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Chapsyn-110/PSD-93 antibody

RRID:AB_2277296

Antibody ID

AB_2277296

Target Antigen

Chapsyn-110/PSD-93 null

Proper Citation

(UC Davis/NIH NeuroMab Facility Cat# 75-057, RRID:AB_2277296)

Clonality

monoclonal antibody

Comments

Originating manufacturer of this product. Applications: IB, ICC, IHC, IP, KO, WB. Validation status: IF or IB (Pass), IB in brain (Pass), IHC in brain (Pass), KO (Pass).

Clone ID

N18/30

Host Organism

mouse

Loss of SynDIG1 Reduces Excitatory Synapse Maturation But Not Formation In Vivo.

  • Chenaux G
  • eNeuro
  • 2017 Oct 31

Literature context:


Abstract:

Modification of the strength of excitatory synaptic connections is a fundamental mechanism by which neural circuits are refined during development and learning. Synapse Differentiation Induced Gene 1 (SynDIG1) has been shown to play a key role in regulating synaptic strength in vitro. Here, we investigated the role of SynDIG1 in vivo in mice with a disruption of the SynDIG1 gene rather than use an alternate loxP-flanked conditional mutant that we find retains a partial protein product. The gene-trap insertion with a reporter cassette mutant mice shows that the SynDIG1 promoter is active during embryogenesis in the retina with some activity in the brain, and postnatally in the mouse hippocampus, cortex, hindbrain, and spinal cord. Ultrastructural analysis of the hippocampal CA1 region shows a decrease in the average PSD length of synapses and a decrease in the number of synapses with a mature phenotype. Intriguingly, the total synapse number appears to be increased in SynDIG1 mutant mice. Electrophysiological analyses show a decrease in AMPA and NMDA receptor function in SynDIG1-deficient hippocampal neurons. Glutamate stimulation of individual dendritic spines in hippocampal slices from SynDIG1-deficient mice reveals increased short-term structural plasticity. Notably, the overall levels of PSD-95 or glutamate receptors enriched in postsynaptic biochemical fractions remain unaltered; however, activity-dependent synapse development is strongly compromised upon the loss of SynDIG1, supporting its importance for excitatory synapse maturation. Together, these data are consistent with a model in which SynDIG1 regulates the maturation of excitatory synapse structure and function in the mouse hippocampus in vivo.

Funding information:
  • NIMH NIH HHS - R01 MH104638(United States)

Hierarchical organization and genetically separable subfamilies of PSD95 postsynaptic supercomplexes.

  • Frank RAW
  • J. Neurochem.
  • 2017 Sep 7

Literature context:


Abstract:

PSD95 is an abundant postsynaptic scaffold protein in glutamatergic synapses that assembles into supercomplexes composed of over 80 proteins including neurotransmitter receptors, ion channels and adhesion proteins. How these diverse constituents are organized into PSD95 supercomplexes in vivo is poorly understood. Here, we dissected the supercomplexes in mice combining endogenous gene-tagging, targeted mutations and quantitative biochemical assays. Generating compound heterozygous mice with two different gene-tags, one on each Psd95 allele, showed that each ~1.5 MDa PSD95-containing supercomplex contains on average two PSD95 molecules. Gene-tagging the endogenous GluN1 and PSD95 with identical Flag tags revealed N-methyl D-aspartic acid receptors (NMDARs) containing supercomplexes that represent only 3% of the total population of PSD95 supercomplexes, suggesting there are many other subtypes. To determine whether this extended population of different PSD95 supercomplexes use genetically defined mechanisms to specify their assembly, we tested the effect of five targeted mouse mutations on the assembly of known PSD95 interactors, Kir2.3, Arc, IQsec2/BRAG1 and Adam22. Unexpectedly, some mutations were highly selective, whereas others caused widespread disruption, indicating that PSD95 interacting proteins are organized hierarchically into distinct subfamilies of ~1.5 MDa supercomplexes, including a subpopulation of Kir2.3-NMDAR ion channel-channel supercomplexes. Kir2.3-NMDAR ion channel-channel supercomplexes were found to be anatomically restricted to particular brain regions. These data provide new insight into the mechanisms that govern the constituents of postsynaptic supercomplexes and the diversity of synapse types. Read the Editorial Highlight for this article on page 500. Cover Image for this issue: doi. 10.1111/jnc.13811.

