A rare autism-associated MINT2/APBA2 mutation disrupts neurexin trafficking and synaptic function.
ABSTRACT: MINT2/APBA2 is a synaptic adaptor protein involved in excitatory synaptic transmission. Several nonsynonymous coding variants in MINT2 have been identified in autism spectrum disorders (ASDs); however, these rare variants have not been examined functionally and the pathogenic mechanisms are unknown. Here, we examined the synaptic effects of rat Mint2 N723S mutation (equivalent to autism-linked human MINT2 N722S mutation) which targets a conserved asparagine residue in the second PDZ domain of Mint2 that binds to neurexin-1? (Nrxn1?), a presynaptic cell-adhesion protein implicated in ASDs. We show the N723S mutation impairs Nrxn1? stabilization and trafficking to the membrane while binding to Nrxn1? remains unaffected. Using time-lapse imaging in primary mouse neurons, we found that the N723S mutant had more immobile puncta at neuronal processes compared to Mint2 wild type. We therefore, reasoned that the N723S mutant may alter the co-transport of Nrxn1? at axonal processes to presynaptic terminals. Indeed, we found the N723S mutation affected Nrxn1? localization at presynaptic terminals which correlated with a decrease in Nrxn-mediated synaptogenesis and miniature event frequency in excitatory synapses. Together, our data reveal Mint2 N723S leads to neuronal dysfunction, in part due to alterations in Nrxn1? surface trafficking and synaptic function of Mint2.
Project description:The differential formation of excitatory (glutamate-mediated) and inhibitory (GABA-mediated) synapses is a critical step for the proper functioning of the brain. An imbalance in these synapses may lead to various neurological disorders such as autism, schizophrenia, Tourette's syndrome and epilepsy. Synapses are formed through communication between the appropriate synaptic partners. However, the molecular mechanisms that mediate the formation of specific synaptic types are not known. Here we show that two members of the fibroblast growth factor (FGF) family, FGF22 and FGF7, promote the organization of excitatory and inhibitory presynaptic terminals, respectively, as target-derived presynaptic organizers. FGF22 and FGF7 are expressed by CA3 pyramidal neurons in the hippocampus. The differentiation of excitatory or inhibitory nerve terminals on dendrites of CA3 pyramidal neurons is specifically impaired in mutants lacking FGF22 or FGF7. These presynaptic defects are rescued by postsynaptic expression of the appropriate FGF. FGF22-deficient mice are resistant to epileptic seizures, and FGF7-deficient mice are prone to them, as expected from the alterations in excitatory/inhibitory balance. Differential effects of FGF22 and FGF7 involve both their distinct synaptic localizations and their use of different signalling pathways. These results demonstrate that specific FGFs act as target-derived presynaptic organizers and help to organize specific presynaptic terminals in the mammalian brain.
Project description:Synaptogenic adhesion molecules play critical roles in synapse formation. SALM5/Lrfn5, a SALM/Lrfn family adhesion molecule implicated in autism spectrum disorders (ASDs) and schizophrenia, induces presynaptic differentiation in contacting axons, but its presynaptic ligand remains unknown. We found that SALM5 interacts with the Ig domains of LAR family receptor protein tyrosine phosphatases (LAR-RPTPs; LAR, PTP?, and PTP?). These interactions are strongly inhibited by the splice insert B in the Ig domain region of LAR-RPTPs, and mediate SALM5-dependent presynaptic differentiation in contacting axons. In addition, SALM5 regulates AMPA receptor-mediated synaptic transmission through mechanisms involving the interaction of postsynaptic SALM5 with presynaptic LAR-RPTPs. These results suggest that postsynaptic SALM5 promotes synapse development by trans-synaptically interacting with presynaptic LAR-RPTPs and is important for the regulation of excitatory synaptic strength.
