mobility enhanced data-independent label-free LC-MS/MS, we established a reference proteome of crude synaptosomes, synaptic junctions, and PSD derived from mouse hippocampus including TOP3-based absolute quantification values for identified proteins. The final dataset across all fractions comprised 49 491 peptides corresponding to 4558 protein groups. Of these, 2102 protein groups were identified in highly purified PSD in at least two biological replicates. Identified proteins play pivotal roles in neurological and synaptic processes providing a rich resource for studies on hippocampal PSD function as well as on the pathogenesis of neuropsychiatric disorders. All MS data have been deposited in the ProteomeXchange with identifier PXD000590 (http://proteomecentral.proteomexchange.org/dataset/PXD000590).
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Scaffold Proteins at the Postsynaptic Density
Advances in experimental medicine and biology, 2012Co-Authors: Chiara Verpelli, Michael J. Schmeisser, Carlo Sala, Tobias M. BoeckersAbstract:Scaffold proteins are abundant and essential components of the Postsynaptic Density (PSD). They play a major role in many synaptic functions including the trafficking, anchoring, and clustering of glutamate receptors and adhesion molecules. Moreover, they link Postsynaptic receptors with their downstream signaling proteins and regulate the dynamics of cytoskeletal structures. By definition, PSD scaffold proteins do not have intrinsic enzymatic activities but are formed by modular and specific domains deputed to form large protein networks. Here, we will discuss the latest findings regarding the structure and functions of major PSD scaffold proteins.
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The Postsynaptic Density.
Cell and tissue research, 2006Co-Authors: Tobias M. BoeckersAbstract:Glutamatergic synapses in the central nervous system are characterized by an electron-dense web underneath the Postsynaptic membrane; this web is called the Postsynaptic Density (PSD). PSDs are composed of a dense network of several hundred proteins, creating a macromolecular complex that serves a wide range of functions. Prominent PSD proteins such as members of the MaGuk or ProSAP/Shank family build up a dense scaffold that creates an interface between clustered membrane-bound receptors, cell adhesion molecules and the actin-based cytoskeleton. Moreover, kinases, phosphatases and several proteins of different signalling pathways are specifically localized within the spine/PSD compartment. Small GTPases and regulating proteins are also enriched in PSDs being the molecular basis for regulated structural changes of cytoskeletal components within the synapse in response to external or internal stimuli, e.g. synaptic activation. This synaptic rearrangement (structural plasticity) is a rapid process and is believed to underlie learning and memory formation. The characterization of synapse/PSD proteins is especially important in the light of recent data suggesting that several mental disorders have their molecular defect at the synapse/PSD level.
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A role for zinc in Postsynaptic Density asSAMbly and plasticity
Trends in biochemical sciences, 2006Co-Authors: Eckart D Gundelfinger, Tobias M. Boeckers, Marisa K Baron, James U. BowieAbstract:Chemical synapses are asymmetric cell junctions that mediate communication between neurons. Multidomain scaffolding proteins of the Shank family act as major organizing elements of the ‘Postsynaptic Density' – that is, the cytoskeletal protein matrix associated with the Postsynaptic membrane. A recent study has shown that the C-terminal sterile α-motif or ‘SAM domain' of Shank3 (also known as ProSAP2) can form two-dimensional sheets of helical fibers. Assembly and packaging of these fibers are markedly enhanced by the presence of Zn 2+ ions. Zn 2+ can be released together with glutamate from synaptic vesicles and can enter the Postsynaptic cell through specific ionotropic receptors. Based on these observations, we propose a new model of synaptic plasticity in which Zn 2+ influx directly and instantly modulates the structure and function of the Postsynaptic Density.
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an architectural framework that may lie at the core of the Postsynaptic Density
Science, 2006Co-Authors: Marisa K Baron, Tobias M. Boeckers, Eckart D Gundelfinger, Mari Gingery, Salem Faham, Bianca Vaida, Michael R Sawaya, Danielle Salyer, James U. BowieAbstract:The Postsynaptic Density (PSD) is a complex assembly of proteins associated with the Postsynaptic membrane that organizes neurotransmitter receptors, signaling pathways, and regulatory elements within a cytoskeletal matrix. Here we show that the sterile alpha motif domain of rat Shank3/ProSAP2, a master scaffolding protein located deep within the PSD, can form large sheets composed of helical fibers stacked side by side. Zn2+, which is found in high concentrations in the PSD, binds tightly to Shank3 and may regulate assembly. Sheets of the Shank protein could form a platform for the construction of the PSD complex.
Junghwa Taocheng - One of the best experts on this subject based on the ideXlab platform.
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stimulation induces gradual increases in the thickness and curvature of Postsynaptic Density of hippocampal ca1 neurons in slice cultures
Molecular Brain, 2019Co-Authors: Junghwa TaochengAbstract:Activity can induce structural changes in glutamatergic excitatory synapses, including increase in thickness and curvature of the Postsynaptic Density (PSD); these structural changes can only be documented by electron microscopy. Here in organotypic hippocampal slice cultures where experimental conditions can be easily manipulated, increases in thickness and curvature of PSDs were noticeable within 30 s of stimulation and progressed with time up to 3 min. These structural changes were reversible upon returning the samples to control medium for 5–10 min. Thus, the Postsynaptic Density is a very dynamic structure that undergoes rapid reorganization of its components upon stimulation, and recovery upon cessation of stimulation. The gradual increase in thickness of PSD could result from a gradual translocation of some PSD proteins to the PSD, and the increase in curvature of the PSD is likely led by Postsynaptic elements.