Membrane-associated guanylate kinase scaffolds organize a horizontal cell synaptic complex restricted to invaginating contacts with photoreceptors.

  • Vila A
  • J. Comp. Neurol.
  • 2017 Mar 1

Literature context:


Abstract:

Synaptic processes and plasticity of synapses are mediated by large suites of proteins. In most cases, many of these proteins are tethered together by synaptic scaffold proteins. Scaffold proteins have a large number and typically a variety of protein interaction domains that allow many different proteins to be assembled into functional complexes. Because each scaffold protein has a different set of protein interaction domains and a unique set of interacting partners, the presence of synaptic scaffolds can provide insight into the molecular mechanisms that regulate synaptic processes. In studies of rabbit retina, we found SAP102 and Chapsyn110 selectively localized in the tips of B-type horizontal cell processes, where they contact cone and rod photoreceptors. We further identified some known SAP102 binding partners, kainate receptor GluR6/7 and inward rectifier potassium channel Kir2.1, closely associated with SAP102 in photoreceptor invaginations. The kainate receptor occupies a position distinct from that of the majority of AMPA receptors that dominate the horizontal cell postsynaptic response. GluR6/7 and Kir2.1 presumably are involved in synaptic processes that govern cell-to-cell communication and could both contribute in different ways to synaptic currents that mediate feedback signaling. Notably, we failed to find evidence for the presence of Cx57 or Cx59 that might be involved in ephaptic feedback signaling in this complex. The presence of SAP102 and its binding partners in both cone and rod invaginating synapses suggests that whatever mechanism is supported by this protein complex is present in both types of photoreceptors. J. Comp. Neurol. 525:850-867, 2017. © 2016 Wiley Periodicals, Inc.

Distribution of the SynDIG4/proline-rich transmembrane protein 1 in rat brain.

  • Kirk LM
  • J. Comp. Neurol.
  • 2016 Aug 1

Literature context:


Abstract:

The modulation of AMPA receptor (AMPAR) content at synapses is thought to be an underlying molecular mechanism of memory and learning. AMPAR content at synapses is highly plastic and is regulated by numerous AMPAR accessory transmembrane proteins such as TARPs, cornichons, and CKAMPs. SynDIG (synapse differentiation-induced gene) defines a family of four genes (SynDIG1-4) expressed in distinct and overlapping patterns in the brain. SynDIG1 was previously identified as a novel transmembrane AMPAR-associated protein that regulates synaptic strength. The related protein SynDIG4 [also known as Prrt1 (proline-rich transmembrane protein 1)] has recently been identified as a component of AMPAR complexes. In this study, we show that SynDIG1 and SynDIG4 have distinct yet overlapping patterns of expression in the central nervous system, with SynDIG4 having especially prominent expression in the hippocampus and particularly within CA1. In contrast to SynDIG1 and other traditional AMPAR auxiliary subunits, SynDIG4 is de-enriched at the postsynaptic density and colocalizes with extrasynaptic GluA1 puncta in primary dissociated neuron culture. These results indicate that, although SynDIG4 shares sequence similarity with SynDIG1, it might act through a unique mechanism as an auxiliary factor for extrasynaptic GluA1-containing AMPARs. J. Comp. Neurol. 524:2266-2280, 2016. © 2015 Wiley Periodicals, Inc.

Synaptic state-dependent functional interplay between postsynaptic density-95 and synapse-associated protein 102.

