discs large 1


EVOLUTIONARY HOMOLOGS


Table of contents

Mammalian Discs large homologs and junctional assembly

Phosphatidylinositol 3-kinase (PI3K) is recruited to and activated by E-cadherin engagement. This PI3K activation is essential for adherens junction integrity and intestinal epithelial cell differentiation. Evidence is provided that hDlg, the homolog of disc-large tumor suppressor, is another key regulator of adherens junction integrity and differentiation in mammalian epithelial cells. This study reports the following: (1) hDlg co-localizes with E-cadherin, but not with ZO-1, at the sites of cell-cell contact in intestinal epithelial cells; (2) reduction of hDlg expression levels by RNAi in intestinal cells not only severely alters adherens junction integrity but also prevents the recruitment of p85/PI3K to E-cadherin-mediated cell-cell contact and inhibits sucrase-isomaltase gene expression; (3) PI3K and hDlg are associated with E-cadherin in a common macromolecular complex in living differentiating intestinal cells; (4) this interaction requires the association of hDlg with E-cadherin and with Src homology domain 2 domains of the p85/PI3K subunit; (5) phosphorylation of hDlg on serine and threonine residues prevents its interaction with the p85 Src homology domain 2 in subconfluent cells, whereas phosphorylation of hDlg on tyrosine residues is essential. It is concluded that hDlg may be a determinant in E-cadherin-mediated adhesion and signaling in mammalian epithelial cells (Laprise, 2004).

Discs large homologs and synaptic development

PSD-95 is a neuronal PDZ protein that associates with receptors and cytoskeletal elements at synapses, but whose function is uncertain. Overexpression of PSD-95 in hippocampal neurons can drive maturation of glutamatergic synapses. PSD-95 expression enhances postsynaptic clustering and activity of glutamate receptors. Postsynaptic expression of PSD-95 also enhances maturation of the presynaptic terminal. These effects require synaptic clustering of PSD-95 but do not rely on the PSD-95 guanylate kinase domain. PSD-95 expression also increases the number and size of dendritic spines. These results demonstrate that PSD-95 can orchestrate synaptic development and are suggestive of roles for PSD-95 in synapse stabilization and plasticity (El-Husseini, 2000).

PSD-95 can drive maturation of synapses, not only of postsynaptic components but also of presynaptic terminals. The selective enhancement of GluR1 versus NMDA receptor clustering correlates with anatomical studies showing that the number of NMDARs remains relatively constant, whereas the number of synaptic GluRs increases during development and the magnitude of GluR quantal response increases as synapses mature. It is not clear whether the increase in size of synaptic spines and the increase in GluR1 clustering induced by PSD-95 are parallel processes or if one triggers the other. Also unclear is the mechanism underlying the enhanced GluR1 clustering, which presumably involves an intermediary protein(s), because PSD-95 does not bind GluR1. The enhanced size of axon terminals contacting neurons transfected with PSD-95 and the increased frequency of mEPSCs can be explained by the hypothesis that PSD-95 conveys a retrograde signal for presynaptic development. The increased frequency of mEPSCs presumably reflects the increased probability of release associated with an increased vesicle pool size. This result may explain why the PSD-95 knockout mouse has augmented paired pulse facilitation, which would be consistent with a decreased probability of release in the mutant. The transsynaptic influence of PSD-95 is reminiscent of rapsyn, a nicotinic acetylcholine receptor clustering protein essential for differentiation of motor neuron terminals. PSD-95 may communicate with the axon through neuroligin, a PSD-95-associated cell adhesion molecule that links to the nerve terminal by means of neurexins. Very recent studies show that neuroligin expression in heterologous cells can trigger presynaptic development (El-Husseini, 2000).