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irsp53 accumulates at the Postsynaptic Density under excitatory conditions
PLOS ONE, 2017Co-Authors: Ayse Dosemeci, Hannah Loo, Dana Toy, Junghwa Taocheng, Amelia BurchAbstract:IRSp53 (BAIAP2) is an abundant protein at the Postsynaptic Density (PSD) that binds to major PSD scaffolds, PSD-95 and Shanks, as well as to F-actin. The distribution of IRSp53 at the PSD in cultured hippocampal neurons was examined under basal and excitatory conditions by immuno-electron microscopy. Under basal conditions, label for IRSp53 is concentrated at the PSD. Upon depolarization by application of a medium containing 90 mM K+, the intensity of IRSp53 label at the PSD increased by 36±7%. Application of NMDA (50 μM) yielded 53±1% increase in the intensity of IRSp53 label at the PSD compared to controls treated with APV, an NMDA antagonist. The accumulation of IRSp53 label upon application of high K+ or NMDA was prominent at the deeper region of the PSD (the PSD pallium, lying 40–120 nm from the Postsynaptic plasma membrane). IRSp53 molecules that accumulate at the distal region of the PSD pallium under excitatory conditions are too far from the plasma membrane to fulfill the generally recognized role of the protein as an effector of membrane-bound small GTPases. Instead, these IRSp53 molecules may have a structural role organizing the Shank scaffold and/or linking the PSD to the actin cytoskeleton.
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camkii mediated displacement of aida 1 out of the Postsynaptic Density core
FEBS Letters, 2016Co-Authors: Ayse Dosemeci, Dana Toy, Junghwa Taocheng, Amelia Burch, Ulrich K BayerAbstract:Ankyrin repeat and sterile alpha motif domain-containing protein 1B (ANKS1B, also known as AIDA-1) is a major component of the Postsynaptic Density (PSD) in excitatory neurons where it concentrates at the electron-dense core under basal conditions and moves out during activity. This study investigates the molecular mechanism underlying activity-induced displacement of AIDA-1. Experiments with PSD fractions from brain indicate phosphorylation of AIDA-1 upon activation of endogenous CaMKII. Immuno-electron microscopy studies show that treatment of hippocampal neurons with NMDA results in an ~ 30 nm shift in the median distance of the AIDA-1 label from the Postsynaptic membrane, an effect that is blocked by the CaMKII inhibitor tatCN21. CaMKII-mediated redistribution of AIDA-1 is similar to that observed for SynGAP. CaMKII-mediated removal of two abundant PSD-95-binding proteins from the PSD core during activity is expected to initiate a molecular reorganization at the PSD.
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zinc stabilizes shank3 at the Postsynaptic Density of hippocampal synapses
PLOS ONE, 2016Co-Authors: Junghwa Taocheng, Thomas S. Reese, Dana Toy, Christine A. Winters, Ayse DosemeciAbstract:Shank3 is a Postsynaptic Density (PSD) scaffold protein of the Shank family. Here we use pre-embedding immunogold electron microscopy to investigate factors influencing the distribution of Shank3 at the PSD. In dissociated rat hippocampal cultures under basal conditions, label for Shank3 was concentrated in a broad layer of the PSD, ~20–80 nm from the Postsynaptic membrane. Upon depolarization with high K+ (90 mM, 2 min), or application of NMDA (50 μM, 2 min), both the labeling intensity at the PSD and the median distance of label from the Postsynaptic membrane increased significantly, indicating that Shank3 molecules are preferentially recruited to the distal layer of the PSD. Incubation in medium supplemented with zinc (50 μM ZnCl2, 1 hr) also significantly increased labeling intensity for Shank3 at the PSD, but this addition of Shank3 was not preferential to the distal layer. When cells were incubated with zinc and then treated with NMDA, labeling intensity of Shank3 became higher than with either treatment alone and manifested a preference for the distal layer of the PSD. Without zinc supplementation, NMDA-induced accumulation of Shank3 at the PSD was transient, reversing within 30 min after return to control medium. However, when zinc was included in culture media throughout the experiment, the NMDA-induced accumulation of Shank3 was largely retained, including Shank3 molecules recruited to the distal layer of the PSD. These results demonstrate that activity induces accumulation of Shank3 at the PSD and that zinc stabilizes PSD-associated Shank3, possibly through strengthening of Shank-Shank association.
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differential distribution of shank and gkap at the Postsynaptic Density
PLOS ONE, 2015Co-Authors: Junghwa Taocheng, Yijung Yang, Thomas S. Reese, Ayse DosemeciAbstract:Shank and GKAP are scaffold proteins and binding partners at the Postsynaptic Density (PSD). The distribution and dynamics of Shank and GKAP were studied in dissociated hippocampal cultures by pre-embedding immunogold electron microscopy. Antibodies against epitopes containing their respective mutual binding sites were used to verify the expected juxtapositioning of Shank and GKAP. If all Shank and GKAP molecules at the PSD were bound to each other, the distribution of label for the two proteins should coincide. However, labels for the mutual binding sites showed significant differences in distribution, with a narrow distribution for GKAP located close to the Postsynaptic membrane, and a wider distribution for Shank extending deeper into the cytoplasm. Upon depolarization with high K+, neither the intensity nor distribution of label for GKAP changed, but labeling intensity for Shank at the PSD increased to ~150% of controls while the median distance of label from Postsynaptic membrane increased by 7.5 nm. These results indicate a preferential recruitment of Shank to more distal parts of the PSD complex. Conversely, upon incubation in Ca2+-free medium containing EGTA, the labeling intensity of Shank at the PSD decreased to ~70% of controls and the median distance of label from Postsynaptic membrane decreased by 9 nm, indicating a preferential loss of Shank molecules in more distal parts of the PSD complex. These observations identify two pools of Shank at the PSD complex, one relatively stable pool, closer to the Postsynaptic membrane that can bind to GKAP, and another more dynamic pool at a location too far away to bind to GKAP.