Project description:Genetic aspects of autism spectrum disorders (ASDs) have recently been extensively explored, but environmental influences that affect ASDs have received considerably less attention. Zinc (Zn) is a nutritional factor implicated in ASDs, but evidence for a strong association and linking mechanism is largely lacking. Here we report that trans-synaptic Zn mobilization rapidly rescues social interaction in two independent mouse models of ASD. In mice lacking Shank2, an excitatory postsynaptic scaffolding protein, postsynaptic Zn elevation induced by clioquinol (a Zn chelator and ionophore) improves social interaction. Postsynaptic Zn is mainly derived from presynaptic pools and activates NMDA receptors (NMDARs) through postsynaptic activation of the tyrosine kinase Src. Clioquinol also improves social interaction in mice haploinsufficient for the transcription factor Tbr1, which accompanies NMDAR activation in the amygdala. These results suggest that trans-synaptic Zn mobilization induced by clioquinol rescues social deficits in mouse models of ASD through postsynaptic Src and NMDAR activation.
Project description:Both genetic and environmental factors are thought to contribute to neurodevelopmental and neuropsychiatric disorders with maternal immune activation (MIA) being a risk factor for both autism spectrum disorders and schizophrenia. Although MIA mouse offspring exhibit behavioral impairments, the synaptic alterations in vivo that mediate these behaviors are not known. Here we employed in vivo multiphoton imaging to determine that in the cortex of young MIA offspring there is a reduction in number and turnover rates of dendritic spines, sites of majority of excitatory synaptic inputs. Significantly, spine impairments persisted into adulthood and correlated with increased repetitive behavior, an ASD relevant behavioral phenotype. Structural analysis of synaptic inputs revealed a reorganization of presynaptic inputs with a larger proportion of spines being contacted by both excitatory and inhibitory presynaptic terminals. These structural impairments were accompanied by altered excitatory and inhibitory synaptic transmission. Finally, we report that a postnatal treatment of MIA offspring with the anti-inflammatory drug ibudilast, prevented both synaptic and behavioral impairments. Our results suggest that a possible altered inflammatory state associated with maternal immune activation results in impaired synaptic development that persists into adulthood but which can be prevented with early anti-inflammatory treatment.
Project description:Heterozygous mutations of the NRXN1 gene, which encodes the presynaptic cell-adhesion molecule neurexin-1, were repeatedly associated with autism and schizophrenia. However, diverse clinical presentations of NRXN1 mutations in patients raise the question of whether heterozygous NRXN1 mutations alone directly impair synaptic function. To address this question under conditions that precisely control for genetic background, we generated human ESCs with different heterozygous conditional NRXN1 mutations and analyzed two different types of isogenic control and NRXN1 mutant neurons derived from these ESCs. Both heterozygous NRXN1 mutations selectively impaired neurotransmitter release in human neurons without changing neuronal differentiation or synapse formation. Moreover, both NRXN1 mutations increased the levels of CASK, a critical synaptic scaffolding protein that binds to neurexin-1. Our results show that, unexpectedly, heterozygous inactivation of NRXN1 directly impairs synaptic function in human neurons, and they illustrate the value of this conditional deletion approach for studying the functional effects of disease-associated mutations.
Project description:Angelman Syndrome is a debilitating neurological disorder caused by mutation of the E3 ubiquitin ligase Ube3A, a gene whose mutation has also recently been associated with autism spectrum disorders (ASDs). The function of Ube3A during nervous system development and how Ube3A mutations give rise to cognitive impairment in individuals with Angleman Syndrome and ASDs are not clear. We report here that experience-driven neuronal activity induces Ube3A transcription and that Ube3A then regulates excitatory synapse development by controlling the degradation of Arc, a synaptic protein that promotes the internalization of the AMPA subtype of glutamate receptors. We find that disruption of Ube3A function in neurons leads to an increase in Arc expression and a concomitant decrease in the number of AMPA receptors at excitatory synapses. We propose that this deregulation of AMPA receptor expression at synapses may contribute to the cognitive dysfunction that occurs in Angelman Syndrome and possibly other ASDs.
