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Pan-MAGUK scaffolding protein family antibody

RRID:AB_10673115

Antibody ID

AB_10673115

Target Antigen

Pan-MAGUK scaffolding protein family null

Proper Citation

(UC Davis/NIH NeuroMab Facility Cat# 75-029, RRID:AB_10673115)

Clonality

monoclonal antibody

Comments

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

Clone ID

K28/86

Host Organism

mouse

Developing high-quality mouse monoclonal antibodies for neuroscience research - approaches, perspectives and opportunities.

  • Gong B
  • N Biotechnol
  • 2016 Sep 25

Literature context:


Abstract:

High-quality antibodies (Abs) are critical to neuroscience research, as they remain the primary affinity proteomics reagent used to label and capture endogenously expressed protein targets in the nervous system. As in other fields, neuroscientists are frequently confronted with inaccurate and irreproducible Ab-based results and/or reporting. The UC Davis/NIH NeuroMab Facility was created with the mission of addressing the unmet need for high-quality Abs in neuroscience research by applying a unique approach to generate and validate mouse monoclonal antibodies (mAbs) optimized for use against mammalian brain (i.e., NeuroMabs). Here we describe our methodology of multi-step mAb screening focused on identifying mAbs exhibiting efficacy and specificity in labeling mammalian brain samples. We provide examples from NeuroMab screens, and from the subsequent specialized validation of those selected as NeuroMabs. We highlight the particular challenges and considerations of determining specificity for brain immunolabeling. We also describe why our emphasis on extensive validation of large numbers of candidates by immunoblotting and immunohistochemistry against brain samples is essential for identifying those that exhibit efficacy and specificity in those applications to become NeuroMabs. We describe the special attention given to candidates with less common non-IgG1 IgG subclasses that can facilitate simultaneous multiplex labeling with subclass-specific secondary antibodies. We detail our recent use of recombinant cloning of NeuroMabs as a method to archive all NeuroMabs, to unambiguously define NeuroMabs at the DNA sequence level, and to re-engineer IgG1 NeuroMabs to less common IgG subclasses to facilitate their use in multiplex labeling. Finally, we provide suggestions to facilitate Ab development and use, as to design, execution and interpretation of Ab-based neuroscience experiments. Reproducibility in neuroscience research will improve with enhanced Ab validation, unambiguous identification of Abs used in published experiments, and end user proficiency in Ab-based assays.

Tonic PKA Activity Regulates SK Channel Nanoclustering and Somatodendritic Distribution.

  • Abiraman K
  • J. Mol. Biol.
  • 2016 Jun 5

Literature context:


Abstract:

Small-conductance calcium-activated potassium (SK) channels mediate a potassium conductance in the brain and are involved in synaptic plasticity, learning, and memory. SK channels show a distinct subcellular localization that is crucial for their neuronal functions. However, the mechanisms that control this spatial distribution are unknown. We imaged SK channels labeled with fluorophore-tagged apamin and monitored SK channel nanoclustering at the single molecule level by combining atomic force microscopy and toxin (i.e., apamin) pharmacology. Using these two complementary approaches, we found that native SK channel distribution in pyramidal neurons, across the somatodendritic domain, depends on ongoing cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) levels, strongly limiting SK channel expression at the pyramidal neuron soma. Furthermore, tonic cAMP-PKA levels also controlled whether SK channels were expressed in nanodomains as single entities or as a group of multiple channels. Our study reveals a new level of regulation of SK channels by cAMP-PKA and suggests that ion channel topography and nanoclustering might be under the control of second messenger cascades.

Postsynaptic density scaffold SAP102 regulates cortical synapse development through EphB and PAK signaling pathway.

  • Murata Y
  • J. Neurosci.
  • 2013 Mar 13

Literature context:


Abstract:

Membrane-associated guanylate kinases (MAGUKs), including SAP102, PSD-95, PSD-93, and SAP97, are scaffolding proteins for ionotropic glutamate receptors at excitatory synapses. MAGUKs play critical roles in synaptic plasticity; however, details of signaling roles for each MAGUK remain largely unknown. Here we report that SAP102 regulates cortical synapse development through the EphB and PAK signaling pathways. Using lentivirus-delivered shRNAs, we found that SAP102 and PSD-95, but not PSD-93, are necessary for excitatory synapse formation and synaptic AMPA receptor (AMPAR) localization in developing mouse cortical neurons. SAP102 knockdown (KD) increased numbers of elongated dendritic filopodia, which is often observed in mouse models and human patients with mental retardation. Further analysis revealed that SAP102 coimmunoprecipitated the receptor tyrosine kinase EphB2 and RacGEF Kalirin-7 in neonatal cortex, and SAP102 KD reduced surface expression and dendritic localization of EphB. Moreover, SAP102 KD prevented reorganization of actin filaments, synapse formation, and synaptic AMPAR trafficking in response to EphB activation triggered by its ligand ephrinB. Last, p21-activated kinases (PAKs) were downregulated in SAP102 KD neurons. These results demonstrate that SAP102 has unique roles in cortical synapse development by mediating EphB and its downstream PAK signaling pathway. Both SAP102 and PAKs are associated with X-linked mental retardation in humans; thus, synapse formation mediated by EphB/SAP102/PAK signaling in the early postnatal brain may be crucial for cognitive development.