Excitatory synapses in the brain exhibit a remarkable degree of functional plasticity, which largely reflects changes in the number of synaptic AMPA receptors. However, mechanisms involved in recruiting AMPA receptors to synapses are unknown. Hippocampal slice cultures and biolistic gene transfections have been used to study the targeting of AMPA receptors to synapses. AMPA receptors are localized to synapses through direct binding of the first two PDZ domains of synaptic PSD-95 to the AMPAR-associated protein, stargazin. Increasing the level of synaptic PSD-95 recruits new AMPA receptors to synapses without changing the number of surface AMPARs. At the same time, stargazin overexpression drastically increases the number of extra-synaptic AMPA receptors, but fails to alter synaptic currents if synaptic PSD-95 levels are kept constant. Finally, compensatory mutations were made to both PSD-95 and stargazin to demonstrate the central role of direct interactions between them in determining the number of synaptic AMPARs (Schnell, 2002).

Three groups of evolutionarily conserved proteins have been implicated in the establishment of epithelial cell polarity: the apically-localized proteins of the Par (Par3-Par6-aPKC-Cdc42) and Crumbs groups (Crb3-PALS1-PATJ) and the basolaterally-localized proteins of the Dlg group (Dlg1-Scribble-Lgl). During epithelial morphogenesis, these proteins participate in a complex network of interdependent interactions that define the position and functional organization of adherens junctions and tight junctions. However, the biochemical pathways through which they control polarity are poorly understood. In this study, an interaction was identified between endogenous hDlg1 and MPP7, a previously uncharacterized MAGUK-p55 subfamily member. MPP7 targets to the lateral surface of epithelial cells via its L27N domain, through an interaction with hDlg1. Loss of either hDlg1 or MPP7 from epithelial Caco-2 cells results in a significant defect in the assembly and maintenance of functional tight junctions. It is concluded that the formation of a complex between hDlg1 and MPP7 promotes epithelial cell polarity and tight junction formation (Stucke, 2007).

The origins and evolution of higher cognitive functions, including complex forms of learning, attention and executive functions, are unknown. A potential mechanism driving the evolution of vertebrate cognition early in the vertebrate lineage (550 million years ago) was genome duplication and subsequent diversification of postsynaptic genes. This study reports the first genetic analysis of a vertebrate gene family in cognitive functions measured using computerized touchscreens. Comparison of mice carrying mutations in each of the four Dlg paralogs showed that simple associative learning required Dlg4, whereas Dlg2 and Dlg3 diversified to have opposing functions in complex cognitive processes. Exploiting the translational utility of touchscreens in humans and mice, testing Dlg2 mutations in both species showed that Dlg2's role in complex learning, cognitive flexibility and attention has been highly conserved over 100 million years. Dlg-family mutations underlie psychiatric disorders, suggesting that genome evolution expanded the complexity of vertebrate cognition at the cost of susceptibility to mental illness (Nithianantharajah, 2013). SynGAP is a Ras/Rap GTPase-activating protein (GAP) that is a major constituent of postsynaptic densities (PSDs) from mammalian forebrain. Its α1 isoform binds to all three PDZ (PSD-95, Discs-large, ZO-1) domains of PSD-95 (see Drosophila Discs large), the principal PSD scaffold, and can occupy as many as 15% of these PDZ domains. Evidence is presented that synGAP-α1 regulates the composition of the PSD by restricting binding to the PDZ domains of PSD-95. Phosphorylation by Ca2+/calmodulin-dependent protein kinase II (CaMKII; see Drosophila CaMKII) and Polo-like kinase-2 (PLK2; see Drosophila Polo) decreases its affinity for the PDZ domains by several fold, which would free PDZ domains for occupancy by other proteins. Finally, three critical postsynaptic signaling proteins that bind to the PDZ domains of PSD-95 are shown to be present in higher concentration in PSDs isolated from mice with a heterozygous deletion of synGAP (Walkup , 2016).

