Autism Spectrum Disorder/Intellectual Disability-Associated Mutations in Trio Disrupt Neuroligin 1-Mediated Synaptogenesis.

We recently identified an autism spectrum disorder/intellectual disability (ASD/ID)-related de novo mutation hotspot in the Rac1-activating GEF1 domain of the protein Trio. Trio is a Rho guanine nucleotide exchange factor (RhoGEF) that is essential for glutamatergic synapse function. An ASD/ID-related mutation identified in Trio's GEF1 domain, Trio D1368V, produces a pathologic increase in glutamatergic synaptogenesis, suggesting that Trio is coupled to synaptic regulatory mechanisms that govern glutamatergic synapse formation. However, the molecular mechanisms by which Trio regulates glutamatergic synapses are largely unexplored. Here, using biochemical methods, we identify an interaction between Trio and the synaptogenic protein Neuroligin 1 (NLGN1) in the brain. Molecular biological approaches were then combined with super-resolution dendritic spine imaging and whole-cell voltage-clamp electrophysiology in hippocampal slices from male and female rats to examine the impact ASD/ID-related Trio mutations have on NLGN1-mediated synaptogenesis. We find that an ASD/ID-related mutation in Trio's eighth spectrin repeat region, Trio N1080I, inhibits Trio's interaction with NLGN1 and prevents Trio D1368V-mediated synaptogenesis. Inhibiting Trio's interaction with NLGN1 via Trio N1080I blocked NLGN1-mediated synaptogenesis and increases in synaptic NMDA Receptor function but not NLGN1-mediated increases in synaptic AMPA Receptor function. Finally, we show that the aberrant synaptogenesis produced by Trio D1368V is dependent on NLGN signaling. Our findings demonstrate that ASD/ID-related mutations in Trio are able to pathologically increase as well as decrease NLGN-mediated effects on glutamatergic neurotransmission, and point to an NLGN1-Trio interaction as part of a key pathway involved in ASD/ID etiology.SIGNIFICANCE STATEMENT A number of genes have been implicated in the development of autism spectrum disorder/intellectual disability (ASD/ID) in humans. It is now important to identify relationships between these genes to uncover specific cellular regulatory pathways that contribute to these disorders. In this study, we discover that two glutamatergic synapse regulatory proteins implicated in ASD/ID, Trio and Neuroligin 1, interact with one another to promote glutamatergic synaptogenesis. We also identify ASD/ID-related mutations in Trio that either inhibit or augment Neuroligin 1-mediated glutamatergic synapse formation. Together, our results identify a synaptic regulatory pathway that, when disrupted, likely contributes to the development of ASD/ID. Going forward, it will be important to determine whether this pathway represents a point of convergence of other proteins implicated in ASD/ID.

IL4I1 augments CNS remyelination and axonal protection by modulating T cell driven inflammation.

SEE PLUCHINO AND PERUZZOTTI-JAMETTI DOI101093/AWW266 FOR A SCIENTIFIC COMMENTARY ON THIS ARTICLE: Myelin regeneration (remyelination) is a spontaneous process that occurs following central nervous system demyelination. However, for reasons that remain poorly understood, remyelination fails in the progressive phase of multiple sclerosis. Emerging evidence indicates that alternatively activated macrophages in central nervous system lesions are required for oligodendrocyte progenitor differentiation into remyelinating oligodendrocytes. Here, we show that an alternatively activated macrophage secreted enzyme, interleukin-four induced one (IL4I1), is upregulated at the onset of inflammation resolution and remyelination in mouse central nervous system lesions after lysolecithin-induced focal demyelination. Focal demyelination in mice lacking IL4I1 or interleukin 4 receptor alpha (IL4Rα) results in increased proinflammatory macrophage density, remyelination impairment, and axonal injury in central nervous system lesions. Conversely, recombinant IL4I1 administration into central nervous system lesions reduces proinflammatory macrophage density, enhances remyelination, and rescues remyelination impairment in IL4Rα deficient mice. We find that IL4I1 does not directly affect oligodendrocyte differentiation, but modulates inflammation by reducing interferon gamma and IL17 expression in lesioned central nervous system tissues, and in activated T cells from splenocyte cultures. Remarkably, intravenous injection of IL4I1 into mice with experimental autoimmune encephalomyelitis at disease onset significantly reversed disease severity, resulting in recovery from hindlimb paralysis. Analysis of post-mortem tissues reveals reduced axonal dystrophy in spinal cord, and decreased CD4+ T cell populations in spinal cord and spleen tissues. These results indicate that IL4I1 modulates inflammation by regulating T cell expansion, thereby permitting the formation of a favourable environment in the central nervous system tissue for remyelination. Therefore, IL4I1 is a potentially novel therapeutic for promoting central nervous system repair in multiple sclerosis.

Kalirin and Trio: RhoGEFs in Synaptic Transmission, Plasticity, and Complex Brain Disorders.

Changes in the actin cytoskeleton are a primary mechanism mediating the morphological and functional plasticity that underlies learning and memory. The synaptic Ras homologous (Rho) guanine nucleotide exchange factors (GEFs) Kalirin and Trio have emerged as central regulators of actin dynamics at the synapse. The increased attention surrounding Kalirin and Trio stems from the growing evidence for their roles in the etiology of a wide range of neurodevelopmental and neurodegenerative disorders. In this Review, we discuss recent findings revealing the unique and diverse functions of these paralog proteins in neurodevelopment, excitatory synaptic transmission, and plasticity. We additionally survey the growing literature implicating these proteins in various neurological disorders.

Synaptic Kalirin-7 and Trio Interactomes Reveal a GEF Protein-Dependent Neuroligin-1 Mechanism of Action.

The RhoGEFs Kalirin-7 and Trio are regulators of synaptic plasticity, and their dysregulation is associated with a range of neurodevelopmental and neurodegenerative disorders. Although studies have implicated both Kalirin and Trio in certain diseases, such as tauopathies, they remarkably differ in their association with other disorders. Using unbiased proteomics, we identified interactomes of Kalirin-7 and Trio to ascertain distinct protein association networks associated with their respective function and revealed groups of proteins that preferentially interact with a particular RhoGEF. In comparison, we find Trio interacts with a range of axon guidance and presynaptic complexes, whereas Kalirin-7 associates with several synaptic adhesion molecules. Specifically, we show Kalirin-7 is an interactor of the cell adhesion molecule neuroligin-1 (NLGN1), and NLGN1-dependent synaptic function is mediated through Kalirin-7 in an interaction-dependent manner. Our data reveal not only the interactomes of two important disease-related proteins, but also provide an intracellular effector of NLGN1 function.

Posttranslational modifications of neuroligins regulate neuronal and glial signaling.

This review covers the dynamic regulation of neuroligin isoforms, focusing on posttranslational events including phosphorylation, glycosylation and activity-dependent cleavage. There is a growing literature on how phosphorylation confers an isoform-specific level of modulation affecting a variety of protein interactions. In addition, recent studies describe activity-dependent proteolytic cleavage of neuroligins, revealing a broader role for neuroligins than just synaptic 'glue'. Interesting new research implicates the cleaved extracellular fragments of neuroligins in promoting glioma. These reports on cell signaling mediated by the cleavage products of neuroligins suggest novel and important roles for neuroligins in neuro-glial signaling.