On "An obese body mass increases the adverse effects..." Bauer LO, Wu Z, Wolfson LI. Phys Ther. 2011;91:1063-1071.

  • Kietrys D
  • Phys Ther
  • 2011 Sep 2

Literature context:


Abstract:

Funding information:
  • NINDS NIH HHS - NS025044(United States)

Kalirin binds the NR2B subunit of the NMDA receptor, altering its synaptic localization and function.

  • Kiraly DD
  • J. Neurosci.
  • 2011 Aug 31

Literature context:


Abstract:

The ability of dendritic spines to change size and shape rapidly is critical in modulating synaptic strength; these morphological changes are dependent upon rearrangements of the actin cytoskeleton. Kalirin-7 (Kal7), a Rho guanine nucleotide exchange factor localized to the postsynaptic density (PSD), modulates dendritic spine morphology in vitro and in vivo. Kal7 activates Rac and interacts with several PSD proteins, including PSD-95, DISC-1, AF-6, and Arf6. Mice genetically lacking Kal7 (Kal7(KO)) exhibit deficient hippocampal long-term potentiation (LTP) as well as behavioral abnormalities in models of addiction and learning. Purified PSDs from Kal7(KO) mice contain diminished levels of NR2B, an NMDA receptor subunit that plays a critical role in LTP induction. Here we demonstrate that Kal7(KO) animals have decreased levels of NR2B-dependent NMDA receptor currents in cortical pyramidal neurons as well as a specific deficit in cell surface expression of NR2B. Additionally, we demonstrate that the genotypic differences in conditioned place preference and passive avoidance learning seen in Kal7(KO) mice are abrogated when animals are treated with an NR2B-specific antagonist during conditioning. Finally, we identify a stable interaction between the pleckstrin homology domain of Kal7 and the juxtamembrane region of NR2B preceding its cytosolic C-terminal domain. Binding of NR2B to a protein that modulates the actin cytoskeleton is important, as NMDA receptors require actin integrity for synaptic localization and function. These studies demonstrate a novel and functionally important interaction between the NR2B subunit of the NMDA receptor and Kalirin, proteins known to be essential for normal synaptic plasticity.

Funding information:
  • NINDS NIH HHS - R01 NS081674(United States)

PSD-95 is required to sustain the molecular organization of the postsynaptic density.

  • Chen X
  • J. Neurosci.
  • 2011 Apr 27

Literature context:


Abstract:

PSD-95, a membrane-associated guanylate kinase, is the major scaffolding protein in the excitatory postsynaptic density (PSD) and a potent regulator of synaptic strength. Here we show that PSD-95 is in an extended configuration and positioned into regular arrays of vertical filaments that contact both glutamate receptors and orthogonal horizontal elements layered deep inside the PSD in rat hippocampal spine synapses. RNA interference knockdown of PSD-95 leads to loss of entire patches of PSD material, and electron microscopy tomography shows that the patchy loss correlates with loss of PSD-95-containing vertical filaments, horizontal elements associated with the vertical filaments, and putative AMPA receptor-type, but not NMDA receptor-type, structures. These observations show that the orthogonal molecular scaffold constructed from PSD-95-containing vertical filaments and their associated horizontal elements is essential for sustaining the three-dimensional molecular organization of the PSD. Our findings provide a structural basis for understanding the functional role of PSD-95 at the PSD.

Funding information:
  • NIMH NIH HHS - R01 MH083588-10A2(United States)

Ribeye is required for presynaptic Ca(V)1.3a channel localization and afferent innervation of sensory hair cells.

  • Sheets L
  • Development
  • 2011 Apr 9

Literature context:


Abstract:

Ribbon synapses of the ear, eye and pineal gland contain a unique protein component: Ribeye. Ribeye consists of a novel aggregation domain spliced to the transcription factor CtBP2 and is one of the most abundant proteins in synaptic ribbon bodies. Although the importance of Ribeye for the function and physical integrity of ribbon synapses has been shown, a specific role in synaptogenesis has not been described. Here, we have modulated Ribeye expression in zebrafish hair cells and have examined the role of Ribeye in synapse development. Knockdown of ribeye resulted in fewer stimulus-evoked action potentials from afferent neurons and loss of presynaptic Ca(V)1.3a calcium channel clusters in hair cells. Additionally, afferent innervation of hair cells was reduced in ribeye morphants, and the reduction was correlated with depletion of Ribeye punctae. By contrast, transgenic overexpression of Ribeye resulted in Ca(V)1.3a channels colocalized with ectopic aggregates of Ribeye protein. Overexpression of Ribeye, however, was not sufficient to create ectopic synapses. These findings reveal two distinct functions of Ribeye in ribbon synapse formation--clustering Ca(V)1.3a channels at the presynapse and stabilizing contacts with afferent neurons--and suggest that Ribeye plays an organizing role in synaptogenesis.