Discs large homologs and receptor/channel signaling complexes

The postsynaptic density (PSD) can be visualized as an ultrastructural thickening of the postsynaptic membrane that is characteristic of excitatory synapses. Among the glutamate receptor complexes, the NMDA receptor/PSD-95 complex is the one most tightly associated with the PSD. In biochemical preparations of the PSD, NMDA receptors and PSD-95 are highly enriched and resistant to extraction by Triton X-100 and sarkosyl detergents, while AMPA receptors/GRIP and mGluRs/Homer (see Drosophila Homer) are relatively soluble. It is possible that the components of the NMDA receptor/PSD-95 complex comprise the major constituents of the core PSD, which remains after extraction with strong detergents. Because they are likely to play critical roles in the structural organization of the synapse and in the transduction of NMDA receptor signals, these core PSD proteins are important to define and study. A family of proteins (termed GKAP, SAPAP, or DAP) has been characterized that is highly concentrated in the PSD and that binds to the guanylate kinase (GK) domain of PSD-95. GKAP appears to be tightly associated with PSD-95; it can be immunoprecipitated from the brain in a complex with PSD-95 family proteins, and it is consistently colocalized with PSD-95 in neurons, even in the absence of associated NMDA receptors. The GKAP family of proteins contains at least four members and undergoes complex alternative splicing, but the physiological roles of these variants are unknown. To gain insight into GKAP function, a screen was carried out for binding partners of GKAP, hoping to extend the network of protein interactions emanating from NMDA receptors into the PSD (Naisbitt, 1999a and references).

A novel family of postsynaptic density (PSD) proteins, termed Shank, is described that binds via its PDZ domain to the C terminus of PSD-95-associated protein GKAP. A ternary complex of Shank/GKAP/PSD-95 assembles in heterologous cells and can be coimmunoprecipitated from rat brain. Synaptic localization of Shank in neurons is inhibited by a GKAP splice variant that lacks the Shank-binding C terminus. In addition to its PDZ domain, Shank contains a proline-rich region that binds to cortactin and a SAM domain that mediates multimerization. Shank may function as a scaffold protein in the PSD, potentially cross-linking NMDA receptor/PSD-95 complexes and coupling them to regulators of the actin cytoskeleton (Naisbitt, 1999a).

Originally identified as a substrate of Src tyrosine kinase, cortactin is an F-actin-binding protein enriched in cell-matrix contact sites, membrane ruffles and lammelipodia of cultured cells, and in growth cones of neurons. The translocation of cortactin to the cell periphery is stimulated by the small GTPase Rac1, and its F-actin cross-linking activity is inhibited by Src tyrosine phosphorylation. Thus, a large body of evidence implicates cortactin in regulation of the actin cytoskeleton in dynamic regions of the cell periphery. This study suggests that cortactin may also play a role in neuronal synapses, based on the following findings: biochemically, cortactin is loosely associated with the PSD, and immunocytochemically, it colocalizes with Shank in a subset of synapses. Most interestingly, a significant redistribution of cortactin to synaptic sites in response to glutamate stimulation has been demonstrated. The glutamate-induced synaptic localization of cortactin is reminiscent of cortactin recruitment to the cortical cytoskeleton by growth factor stimulation of nonneural cells. Their coexistence in growth cones supports the suggestion that Shank and cortactin may function at sites of active cytoskeletal remodeling in neurons. In mature synapses, it is speculated that a regulated Shank-cortactin interaction may be a mechanism for linking NMDA receptor activation to the control of the postsynaptic actin cytoskeleton. Shank is highly related to CortBP1, a protein isolated by yeast two-hybrid screening with the SH3 domain of cortactin. CortBP1 has been shown to colocalize with cortactin in membrane ruffles of cultured cells and in growth cones of cultured neurons, analogous to the colocalization of Shank and cortactin in growth cones and synapses. Based on their similarity in primary structure and cell biological properties, it seems reasonable to consider CortBP1 and Shank as members of the same family of proteins (Naisbitt, 1999a and references).

