Biology subjects

Giustetto, M.

Publications and source records attributed to Giustetto, M..

6 recordsLinked to original sources

bioRxiv2026

ARHGEF6-dependent cytoskeletal regulation underlies a conserved program of forebrain interneuron development

The molecular programs coordinating inhibitory interneuron migration, maturation, and survival during forebrain development remain incompletely understood. Here we investigate ARHGEF6, a RAC1/CDC42 guanine nucleotide exchange factor linked to X-linked intellectual disability (XLID46) and previously studied only at postsynaptic compartments, and reveal an earlier, conserved role in forebrain interneuron development. ARHGEF6 is selectively enriched in the inhibitory lineage during the peak of interneuron generation and migration. Its loss in mice reduces the number of cortical and hippocampal interneurons, disrupts tangential migration, increases developmental cell death, and impairs morphological and electrophysiological maturation. Strikingly, ARHGEF6-knockout human iPSC-derived organoids and assembloids mirror these deficits exhibiting increased apoptosis, reduced neuronal output, disorganized growth cones, impaired neurite branching, and disrupted migratory dynamics. These cross-species findings reframe ARHGEF6 as an early, essential orchestrator of inhibitory circuit assembly and reveal a conserved cytoskeletal program whose disruption produces the excitatory-inhibitory imbalance linked to cognitive dysfunction.

developmental biology↗

bioRxiv2024

Postnatal Sensory Experience and Barrel Cortex Alterations Anticipate Autistic Traits in a Mouse Model of Cdkl5 Deficiency Disorder

Autistic traits may arise from atypical sensory experience during postnatal life, but whether there is a causal link between defects in cortical circuitry in the brain, altered sensory processing and social behavior remains unknown. Here, we studied tactile stimuli processing in the barrel cortex (BC) and social interactions in juvenile male mice lacking Cyclin-dependent kinase-like 5 (CDKL5), a model of a severe neurodevelopmental disease showing autistic traits and sensory impairments. We identified in these mice defects of whisker-dependent postnatal sensorimotor reflexes, NMDA receptors-dependent signaling, and dendritic orientation in thalamic inputs-receiving spiny stellate neurons. We also found that CDKL5 is required for mapping and processing whisker-derived tactile stimuli in the BC. Intriguingly, KO mice show autistic traits at p21 that are rescued by neonatal CDKL5 replacement in the BC. Our data suggest that CDKL5 is required to link tactile processing in the BC to the onset of social interaction abilities.

neuroscience↗

bioRxiv2024

The CDKL5 kinase undergoes liquid-liquid phase separation driven by a serine-rich C-terminal region and impaired by neurodevelopmental disease-related truncations

Mutations of the cyclin-dependent kinase-like 5 (CDKL5) gene, which encodes a serine/threonine protein kinase, can cause the CDKL5 deficiency disorder (CDD), a severe neurodevelopmental disease characterized by epileptic encephalopathy and neurocognitive impairment. The CDKL5 kinase consists of a catalytic N-terminal domain (NTD) and a less characterized C-terminal domain (CTD). Numerous disease-related mutations truncate CDKL5, leaving the NTD intact while variably shortening the CTD, which highlights the importance of the CTD for CDKL5 function. By systematically analyzing CDKL5 compositional features and evolutionary dynamics, we found that the CTD is a low-complexity region (LCR) highly enriched in serine residues and with a high propensity to undergo liquid-liquid phase separation (LLPS), a biophysical process of condensation controlling protein localization and function. Using a combination of super-resolution imaging, electron microscopy, and molecular and cellular approaches, including optogenetic LLPS induction, we discovered that CDKL5 undergoes LLPS, predominantly driven by its CTD, forming membraneless condensates in neuronal and non-neuronal cells. A CTD internal fragment (CTIF) plays a pivotal LLPS-promoting role, along with the distal portion of the protein. Indeed, two disease-related truncating mutations (S726X and R781X), eliding variable portions of the CTIF, significantly impair LLPS. This impairment is paralleled at the functional level by a reduction in the CDKL5-dependent phosphorylation of EB2, a known CDKL5 target. These findings demonstrate that CDKL5 undergoes LLPS, driven by a CTD region elided by most disease-related truncating mutations. Its loss--through the impairment of CDKL5 LLPS and functional activity--may play a key role in the molecular pathogenesis of CDD.

molecular biology↗

bioRxiv2022

mGluR5 PAMs rescue cortical and behavioural defects in a mouse model of CDKL5 deficiency disorder.