Project description:Autism spectrum disorders (ASDs) comprise a highly heritable, multifarious group of neurodevelopmental disorders, which are characterized by repetitive behaviors and impairments in social interactions. Point mutations have been identified in X-linked Neuroligin (NLGN) 3 and 4X genes in patients with ASDs and all of these reside in their extracellular domains except for a single point mutation in the cytoplasmic domain of NLGN4X in which an arginine is mutated to a cysteine (R704C). Here we show that endogenous NLGN4X is robustly phosphorylated by protein kinase C (PKC) at T707, and R704C completely eliminates T707 phosphorylation. Endogenous NLGN4X is intensely phosphorylated on T707 upon PKC stimulation in human neurons. Furthermore, a phospho-mimetic mutation at T707 has a profound effect on NLGN4X-mediated excitatory potentiation. Our results now establish an important interplay between a genetic mutation, a key posttranslational modification, and robust synaptic changes, which can provide insights into the synaptic dysfunction of ASDs.
Project description:The generation of precise synaptic connections between developing neurons is critical to the formation of functional neural circuits. Astrocyte-secreted glypican 4 induces formation of active excitatory synapses by recruiting AMPA glutamate receptors to the postsynaptic cell surface. We now identify the molecular mechanism of how glypican 4 exerts its effect. Glypican 4 induces release of the AMPA receptor clustering factor neuronal pentraxin 1 from presynaptic terminals by signaling through presynaptic protein tyrosine phosphatase receptor ?. Pentraxin then accumulates AMPA receptors on the postsynaptic terminal forming functional synapses. Our findings reveal a signaling pathway that regulates synaptic activity during central nervous system development and demonstrates a role for astrocytes as organizers of active synaptic connections by coordinating both pre and post synaptic neurons. As mutations in glypicans are associated with neurological disorders, such as autism and schizophrenia, this signaling cascade offers new avenues to modulate synaptic function in disease.
Project description:Autism spectrum disorders (ASDs) are characterized by impairments in social behaviors that are sometimes coupled to specialized cognitive abilities. A small percentage of ASD patients carry mutations in genes encoding neuroligins, which are postsynaptic cell-adhesion molecules. We introduced one of these mutations into mice: the Arg451-->Cys451 (R451C) substitution in neuroligin-3. R451C mutant mice showed impaired social interactions but enhanced spatial learning abilities. Unexpectedly, these behavioral changes were accompanied by an increase in inhibitory synaptic transmission with no apparent effect on excitatory synapses. Deletion of neuroligin-3, in contrast, did not cause such changes, indicating that the R451C substitution represents a gain-of-function mutation. These data suggest that increased inhibitory synaptic transmission may contribute to human ASDs and that the R451C knockin mice may be a useful model for studying autism-related behaviors.
Project description:Abnormal pain sensitivity is commonly associated with autism spectrum disorders (ASDs) and affects the life quality of ASD individuals. SHANK3 deficiency was implicated in ASD and pain dysregulation. Here, we report functional expression of SHANK3 in mouse dorsal root ganglion (DRG) sensory neurons and spinal cord presynaptic terminals. Homozygous and heterozygous Shank3 complete knockout (?e4-22) results in impaired heat hyperalgesia in inflammatory and neuropathic pain. Specific deletion of Shank3 in Nav1.8-expressing sensory neurons also impairs heat hyperalgesia in homozygous and heterozygous mice. SHANK3 interacts with transient receptor potential subtype V1 (TRPV1) via Proline-rich region and regulates TRPV1 surface expression. Furthermore, capsaicin-induced spontaneous pain, inward currents in DRG neurons, and synaptic currents in spinal cord neurons are all reduced after Shank3 haploinsufficiency. Finally, partial knockdown of SHANK3 expression in human DRG neurons abrogates TRPV1 function. Our findings reveal a peripheral mechanism of SHANK3, which may underlie pain deficits in SHANK3-related ASDs.