The structure of central synapses at the molecular level is poorly understood. A recent advance came with the identification of the postsynaptic density-95 (PSD-95)/synapse-associated protein 90 family of proteins. Members of this family are important mediators of the synaptic clustering of certain classes of ion channels. By yeast two-hybrid screening, a novel protein termed guanylate kinase-associated protein (GKAP) has been isolated that binds to the GK-like domain of PSD-95. Reported here is a detailed characterization of GKAP expression in the rat brain as well as the cloning of a novel GKAP splice variant. By Northern blot, GKAP mRNAs (4, 6.5, and 8 kB) are expressed predominantly in the rat brain. By in situ hybridization, GKAP is expressed widely in neurons of cortex and hippocampus and in the Purkinje and granule cells of the cerebellum. On brain immunoblots, two prominent bands of 95 and 130 kDa are detected that correspond to products of short and long N-terminal splice variants of GKAP. Two independent GKAP antibodies label somatodendritic puncta in neocortical and hippocampal neurons, in a pattern consistent with synaptic elements. Immunogold electron microscopy reveals GKAP to be predominantly postsynaptic and present at asymmetric synapses and in dendritic spines. The distribution of GKAP immunogold particles is uniform in the lateral plane of the PSD but peaks in the perpendicular axis approximately 20 nm from the postsynaptic membrane. In cultured hippocampal neurons GKAP immunoreactive puncta colocalize with the AMPA receptor subunit Glu receptor 1 but not with the GABAA receptor subunits beta2 and beta3. Thus GKAP is a widely expressed neuronal protein localized specifically in the PSD of glutamatergic synapses, consistent with its direct interaction with PSD-95 family proteins (Naisbitt, 1999b).

The human homolog of the Drosophila discs large tumor suppressor protein (hDLG) and closely related proteins such as postsynaptic density protein 95 kDa (PSD-95) are associated with N-methyl-D-aspartate receptors (NMDA-R) and Shaker-type K+ channels, and are thought to be involved in the clustering of these types of channel proteins. A protein named DAP-1 has been identified that binds to the guanylate kinase-like domains of hDLG and PSD-95. DAP-1 associates with hDLG, PSD-95, NMDA-R and adenomatous polyposis coli protein (APC). Furthermore, DAP-1 is specifically expressed in the brain and colocalizes with PSD-95 and APC in mouse cerebellum. DAP-1 is colocalized with PSD-95 and NMDA-R at the synapses in cultured rat hippocampal neurons. These findings suggest that DAP-1 may play several roles in the molecular organization of synapses and neuronal cell signaling by interacting with hDLG and PSD-95, which in turn are associated with receptors, ion channels and APC (Satoh, 1999).

Shank cross-links Homer and PSD-95 complex in the postsynaptic density (PSD) and plays a role in the signaling mechanisms of both metabotropic receptors and NMDA receptors. Homer is a neuronal immediate early gene (IEG) that selectively binds the C terminus of group 1 metabotropic receptors (mGluR1a and mGluR5) and is enriched at excitatory synapses. In the adult brain, Homer is rapidly and transiently induced by physiological synaptic stimuli that evoke long-term potentiation in the hippocampus and is also induced in the striatum by dopamimetic drugs of addiction. The Homer IEG, now termed Homer1a, is a member of a family of closely related Homer proteins which are constitutively expressed in brain and enriched at excitatory synapses. Currently, there are three mammalian Homer genes and at least six distinct transcripts expressed in brain. Like Homer1a, all new family members contain an N-terminal: an approximately 110-amino acid domain that binds mGluR1a/5. The region of Homer that interacts with mGluR1a/5 is termed an EVH1 domain based on homology to domains in a family of proteins that include Drosophilia Enabled, mammalian VASP, and the Wiscott-Aldridge syndrome protein (WASP). The EVH1 domain of Homer is conserved at a level of 80% identity between Drosophilia, rodent, and human. The three-dimensional structures of the mammalian Enabled (Mena) EVH1 domain and the Homer1 EVH1 domain have recently been shown to be similar to the pleckstrin homology (PH) and phosphotyrosine-binding protein (PTB) domains. Other than the IEG Homer1a, all forms of Homer encode an additional C-terminal domain with predicted coiled-coil (CC) structure. The CC domain mediates homo- and heteromultimerization between Homer proteins, and such multimers are naturally present in brain. Another distinction from Homer1a is that all CC-containing Homer family members (CC-Homers) are constitutively expressed in brain. CC-Homers are enriched in postsynaptic density fractions, are ultrastructurally localized at the PSD, and coimmunoprecipitate with group 1 mGluRs from brain. These observations suggest that CC-Homer proteins function as multivalent adapter complexes that bind mGluRs at postsynaptic sites (Tu, 1999 and references).