  • Bonnet SA
  • J. Neurosci.
  • 2013 Aug 14

Literature context:


Abstract:

Activity-dependent regulation of AMPA receptor (AMPAR)-mediated synaptic transmission is the basis for establishing differences in synaptic weights among individual synapses during developmental and experience-dependent synaptic plasticity. Synaptic signaling scaffolds of the Discs large (DLG)-membrane-associated guanylate kinase (MAGUK) protein family regulate these processes by tethering signaling proteins to receptor complexes. Using a molecular replacement strategy with RNAi-mediated knockdown in rat and mouse hippocampal organotypic slice cultures, a postsynaptic density-95 (PSD-95) knock-out mouse line and electrophysiological analysis, our current study identified a functional interplay between two paralogs, PSD-95 and synapse-associated protein 102 (SAP102) to regulate synaptic AMPARs. During synaptic development, the SAP102 protein levels normally plateau but double if PSD-95 expression is prevented during synaptogenesis. For an autonomous function of PSD-95 in regulating synaptic AMPARs, in addition to the previously demonstrated N-terminal multimerization and the first two PDZ (PSD-95, Dlg1, zona occludens-1) domains, the PDZ3 and guanylate kinase domains were required. The Src homology 3 domain was dispensable for the PSD-95-autonomous regulation of basal synaptic transmission. However, it mediated the functional interaction with SAP102 of PSD-95 mutants to enhance AMPARs. These results depict a protein domain-based multifunctional aspect of PSD-95 in regulating excitatory synaptic transmission and unveil a novel form of domain-based interplay between signaling scaffolds of the DLG-MAGUK family.

Funding information:
  • Wellcome Trust - 077046(United Kingdom)

Eye opening and PSD95 are required for long-term potentiation in developing superior colliculus.

  • Zhao JP
  • Proc. Natl. Acad. Sci. U.S.A.
  • 2013 Jan 8

Literature context:


Abstract:

The only major glutamate receptor membrane-associated guanylate kinase scaffolds expressed in the young superficial superior colliculus (SC) are synapse-associated protein 102 (SAP102) and postsynaptic density protein 95 (PSD95). In this, as in all visual brain regions examined, synaptic PSD95 increases rapidly following simultaneous eyelid opening (EO). We show that EO and PSD95 are necessary for SC NMDA receptor (NMDAR)-dependent long-term potentiation (LTP) and this LTP is eliminated or reinstated by manipulating EO. PSD95 knockdown (KD) in vivo blocks this LTP, but not long-term depression, and reduces frequencies of miniature AMPA receptor and NMDAR currents with no change in presynaptic release. Furthermore, miniature NMDAR currents after PSD95 KD show an activity-triggered calcineurin sensitivity that is normally only found in the pre-EO period when SAP102 binds mixed GluN2A/GluN2B NMDARs. These data indicate that young SC LTP arises from PSD95 unsilencing of silent synapses, that unsilencing is labile in young brain, and that even though SAP102 and PSD95 can bind the same NMDARs, only PSD95 enables SC synaptic maturation.

Funding information:
  • NIGMS NIH HHS - GM070064(United States)

PSD-95 is post-transcriptionally repressed during early neural development by PTBP1 and PTBP2.

  • Zheng S
  • Nat. Neurosci.
  • 2012 Jan 15

Literature context:


Abstract:

Postsynaptic density protein 95 (PSD-95) is essential for synaptic maturation and plasticity. Although its synaptic regulation has been widely studied, the control of PSD-95 cellular expression is not understood. We found that Psd-95 was controlled post-transcriptionally during neural development. Psd-95 was transcribed early in mouse embryonic brain, but most of its product transcripts were degraded. The polypyrimidine tract binding proteins PTBP1 and PTBP2 repressed Psd-95 (also known as Dlg4) exon 18 splicing, leading to premature translation termination and nonsense-mediated mRNA decay. The loss of first PTBP1 and then of PTBP2 during embryonic development allowed splicing of exon 18 and expression of PSD-95 late in neuronal maturation. Re-expression of PTBP1 or PTBP2 in differentiated neurons inhibited PSD-95 expression and impaired the development of glutamatergic synapses. Thus, expression of PSD-95 during early neural development is controlled at the RNA level by two PTB proteins whose sequential downregulation is necessary for synapse maturation.

Funding information:
  • NIDCR NIH HHS - R01 DE023090(United States)

Characterization of the axon initial segment (AIS) of motor neurons and identification of a para-AIS and a juxtapara-AIS, organized by protein 4.1B.