Cyclin-dependent kinase-like 5 (CDKL5) deficiency disorder (CDD) is a devastating rare neurodevelopmental disease without a cure, caused by mutations of the serine/threonine kinase CDKL5 highly expressed in the forebrain. CDD is characterized by early-onset seizures, severe intellectual disabilities, autistic-like traits, sensorimotor and cortical visual impairments (CVI). The lack of an effective therapeutic strategy for CDD urgently demands the identification of novel druggable targets potentially relevant for CDD pathophysiology. To this aim, we studied metabotropic glutamate receptors 5 (mGluR5) for their important role in critical mechanisms involved in CDD, i.e.: synaptogenesis, dendritic spines formation/maturation and synaptic plasticity, and because mGluR5 function depends on the postsynaptic protein Homer1bc that is downregulated in the cerebral cortex of CDKL5-/y mice. In this study, we reveal that CDKL5 loss tampers with (i) the strength of Homer1bc-mGluR5 binding, (ii) the synaptic localization of mGluR5 and (iii) the mGluR5-mediated enhancement of NMDA-induced neuronal responses. Importantly, we showed that the stimulation of mGluR5 activity by administering in mice specific positive-allosteric-modulators, i.e.: 3-Cyano-N-(1,3-diphenyl-1H-pyrazol-5-yl)benzamide (CDPPB) or RO6807794, corrected the synaptic, functional and behavioural defects shown by CDKL5-/y mice. Notably, the cerebral cortex of 2 CDD patients show similar changes in the synaptic organization to mutant CDKL5 mice, including a reduced mGluR5 expression, suggesting that mGluR5 represent a promising therapeutic target for CDD patients.

neuroscience↗

bioRxiv2021

EXPRESSION OF A SECRETABLE, CELL-PENETRATING CDKL5 PROTEIN ENHANCES THE EFFICACY OF AAV VECTOR-MEDIATED GENE THERAPY FOR CDKL5 DEFICIENCY DISORDER.

No therapy is currently available for CDKL5 (cyclin-dependent kinase-like 5) deficiency disorder (CDD), a severe neurodevelopmental disorder caused by mutations in the CDKL5 gene. Although delivery of a wild-type copy of the mutated gene to cells represents the most curative approach for a monogenic disease, proof-of-concept studies highlight significant efficacy caveats for brain gene therapy. Herein, we used a secretable TATk-CDKL5 protein to enhance the efficiency of a gene therapy for CDD. We found that, although AAVPHP.B_Igk-TATk-CDKL5 and AAVPHP.B_CDKL5 vectors had similar brain infection efficiency, the AAVPHP.B_Igk-TATk-CDKL5 vector led to a higher CDKL5 protein replacement due to secretion and transduction of the TATk-CDKL5 protein into the neighboring cells. Importantly, Cdkl5 KO mice treated with the AAVPHP.B_Igk-TATk-CDKL5 vector showed a behavioral and neuroanatomical improvement in comparison with vehicle-treated Cdkl5 KO mice or Cdkl5 KO mice treated with the AAVPHP.B_CDKL5 vector, indicating that a gene therapy based on a secretable recombinant TATk-CDKL5 protein is more effective at compensating Cdkl5-null brain defects than gene therapy based on the expression of the native CDKL5.

neuroscience↗

bioRxiv2018

Modulation of ERK1/MAPK3 potentiates ERK nuclear signalling, facilitates neuronal cell survival and improves memory in mouse models of neurodegenerative disorders

Cell signalling mechanisms are central to neuronal activity and their dysregulation may lead to neurodegenerative processes and associated cognitive decline. So far, a major effort has been directed toward the dissection of disease specific pathways with the still unmet promise to develop precision medicine strategies. With a different approach, here we show that a selective genetic potentiation of neuronal ERK signalling prevents cell death in vitro and in vivo in the mouse brain while ERK attenuation does the opposite. This neuroprotective effect can also be induced pharmacologically by a cell permeable peptide mimicking the loss of ERK1 MAP kinase, leading to a selective enhancement of ERK2 mediated nuclear cell signalling. The drug treatment prevents neurodegeneration in mouse models of Huntingtons (HD), Alzheimers (AD), and Parkinsons disease (PD). Importantly, the selective potentiation of ERK2 signalling facilitates both structural and synaptic plasticity, enhances cognition in healthy mice and rescues mild cognitive impairments in both models of AD and HD. Altogether, our observation truly represents a remarkable example of a shared molecular mechanism across multiple neurodegenerative disorders and a potentially valuable therapeutic target for neuro-enhancement.

neuroscience↗