Analysis of Homer EVH1-binding specificity led to the discovery of other physiological binding partners of Homer. The Homer EVH1 domain binds an internal, proline-rich sequence that is approximately 50 residues from the C terminus of both mGluR1a and mGluR5. A similar peptide sequence was identified in the cytosolically disposed N terminus of inositol trisphosphate receptors (IP3R). In brain, IP3R are enriched in a specialization of the endoplasmic reticulum that extends into dendritic spines, termed the spine apparatus. The spine apparatus appears throughout the spine and conspiciously approaches the lateral margin of the PSD, precisely where group 1 mGluRs are concentrated. This ultrastructure is consistent with the hypothesis that CC-Homers couple mGluR in the postsynaptic membrane to IP3R in the spine apparatus. Synaptic mGluR-mediated release of Ca2+ in dendrites of Purkinje neurons is highly localized, consistent with spatially localized coupling of mGluR and IP3R. The CC-Homer-linked signaling complex is additionally notable in that it can be dynamically regulated by Homer1a. Homer1a can bind either mGluR1a/5 or IP3R, but does not encode a CC domain, and cannot cross-link the receptors. Accordingly, Homer1a functions as a natural dominant negative. Consistent with this model, expression of the IEG form of Homer in cerebellar Purkinje cells reduces and delays mGluR-mediated release of intracellular Ca2+. These observations suggest that upregulation of Homer1a, which occurs in response to synaptic activity, dampens glutamate-induced release of Ca2+ from intracellular IP3R-gated stores (Tu, 1999).

Shank is a recently described family of postsynaptic proteins that function as part of the NMDA receptor-associated PSD-95 complex. Shank proteins also bind to Homer. Homer proteins form multivalent complexes that bind proline-rich motifs in group 1 metabotropic glutamate receptors and inositol trisphosphate receptors, thereby coupling these receptors in a signaling complex. A single Homer-binding site is identified in Shank, and Shank and Homer coimmunoprecipitate from brain and colocalize at postsynaptic densities. Moreover, Shank clusters mGluR5 in heterologous cells in the presence of Homer and mediates the coclustering of Homer with PSD-95/GKAP. Thus, Shank may cross-link Homer and PSD-95 complexes in the PSD and play a role in the signaling mechanisms of both mGluRs and NMDA receptors (Tu, 1999).

PSD-95/SAP90 is a member of membrane-associated guanylate kinases localized at the postsynaptic density (PSD) in neuronal cells. Membrane-associated guanylate kinases are a family of signaling molecules expressed at various submembrane domains which have the PDZ (DHR) domains, the SH3 domain, and the guanylate kinase domain. PSD-95/SAP90 interacts with N-methyl-D-aspartate receptors 2A/B, Shaker-type potassium channels, and brain nitric oxide synthase through the PDZ (DHR) domains and clusters these molecules at synaptic junctions. However, neither the function of the SH3 domain or the guanylate kinase domain of PSD-95/SAP90, nor the protein interacting with these domains has been identified. A novel protein family has been isolated consisting of at least four members that specifically interact with PSD-95/SAP90 and its related proteins through the guanylate kinase domain. These proteins have been named SAPAPs (SAP90/PSD-95-associated proteins). SAPAPs are specifically expressed in neuronal cells and enriched in the PSD fraction. SAPAPs induce the enrichment of PSD-95/SAP90 to the plasma membrane in transfected cells. Thus, SAPAPs may have a potential activity to maintain the structure of PSD by concentrating its components to the membrane area (Takeuchi, 1997).