Funding information:
  • NINDS NIH HHS - R01 NS054814-05(United States)

Long-distance control of synapse assembly by target-derived NGF.

  • Sharma N
  • Neuron
  • 2010 Aug 12

Literature context:


Abstract:

We report a role for long-distance retrograde neurotrophin signaling in the establishment of synapses in the sympathetic nervous system. Target-derived NGF is both necessary and sufficient for formation of postsynaptic specializations on dendrites of sympathetic neurons. This, in turn, is a prerequisite for formation of presynaptic specializations, but not preganglionic axonal ingrowth from the spinal cord into sympathetic ganglia. We also find that NGF-TrkA signaling endosomes travel from distal axons to cell bodies and dendrites where they promote PSD clustering. Furthermore, the p75 neurotrophin receptor restricts PSD formation, suggesting an important role for antagonistic NGF-TrkA and p75 signaling pathways during retrograde control of synapse establishment. Thus, in addition to defining the appropriate number of sympathetic neurons that survive the period of developmental cell death, target-derived NGF also exerts control over the degree of connectivity between the spinal cord and sympathetic ganglia through retrograde control of synapse assembly.

Funding information:
  • NHGRI NIH HHS - U54 HG004555(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.

Wnts acting through canonical and noncanonical signaling pathways exert opposite effects on hippocampal synapse formation.

  • Davis EK
  • Neural Dev
  • 2008 Nov 5

Literature context:


Abstract:

BACKGROUND: Wnt proteins comprise a large class of signaling molecules that regulate a variety of developmental processes, including synapse formation. Previous studies have shown Wnts to be involved in both the induction and prevention of synapses in a number of different organisms. However, it is not clear whether the influence of Wnts on synapses is a result of Wnts' behavior in different organisms or differences in the activity of different Wnt ligands. RESULTS: We used in situ hybridization to show that several Wnt ligands (Wnt3, Wnt5a, Wnt7a, and Wnt7b) and their receptors, Frizzled, are expressed in the developing hippocampus during the period of synapse formation in rodents. We used recombinant Wnt protein or Wnt conditioned media to explore the effects of Wnts on synapses in hippocampal cultures. We found that Wnt7a and Wnt7b activate canonical signaling, whereas Wnt5a activates a noncanonical pathway. The activation of the canonical pathway, either through pathway manipulations or through Wnt stimulation, increases presynaptic inputs. In contrast, exposure to Wnt5a, which activates a noncanonical signaling pathway, decreases the number of presynaptic terminals. CONCLUSION: Our observations suggest that the pro- and antisynaptogenic effects of Wnt proteins are associated with the activation of the canonical and noncanonical Wnt signaling pathways.

Funding information:
  • NIDDK NIH HHS - R01 DK021344(United States)

Capabilities of neurexins in the chick ciliary ganglion.

  • Ross BS
  • Dev Neurobiol
  • 2008 Feb 15

Literature context:


Abstract:

Transcellular interactions between neuroligins (NL) and beta-neurexin have been widely documented to promote maturation and function of both glutamatergic and GABAergic synapses. Recently it has been shown that neuroligin-1 plays a similar role at nicotinic synapses on chick ciliary ganglion neurons in culture, acting from the postsynaptic side to enhance transmitter release from adjacent cholinergic terminals and boost nicotinic input to the cells. We show here that the ciliary ganglion expresses three forms of neuroligin as well as two beta-neurexins and an alpha-neurexin. Overexpression of the beta-neurexins, but not the alpha-neurexin, can induce clustering of endogenous PSD-95 in adjacent neurons, presumably engaging neuroligin in the postsynaptic cell. The trans effects of beta-neurexins are selective; though both alpha3- and alpha7-containing nicotinic receptors are available on opposing cells, beta-neurexins induce coclustering of alpha3- but not alpha7-containing nicotinic receptors. Overexpression of other putative synaptogenic molecules, including SynCAM and L1, are ineffective at trans-clustering of PSD-95 on adjacent neurons. The beta-neurexins also exert a cis effect, coclustering presynaptic markers along with beta-neurexin in neurites juxtaposed to postsynaptic proteins, consistent with organizing presynaptic components as well. Striated muscle, the synaptic target of ciliary neurons in vivo, also expresses neuroligin. The results demonstrate that NL and neurexins are present at multiple sites in nicotinic cholinergic pathways and suggest the possibility of both cis- and trans-interactions to influence nicotinic signaling.

Funding information:
  • NICHD NIH HHS - P01 HD075750(United States)