  • Duflocq A
  • BMC Biol.
  • 2011 Sep 29

Literature context:


Abstract:

BACKGROUND: The axon initial segment (AIS) plays a crucial role: it is the site where neurons initiate their electrical outputs. Its composition in terms of voltage-gated sodium (Nav) and voltage-gated potassium (Kv) channels, as well as its length and localization determine the neuron's spiking properties. Some neurons are able to modulate their AIS length or distance from the soma in order to adapt their excitability properties to their activity level. It is therefore crucial to characterize all these parameters and determine where the myelin sheath begins in order to assess a neuron's excitability properties and ability to display such plasticity mechanisms. If the myelin sheath starts immediately after the AIS, another question then arises as to how would the axon be organized at its first myelin attachment site; since AISs are different from nodes of Ranvier, would this particular axonal region resemble a hemi-node of Ranvier? RESULTS: We have characterized the AIS of mouse somatic motor neurons. In addition to constant determinants of excitability properties, we found heterogeneities, in terms of AIS localization and Nav composition. We also identified in all α motor neurons a hemi-node-type organization, with a contactin-associated protein (Caspr)+ paranode-type, as well as a Caspr2+ and Kv1+ juxtaparanode-type compartment, referred to as a para-AIS and a juxtapara (JXP)-AIS, adjacent to the AIS, where the myelin sheath begins. We found that Kv1 channels appear in the AIS, para-AIS and JXP-AIS concomitantly with myelination and are progressively excluded from the para-AIS. Their expression in the AIS and JXP-AIS is independent from transient axonal glycoprotein-1 (TAG-1)/Caspr2, in contrast to juxtaparanodes, and independent from PSD-93. Data from mice lacking the cytoskeletal linker protein 4.1B show that this protein is necessary to form the Caspr+ para-AIS barrier, ensuring the compartmentalization of Kv1 channels and the segregation of the AIS, para-AIS and JXP-AIS. CONCLUSIONS: α Motor neurons have heterogeneous AISs, which underlie different spiking properties. However, they all have a para-AIS and a JXP-AIS contiguous to their AIS, where the myelin sheath begins, which might limit some AIS plasticity. Protein 4.1B plays a key role in ensuring the proper molecular compartmentalization of this hemi-node-type region.

Funding information:
  • NCI NIH HHS - CA173903(United States)
  • NIGMS NIH HHS - 2R01 GM063891(United States)

Cdk-mediated phosphorylation of the Kvβ2 auxiliary subunit regulates Kv1 channel axonal targeting.

  • Vacher H
  • J. Cell Biol.
  • 2011 Mar 7

Literature context:


Abstract:

Kv1 channels are concentrated at specific sites in the axonal membrane, where they regulate neuronal excitability. Establishing these distributions requires regulated dissociation of Kv1 channels from the neuronal trafficking machinery and their subsequent insertion into the axonal membrane. We find that the auxiliary Kvβ2 subunit of Kv1 channels purified from brain is phosphorylated on serine residues 9 and 31, and that cyclin-dependent kinase (Cdk)-mediated phosphorylation at these sites negatively regulates the interaction of Kvβ2 with the microtubule plus end-tracking protein EB1. Endogenous Cdks, EB1, and Kvβ2 phosphorylated at serine 31 are colocalized in the axons of cultured hippocampal neurons, with enrichment at the axon initial segment (AIS). Acute inhibition of Cdk activity leads to intracellular accumulation of EB1, Kvβ2, and Kv1 channel subunits within the AIS. These studies reveal a new regulatory mechanism for the targeting of Kv1 complexes to the axonal membrane through the reversible Cdk phosphorylation-dependent binding of Kvβ2 to EB1.

Funding information:
  • NEI NIH HHS - N01-EY-5-0007(United States)

TARP phosphorylation regulates synaptic AMPA receptors through lipid bilayers.