PSD-95/SAP90 is a synaptic membrane-associated guanylate kinase with three PDZ, one SH3, and one guanylate kinase (GK) domain. PSD-95/SAP90 binds various proteins through the PDZ domains and organizes synaptic junctions. PSD-95/SAP90 also interacts with the postsynaptic density (PSD) fraction-enriched protein, named SAPAP (also called GKAP and DAP), through the GK domain. SAPAP is Triton X-100-insoluble and recruits PSD-95/SAP90 into the Triton X-100-insoluble fraction in the transfected cells, suggesting that SAPAP may fix PSD-95/SAP90 to the PSD. BEGAIN, a novel protein interacting with the GK domain of PSD-95/SAP90, is described. BEGAIN is specifically expressed in brain and enriched in the PSD fraction. BEGAIN is Triton X-100-soluble in the transfected cells but is recruited to the Triton X-100-insoluble fraction by SAPAP when coexpressed with PSD-95/SAP90. BEGAIN may be a novel PSD component associated with the core complex of PSD-95/SAP90 and SAPAP (Deguchi, 1998).

Guanylate kinase-associated protein (GKAP)/SAP90/PSD-95-associated protein (SAPAP)/DLG-associated protein (DAP) is a protein of the postsynaptic density (PSD), and binds to the guanylate kinase domain of PSD-95/synapse-associated protein (SAP) 90 and synaptic scaffolding molecule. GKAP/SAPAP/DAP recruits PSD-95/SAP90 and its interacting protein, brain-enriched guanylate kinase-interacting protein, into the Triton X-100-insoluble fraction in transfected cells, suggesting that GKAP/SAPAP/DAP may link several PSD components to the Triton X-100-insoluble structures in the PSD. A novel neuronal GKAP/SAPAP/DAP-binding protein has been identified and named synamon. Synamon has seven ankyrin repeats at the NH(2) terminus followed by one src homology 3 domain and one PSD-95/Dlg-A/ZO-1 domain, and several proline-rich regions at the carboxyl terminus. Synamon interacts with the COOH-terminal region of GKAP/SAPAP/DAP via the middle region containing a PSD-95/Dlg-A/ZO-1 domain. Synamon was coimmunoprecipitated with SAPAP from rat crude synaptosomes and colocalized with SAPAP in primary cultured rat hippocampal neurons. Because synamon is composed of various protein-interacting modules, it may also interact with proteins other than GKAP/SAPAP/DAP to organize the architecture of the PSD (Yao, 1999).

Neuregulins (NRGs) and their receptors, the ErbB protein tyrosine kinases, are essential for neuronal development, but their functions in the adult CNS are unknown. The neuregulin receptor ErbB4 is enriched in the postsynaptic density (PSD) and associates with PSD-95. Heterologous expression of PSD-95 enhances NRG activation of ErbB4 and MAP kinase. Conversely, inhibiting expression of PSD-95 in neurons attenuates NRG-mediated activation of MAP kinase. PSD-95 forms a ternary complex with two molecules of ErbB4, suggesting that PSD-95 facilitates ErbB4 dimerization. NRG suppresses induction of long-term potentiation in the hippocampal CA1 region without affecting basal synaptic transmission. Thus, NRG signaling may be synaptic and regulated by PSD-95. A role of NRG signaling in the adult CNS may be modulation of synaptic plasticity (Huang, 2000).

