2023
SELECTED PUBLICATIONS

DECEMBER 2023

The RhoGEF Trio is transported by microtubules and affects microtubule stability in migrating neural crest cells

Stefanie Gossen; Sarah Gerstner, Annette Borchers

Abstract

Directed cell migration requires a local fine-tuning of Rho GTPase activity to control protrusion formation, cell-cell contraction, and turnover of cellular adhesions. The Rho guanine nucleotide exchange factor (GEF) TRIO is ideally suited to control RhoGTPase activity because it combines two distinct catalytic domains to control Rac1 and RhoA activity in one molecule. However, at the cellular level, this molecular feature also requires a tight spatiotemporal control of TRIO activity. Here, we analyze the dynamic localization of Trio in Xenopus cranial neural crest (NC) cells, where we have recently shown that Trio is required for protrusion formation and migration. Using live cell imaging, we find that the GEF2 domain, but not the GEF1 domain of Trio, dynamically colocalizes with EB3 at microtubule plus-ends. Microtubule-mediated transport of Trio appears to be relevant for its function in NC migration, as a mutant GEF2 construct lacking the SxIP motif responsible for microtubule plus-end localization was significantly impaired in its ability to rescue the Trio loss-of-function phenotype compared to wild-type GEF2. Furthermore, by analyzing microtubule dynamics in migrating NC cells, we observed that loss of Trio function stabilized microtubules at cell-cell contact sites compared to controls, whereas they were destabilized at the leading edge of NC cells. Our data suggest that Trio is transported by microtubules to distinct subcellular locations where it has different functions in controlling microtubule stability, cell morphology, and cell-cell interaction during directed NC migration.

MAY 2023

Comparing synaptic proteomes across five mouse models for autism reveals converging molecular similarities including deficits in oxidative phosphorylation and Rho GTPase signaling

Abigail U. Carbonell, Carmen Freire-Cobo, Ilana V. Deyneko, Saunil Dobariya, Hediye Erdjument-Bromage, Amy E. Clipperton-Allen, Damon T. Page, Thomas A. Neubert and Bryen A. Jordan

Abstract

Specific and effective treatments for autism spectrum disorder (ASD) are lacking due to a poor understanding of disease mechanisms. Here we test the idea that similarities between diverse ASD mouse models are caused by deficits in common molecular pathways at neuronal synapses. To do this, we leverage the availability of multiple genetic models of ASD that exhibit shared synaptic and behavioral deficits and use quantitative mass spectrometry with isobaric tandem mass tagging (TMT) to compare their hippocampal synaptic proteomes. Comparative analyses of mouse models for Fragile X syndrome (Fmr1 knockout), cortical dysplasia focal epilepsy syndrome (Cntnap2 knockout), PTEN hamartoma tumor syndrome (Pten haploinsufficiency), ANKS1B syndrome (Anks1b haploinsufficiency), and idiopathic autism (BTBR+) revealed several common altered cellular and molecular pathways at the synapse, including changes in oxidative phosphorylation, and Rho family small GTPase signaling. Functional validation of one of these aberrant pathways, Rac1 signaling, confirms that the ANKS1B model displays altered Rac1 activity counter to that observed in other models, as predicted by the bioinformatic analyses. Overall similarity analyses reveal clusters of synaptic profiles, which may form the basis for molecular subtypes that explain genetic heterogeneity in ASD despite a common clinical diagnosis. Our results suggest that ASD-linked susceptibility genes ultimately converge on common signaling pathways regulating synaptic function and propose that these points of convergence are key to understanding the pathogenesis of this disorder.

March 2023

TRIO-Related Neurodevelopmental Disorder

Konstantinos Varvagiannis, Diana Baralle, Bert BA de Vries, Gabriella Gazdagh.

Extending the phenotypes associated with TRIO gene variants in a cohort of 25 patients and review of the literature

Gabriella Gazdagh, David Hunt, Anna Maria Cueto Gonzalez, Monserrat Pons Rodriguez, Ayeshah Chaudhry, Marcos Madruga, Fleur Vansenne, Deborah Shears, Aurore Curie, Eva-Lena Stattin, Britt-Marie Anderlid, Slavica Trajkova, Elena Sukarova Angelovska, Catherine McWilliam, Philip R. Wyatt, Mary O'Driscoll, Giles Atton, Anke K. Bergman, Pia Zacher, Leena D. Mewasingh, Antonio Gonzalez-Meneses Lopez, Olga Alonso-Luengo, Htoo A. Wai, Ottilie Rohde, Pauline Boiroux, Anne Debant, Susanne Schmidt, Diana Baralle

Abstract

The TRIO gene encodes a rho guanine exchange factor, the function of which is to exchange GDP to GTP, and hence to activate Rho GTPases, and has been described to impact neurodevelopment. Specific genotype-to-phenotype correlations have been established previously describing striking differentiating features seen in variants located in specific domains of the TRIO gene that are associated with opposite effects on RAC1 activity. Currently, 32 cases with a TRIO gene alteration have been published in the medical literature. Here, we report an additional 25, previously unreported individuals who possess heterozygous TRIO variants and we review the literature. In addition, functional studies were performed on the c.4394A > G (N1465S) and c.6244-2A > G TRIO variants to provide evidence for their pathogenicity. Variants reported by the current study include missense variants, truncating nonsense variants, and an intragenic deletion. Clinical features were previously described and included developmental delay, learning difficulties, microcephaly, macrocephaly, seizures, behavioral issues (aggression, stereotypies), skeletal problems including short, tapering fingers and scoliosis, dental problems (overcrowding/delayed eruption), and variable facial features. Here, we report clinical features that have not been described previously, including specific structural brain malformations such as abnormalities of the corpus callosum and ventriculomegaly, additional psychological and dental issues along with a more recognizable facial gestalt linked to the specific domains of the TRIO gene and the effect of the variant upon the function of the encoded protein. This current study further strengthens the genotype-to-phenotype correlation that was previously established and extends the range of phenotypes to include structural brain abnormalities, additional skeletal, dental, and psychiatric issues.Abstract