  • Sumioka A
  • Neuron
  • 2010 Jun 10

Literature context:


Abstract:

Neurons use neurotransmitters to communicate across synapses, constructing neural circuits in the brain. AMPA-type glutamate receptors are the predominant excitatory neurotransmitter receptors mediating fast synaptic transmission. AMPA receptors localize at synapses by forming protein complexes with transmembrane AMPA receptor regulatory proteins (TARPs) and PSD-95-like membrane-associated guanylate kinases. Among the three classes of ionotropic glutamate receptors (AMPA, NMDA, and kainate type), AMPA receptor activity is most regulatable by neuronal activity to adjust synaptic strength. Here, we mutated the prototypical TARP, stargazin, and found that TARP phosphorylation regulates synaptic AMPA receptor activity in vivo. We also found that stargazin interacts with negatively charged lipid bilayers in a phosphorylation-dependent manner and that the lipid interaction inhibited stargazin binding to PSD-95. Cationic lipids dissociated stargazin from lipid bilayers and enhanced synaptic AMPA receptor activity in a stargazin phosphorylation-dependent manner. Thus, TARP phosphorylation plays a critical role in regulating AMPA receptor-mediated synaptic transmission via a lipid bilayer interaction.

Funding information:
  • NCRR NIH HHS - M01-RR00084(United States)

Synaptic localization and function of Sidekick recognition molecules require MAGI scaffolding proteins.

  • Yamagata M
  • J. Neurosci.
  • 2010 Mar 10

Literature context:


Abstract:

Four transmembrane adhesion molecules-Sidekick-1, Sidekick-2, Down's syndrome cell adhesion molecule (Dscam), and Dscam-like-are determinants of lamina-specific synapse formation in the vertebrate retina. Their C termini are predicted to bind postsynaptic density (PSD)-95/Discs Large/ZO-1 (PDZ) domains, which are present in many synaptic scaffolding proteins. We identify members of the membrane-associated guanylate kinase with inverted orientation (MAGI) and PSD-95 subfamilies of multi-PDZ domain proteins as binding partners for Sidekicks and Dscams. Specific MAGI and PSD-95 family members are present in distinct subsets of retinal synapses, as are Sidekicks and Dscams. Using Sidekick-2 as an exemplar, we show that its PDZ-binding C terminus is required for both its synaptic localization in photoreceptors and its ability to promote lamina-specific arborization of presynaptic and postsynaptic processes in the inner plexiform layer. In photoreceptor synapses that contain both MAGI-1 and PSD-95, Sidekick-2 preferentially associates with MAGI-1. Depletion of MAGI-1 from photoreceptors by RNA interference blocks synaptic localization of Sidekick-2 in photoreceptors without affecting localization of PSD-95. Likewise, depletion of MAGI-2 from retinal ganglion cells and interneurons interferes with Sidekick-2-dependent laminar targeting of processes. These results demonstrate that localization and function of Sidekick-2 require its incorporation into a MAGI-containing synaptic scaffold.

ADAM22, a Kv1 channel-interacting protein, recruits membrane-associated guanylate kinases to juxtaparanodes of myelinated axons.

  • Ogawa Y
  • J. Neurosci.
  • 2010 Jan 20

Literature context:


Abstract:

Clustered Kv1 K(+) channels regulate neuronal excitability at juxtaparanodes of myelinated axons, axon initial segments, and cerebellar basket cell terminals (BCTs). These channels are part of a larger protein complex that includes cell adhesion molecules and scaffolding proteins. To identify proteins that regulate assembly, clustering, and/or maintenance of axonal Kv1 channel protein complexes, we immunoprecipitated Kv1.2 alpha subunits, and then used mass spectrometry to identify interacting proteins. We found that a disintegrin and metalloproteinase 22 (ADAM22) is a component of the Kv1 channel complex and that ADAM22 coimmunoprecipitates Kv1.2 and the membrane-associated guanylate kinases (MAGUKs) PSD-93 and PSD-95. When coexpressed with MAGUKs in heterologous cells, ADAM22 and Kv1 channels are recruited into membrane surface clusters. However, coexpression of Kv1.2 with ADAM22 and MAGUKs does not alter channel properties. Among all the known Kv1 channel-interacting proteins, only ADAM22 is found at every site where Kv1 channels are clustered. Analysis of Caspr-null mice showed that, like other previously described juxtaparanodal proteins, disruption of the paranodal junction resulted in redistribution of ADAM22 into paranodal zones. Analysis of Caspr2-, PSD-93-, PSD-95-, and double PSD-93/PSD-95-null mice showed ADAM22 clustering at BCTs requires PSD-95, but ADAM22 clustering at juxtaparanodes requires neither PSD-93 nor PSD-95. In direct contrast, analysis of ADAM22-null mice demonstrated juxtaparanodal clustering of PSD-93 and PSD-95 requires ADAM22, whereas Kv1.2 and Caspr2 clustering is normal in ADAM22-null mice. Thus, ADAM22 is an axonal component of the Kv1 K(+) channel complex that recruits MAGUKs to juxtaparanodes.