In the CA1 region of the hippocampus, a brief train of high-frequency stimulation produces the lasting enhancement of synaptic transmission known as LTP. Stimulation of the NMDA class of glutamate receptors is a key step in the mechanism of LTP. The findings that NRG suppresses induction of LTP in CA1 but has no effect on paired-pulse facilitation together with the localization of ErbB4 in the PSDs suggest that NRG acts via a postsynaptic mechanism. The activity of NMDARs is subject to modulation by tyrosine phosphorylation, but the suppression of LTP could not be attributed to an effect on the amplitude of NMDA currents because NRG does not alter NMDA receptor–mediated synaptic responses. Thus, NRG/ErbB4 signaling may interrupt LTP induction at a step beyond NMDAR stimulation. ErbB4 has an extended C-terminal region that contains numerous tyrosine residues. Upon phosphorylation, these tyrosine residues bind to adapter proteins and activate a diversity of signaling pathways including ERK, JNK, and PI3 kinase. One of these kinase signaling cascades may have a role in the suppression of synaptic plasticity by NRG (Huang, 2000).

NRG-2 and NRG-3 are expressed in adult hippocampus. At the cellular level, nrg mRNAs are detected in granule cells of the hippocampus and the dentate gyrus. There are at least 14 different splice variants of the nrg-1 mRNAs that encode proteins containing an alpha- or beta-type epidermal growth factor (EGF) domain and either an immunoglobulin- (Ig-)like domain or a cysteine-rich domain (CRD) at the N terminus. The Ig-containing, but not CRD-containing, NRG-1 is expressed in adult hippocampus. All NRG functions known to date can be duplicated by the EGF domain. The NRG used in this study, rHRGbeta177–244, is a recombinant polypeptide containing the entire EGF domain (amino acids 177–244) of the beta-type NRG-1, a potent isoform. rHRGbeta177–244 binds to ErbB3 and ErbB4 and thus induces tyrosine phosphorylation of ErbB2, ErbB3, and ErbB4, but not of the EGF receptor. Thus, the effect of rHRGbeta177–244 on LTP is believed to be mediated via activation of the NRG signaling pathway. Exactly which NRGs or NRG isoforms are involved in this event requires further studies (Huang, 2000).

Chronic pain due to nerve injury is resistant to current analgesics. Animal models of neuropathic pain show neuronal plasticity and behavioral reflex sensitization in the spinal cord that depends on the NMDA receptor. Complexes of NMDA receptors with the multivalent adaptor protein PSD-95 are found in the dorsal horn of spinal cord; PSD-95 plays a key role in neuropathic reflex sensitization. Mutant mice expressing a truncated form of the PSD-95 molecule fail to develop the NMDA receptor-dependent hyperalgesia and allodynia seen in the CCI model of neuropathic pain, but develop normal inflammatory nociceptive behavior following the injection of formalin. In wild-type mice following CCI, CaM kinase II inhibitors attenuate sensitization of behavioral reflexes; elevated constitutive (autophosphorylated) activity of CaM kinase II is detected in spinal cord, and increased amounts of phospho-Thr286 CaM kinase II coimmunoprecipitate with NMDA receptor NR2A/B subunits. Each of these changes is prevented in PSD-95 mutant mice although CaM kinase II is present and can be activated. Disruption of CaM kinase II docking to the NMDA receptor and activation may be responsible for the lack of neuropathic behavioral reflex sensitization in PSD-95 mutant mice (Garry, 2003).