JANUARY 2023

Pathogenic TRIO variants associated with neurodevelopmental disorders perturb the molecular regulation of TRIO and axon pathfinding in vivo

Maxime Bonnet, Fiona Roche, Christine Fagotto-Kaufmann, Gabriella Gazdagh, Iona Truong, Franck Comunale, Sonia Barbosa, Marion Bonhomme, Nicolas Nafati, David Hunt, Monserrat Pons Rodriguez, Ayeshah Chaudhry, Deborah Shears, Marcos Madruga, Fleur Vansenne, Aurore Curie, Andrey V. Kajava, Diana Baralle, Coralie Fassier, Anne Debant & Susanne Schmidt 

Abstract

The RhoGEF TRIO is known to play a major role in neuronal development by controlling actin cytoskeleton remodeling, primarily through the activation of the RAC1 GTPase. Numerous de novo mutations in the TRIO gene have been identified in individuals with neurodevelopmental disorders (NDDs). We have previously established the first phenotype/genotype correlation in TRIO-associated diseases, with striking correlation between the clinical features of the individuals and the opposite modulation of RAC1 activity by TRIO variants targeting different domains. The mutations hyperactivating RAC1 are of particular interest, as they are recurrently found in patients and are associated with a severe form of NDD and macrocephaly, indicating their importance in the etiology of the disease. Yet, it remains unknown how these pathogenic TRIO variants disrupt TRIO activity at a molecular level and how they affect neurodevelopmental processes such as axon outgrowth or guidance. Here we report an additional cohort of individuals carrying a pathogenic TRIO variant that reinforces our initial phenotype/genotype correlation. More importantly, by performing conformation predictions coupled to biochemical validation, we propose a model whereby TRIO is inhibited by an intramolecular fold and NDD-associated variants relieve this inhibition, leading to RAC1 hyperactivation. Moreover, we show that in cultured primary neurons and in the zebrafish developmental model, these gain-of-function variants differentially affect axon outgrowth and branching in vitro and in vivo, as compared to loss-of-function TRIO variants. In summary, by combining clinical, molecular, cellular and in vivo data, we provide compelling new evidence for the pathogenicity of novel genetic variants targeting the TRIO gene in NDDs. We report a novel mechanism whereby the fine-tuned regulation of TRIO activity is critical for proper neuronal development and is disrupted by pathogenic mutations.

Both GEF domains of the autism and epilepsy-associated Trio protein are required for proper tangential migration of GABAergic interneurons

Lara Eid, Ludmilla Lokmane, Praveen K. Raju, Samuel Boris Tene Tadoum, Xiao Jiang, Karolanne Toulouse, Alexis Lupien-Meilleur, François Charron-Ligez, Asmaa Toumi, Stéphanie Backer, Mathieu Lachance, Marisol Lavertu-Jolin, Marie Montseny, Jean-Claude Lacaille, Evelyne Bloch-Gallego, Elsa Rossignol

Abstract

Recessive mutations in the TRIO gene are associated with intellectual deficiency (ID), autism spectrum disorder (ASD) and developmental epileptic encephalopathies (DEE). TRIO is a dual guanine nucleotide exchange factor (GEF) that activates Rac1, Cdc42 and RhoA.

Trio has been extensively studied in excitatory neurons, and has recently been found to regulate the switch from tangential to radial migration in GABAergic interneurons (INs), through GEFD1-Rac1-dependent SDF1α/CXCR4 signalling. Given the central role of Rho-GTPases during neuronal migration and the implication of IN pathologies in ASD and DEE, we investigated the relative roles of both Trio’s GEF domains in regulating the dynamics of INs tangential migration. In Trio-/- mice, we observed reduced numbers of tangentially migrating INs, with intact progenitor proliferation. Further, we noted increased growth cone collapse in developing INs, suggesting altered cytoskeleton dynamics. To bypass the embryonic mortality of Trio-/- mice, we generated Dlx5/6Cre;Trioc/c conditional mutant mice, which develop spontaneous seizures and behavioral deficits reminiscent of ASD and ID. These phenotypes are associated with reduced cortical IN density and functional cortical inhibition. Mechanistically, this reduction of cortical IN numbers reflects a premature switch to radial migration, with an aberrant early entry in the cortical plate, as well as major deficits in cytoskeletal dynamics, including enhanced leading neurite branching and slower nucleokinesis reflecting reduced actin filament condensation and turnover. Further, we show that both Trio GEFD1 and GEFD2 domains are required for proper IN migration, with a dominant role of the RhoA-activating GEFD2 domain.

Altogether, our data show a critical role of the DEE/ASD-associated Trio gene in the establishment of cortical inhibition and the requirement of both GEF domains in regulating IN migration dynamics.