Activity-dependent anchoring of importin alpha at the synapse involves regulated binding to the cytoplasmic tail of the NR1-1a subunit of the NMDA receptor.

  • Jeffrey RA
  • J. Neurosci.
  • 2009 Dec 16

Literature context:


Abstract:

Synaptic plasticity, the capacity of neurons to change the strength of their connections with experience, provides a mechanism for learning and memory in the brain. Long-term plasticity requires new transcription, indicating that synaptically generated signals must be transported to the nucleus. Previous studies have described a role for importin nuclear transport adaptors in mediating the retrograde transport of signals from synapse to nucleus during plasticity. Here, we investigated the possibility that stimulus-induced translocation of importins from synapse to nucleus involves activity-dependent anchoring of importins at the synapse. We show that importin alpha binds to a nuclear localization signal (NLS) present in the cytoplasmic tail of NR1-1a. This interaction is disrupted by activation of NMDA receptors in cultured neurons and by stimuli that trigger late-phase, but not early-phase, long-term potentiation of CA3-CA1 synapses in acute hippocampal slices. In vitro PKC phosphorylation of GST-NR1-1a abolishes its ability to bind importin alpha in brain lysates, and the interaction of importin alpha and NR1 in neurons is modulated by PKC activity. Together, our results indicate that importin alpha is tethered at the postsynaptic density by binding to the NLS present in NR1-1a. This interaction is activity dependent, with importin alpha being released following NMDA receptor activation and phosphorylation rendering it available to bind soluble cargoes and transport them to the nucleus during transcription-dependent forms of neuronal plasticity.

NMDA di-heteromeric receptor populations and associated proteins in rat hippocampus.

  • Al-Hallaq RA
  • J. Neurosci.
  • 2007 Aug 1

Literature context:


Abstract:

Subunit composition of NMDA receptors (NMDARs) determines a range of physiological properties, downstream signaling effects, and binding partners. Differential localization of NR2A- or NR2B-containing NMDARs within the neuron and subunit-specific protein associations may explain differences in NR2A and NR2B contributions to synaptic plasticity and excitotoxic cell death. This question is complicated by the existence of tri-heteromeric complexes (NR1/NR2A/NR2B). To date, no quantitative biochemical determinations have been made of the relative abundance of different NMDAR populations in intact hippocampus, the region extensively correlated with NMDAR-dependent long-term potentiation. We investigated subunit composition and subunit-specific interactions in CA1/CA2 of rat hippocampus. Using sequential immunoprecipitations to deplete either NR2B or NR2A, di-heteromeric NR1/NR2A and NR1/NR2B receptor populations were isolated from postnatal day 7 (P7) hippocampus and P42 and 6-month-old CA1/CA2. Quantitative Western blot analysis revealed that 60-70% of NR2A and 70-85% of NR2B subunits were associated in NR1/NR2A or NR1/NR2B di-heteromeric complexes. Isolated di-heteromeric receptor fractions were used to examine NR2A- or NR2B-specific interactions with synapse-associated proteins. Our results indicate that NR2A- or NR2B-containing NMDARs associate similarly with postsynaptic density-95 (PSD-95), synapse-associated protein 102, and PSD-93 at P42. However, NR2A-containing receptors coimmunoprecipitated a greater proportion of the synaptic proteins neuronal nitric oxide synthase, Homer, and beta-catenin. Finally, mass spectrometry analysis of isolated di-heteromeric receptors identified a novel NMDAR interactor, collapsin response mediator protein 2, which preferentially associates with NR2B-containing di-heteromeric NMDARs. In summary, in rat hippocampus, NR2A and NR2B exist primarily in di-heteromeric complexes that interact similarly with PSD-95-related proteins but are associated with different protein complexes.

Funding information:
  • Wellcome Trust - 087618/Z/08/Z(United Kingdom)