PSD-95 is a major scaffolding protein of the postsynaptic density, tethering NMDA- and AMPA-type glutamate receptors to signaling proteins and the neuronal cytoskeleton. PSD-95 is regulated by the ubiquitin-proteasome pathway. PSD-95 interacts with and is ubiquitinated by the E3 ligase Mdm2. In response to NMDA receptor activation, PSD-95 is ubiquitinated and rapidly removed from synaptic sites by proteasome-dependent degradation. Mutations that block PSD-95 ubiquitination prevent NMDA-induced AMPA receptor endocytosis. Likewise, proteasome inhibitors prevent NMDA-induced AMPA receptor internalization and synaptically induced long-term depression. This is consistent with the notion that PSD-95 levels are an important determinant of AMPA receptor number at the synapse. These data suggest that ubiquitination of PSD-95 through an Mdm2-mediated pathway is critical in regulating AMPA receptor surface expression during synaptic plasticity (Colledge, 2003).

A novel rat gene, tanc (GenBank Accession No. AB098072), has been cloned that encodes a protein containing three tetratricopeptide repeats (TPRs), ten ankyrin repeats and a coiled-coil region, and is possibly a rat homolog of Drosophila rolling pebbles. The tanc gene is expressed widely in the adult rat brain. Subcellular distribution, immunohistochemical study of the brain and immunocytochemical studies of cultured neuronal cells indicate the postsynaptic localization of TANC protein of 200 kDa. Pull-down experiments have shown that TANC protein binds PSD-95, SAP97, and Homer via its C-terminal PDZ-binding motif, -ESNV, and fodrin via both its ankyrin repeats and the TPRs together with the coiled-coil domain. TANC also binds the alpha subunit of Ca2+/calmodulin-dependent protein kinase II. An immunoprecipitation study shows TANC association with various postsynaptic proteins, including guanylate kinase-associated protein (GKAP), alpha-internexin, and N-methyl-D-aspartate (NMDA)-type glutamate receptor 2B and AMPA-type glutamate receptor (GluR1) subunits. These results suggest that TANC protein may work as a postsynaptic scaffold component by forming a multiprotein complex with various postsynaptic density proteins (Suzuki, 2004).

Oncogenic functions for Disc large homologs

Several different human virus oncoproteins, including adenovirus type 9 E4-ORF1, evolved to target the Dlg1 mammalian homolog of the membrane-associated Drosophila discs-large tumor suppressor. This fact has implicated this cellular factor in human cancer. Despite a general belief that such interactions function solely to inactivate this suspected human tumor suppressor protein, this study demonstrates that E4-ORF1 specifically requires endogenous Dlg1 to provoke oncogenic activation of phosphatidylinositol 3-kinase (PI3K) in cells. Based on these results, a model is proposed wherein E4-ORF1 binding to Dlg1 triggers the resulting complex to translocate to the plasma membrane and, at this site, to promote Ras-mediated PI3K activation. These findings establish the first known function for Dlg1 in virus-mediated cellular transformation and also surprisingly expose a previously unrecognized oncogenic activity encoded by this suspected cellular tumor suppressor gene (Frese, 2006).

It is important to point out that, though unexpected, the finding that Dlg1 possesses an oncogenic activity does not refute a substantial body of evidence supporting its designation as a candidate tumor suppressor gene. Viral oncoproteins typically disregulate normal functions of their cellular targets, so it may be pertinent that Dlg1 was recently reported to recruit PI3K into a membrane-associated complex with E-cadherin to promote both adherens junction formation and terminal differentiation in enterocytes. Since this activity is predicted to induce an antiproliferative state in cells, it may represent a tumor suppressor function for Dlg1. Therefore, an intriguing possibility presents itself: whereas Dlg1 normally functions to promote temporally and spatially restricted E-cadherin-dependent PI3K activation during adherens junction formation in cells, Dlg1 modified by E4-ORF1 instead supports constitutive and spatially unrestricted, Ras-dependent PI3K activation at the plasma membrane. This interesting model hints that E4-ORF1 may simultaneously inactivate a tumor suppressor function and stimulate an oncogenic function of Dlg1 (Frese, 2006).


Table of contents


discs large 1: Biological Overview | Regulation | Developmental Biology | Effects of Mutation | References

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