Biology subjects

Comoletti, D.

Publications and source records attributed to Comoletti, D..

7 recordsLinked to original sources

bioRxiv2026

Engineering antigenic breadth against SARS-CoV-2 by pairing divergent RBDs within a single mRNA immunogen

Vaccines capable of eliciting broadly neutralising antibodies (bnAbs) are a major goal for pandemic preparedness. A persistent challenge across vaccine Wields is how to deliberately recruit the rare B cell clones that recognise conserved epitopes shared across diverse viral variants. BnAbs have been known to frequently emerge through extensive somatic hypermutation during afWinity maturation, here we describe an alternative, structure-driven mechanism for bnAb selection. We designed an mRNA vaccine in which two antigenically distinct SARS-CoV-2 variants (Omicron and Delta; O-{Delta}) receptor binding domains (RBDs) are physically fused on a single polypeptide. This design is predicted to favour B cell antigen receptors capable of engaging conserved epitopes on both RBDs with enhanced avidity. A matched non-divergent tandem RBD (Delta-Delta; {Delta}-{Delta}) served as a control. The divergent (O-{Delta}) immunogen was robustly expressed and retained high-afWinity ACE2 binding. In mice, immunisation elicited potent antibody responses and increased the frequency of antigen-speciWic cross-reactive B cells, recognising Delta, Omicron, and the 2002 pandemic strain SARS-CoV RBDs. Using multicolour RBD tetramers and single-cell B cell receptor sequencing, we show that breadth arises via two distinct pathways. The divergent vaccine preferentially enriches clonally distinct cross-reactive B cells (not present within non-cross-reactive B cell pools) with low levels of somatic hypermutation (SHM), consistent with selection of germline-biased precursors. In contrast, the matched control vaccine yields cross-reactivity primarily within existing clonal lineages (clonal overlap between cross-reactive and non-cross-reactive cells) and at higher mutational burdens, consistent with afWinity-maturation-driven acquisition of breadth. Together, these Windings demonstrate that antigen structure can bias B cell selection towards cross-reactive speciWicities without requiring extensive SHM. This work establishes a simple, modular antigen-design principle in which juxtaposing appropriately divergent antigens on a single scaffold promotes the enrichment of bnAb-prone B cells, providing a scalable strategy for vaccine development against rapidly evolving pathogens.

immunology↗

bioRxiv2025

The trimeric structures of the extracellular domains of FAM171A1 and FAM171A2 neuronal proteins belong to a novel structural superfamily

Cell surface molecules play fundamental roles in cell-cell communication, attraction, or repulsion, and when expressed in neurons they are often implicated in neurological disorders. FAM171 is a family of three type-I transmembrane domain cell surface proteins (FAM171A1, FAM171A2, and FAM171B) expressed in several human tissues and especially enriched in the brain. Recent findings suggest that FAM171A1 transduces signals between the cell surface and the cytoskeleton. Genetic evidence links FAM171A1 to multiple cancers and FAM171A2 to neurodegenerative diseases, including Alzheimers and Parkinsons diseases. Despite multiple connections with severe human diseases, no information is currently available on their monomeric structure or oligomerization. Here we show that, structurally, the monomeric ectodomains of human FAM171A1 and FAM171A2 have a new architecture with a novel combination of two domains. Furthermore, their ectodomains oligomerize to form an equilateral trimer. In addition, the ectodomain of FAM171A1 has the propensity to form larger trimer-trimer assemblies at high concentrations. Together, these results provide novel insights into the structure and oligomerization of the extracellular domain of FAM171A1 and FAM171A2, suggesting important roles in ligand binding and signaling.

neuroscience↗

bioRxiv2022

LRRC37B is a species-specific regulator of voltage-gated channels and excitability in human cortical neurons

The enhanced cognitive abilities characterizing the human species result from specialized features of neurons and circuits, but the underlying molecular mechanisms remain largely unknown. Here we report that the hominid-specific gene LRRC37B encodes a novel receptor expressed in a subset of human cortical pyramidal neurons (CPNs). LRRC37B protein localizes at the axon initial segment (AIS), the specialized domain triggering action potentials. LRRC37B ectopic expression in mouse CPNs in vivo leads to reduced intrinsic excitability, a distinctive feature of some classes of human CPNs. At the molecular level, LRRC37B acts as a receptor for the secreted ligand FGF13A and interacts with the voltage gated sodium channel (VGSC) beta subunit SCN1B, thereby inhibiting the channel function of VGSC, specifically at the AIS. Electrophysiological recordings in adult human cortical slices reveals that endogenous expression of LRRC37B in human CPNs reduces neuronal excitability. LRRC37B thus acts as a species-specific modifier of human cortical neuron function, with important implications for human brain evolution and diseases.

neuroscience↗

bioRxiv2022

GPC3-Unc5D complex structure and role in cell migration

Neural migration is a critical step during brain development that requires the interactions of cell-surface guidance receptors. Cancer cells often hijack these mechanisms to disseminate. Here we reveal crystal structures of Uncoordinated-5 receptor D (Unc5D) in complex with morphogen receptor glypican-3 (GPC3), forming an octameric glycoprotein complex. In the complex, four Unc5D molecules pack into an antiparallel bundle, flanked by four GPC3 molecules. Central glycan-glycan interactions are formed by N-linked glycans emanating from GPC3 (N241 in human) and C-mannosylated tryptophans of the Unc5D thrombospondin-like domains. MD simulations, mass-spectrometry and structure-based mutants validate the crystallographic data. Anti-GPC3 nanobodies enhance or weaken Unc5-GPC3 binding. Using these tools in vivo, we show that Unc5/GPC3 guide migrating pyramidal neurons in the mouse cortex, and cancer cells in an embryonic xenograft neuroblastoma model. The results demonstrate a conserved structural mechanism of cell-guidance, with the potential for wide- ranging biomedical implications in development and cancer biology.

neuroscience↗

bioRxiv2022

MAIT cells activate dendritic cells to promote T follicular helper cell differentiation and humoral immunity

Protective immune responses against respiratory pathogens, including influenza virus are initiated by the mucosal immune system. However, most licensed vaccines are administered parenterally and are largely ineffective at inducing mucosal immunity. The development of safe and effective mucosal vaccines has largely been hampered by the lack of a suitable mucosal adjuvant. In this study we explore a novel class of adjuvant that harness mucosal-associated invariant T (MAIT) cells. We show evidence that intranasal immunisation of MAIT cell agonists co-administered with protein, including haemagglutinin from influenza A virus induced potent humoral immunity and immunoglobulin (Ig)A production, which protected mice against infection. MAIT cell adjuvant activity was mediated by CD40L-dependent activation of dendritic cells and subsequent priming of CD4+ T follicular helper cells. In summary, we show that MAIT cells are promising vaccine targets that can be utilised as cellular adjuvants in mucosal vaccines.

immunology↗

bioRxiv2021

Analyses of the Autism-associated Neuroligin-3 R451C Mutation in Human Neurons Reveals a Gain-of-Function Synaptic Mechanism

Mutations in many synaptic genes are associated with autism spectrum disorders (ASDs), suggesting that synaptic dysfunction is a key driver of ASD pathogenesis. Among these mutations, the R451C-substitution in the NLGN3 gene that encodes the postsynaptic adhesion molecule Neuroligin-3 is noteworthy because it was the first specific mutation linked to ASDs. In mice, the corresponding Nlgn3 R451C-knockin mutation recapitulates social interaction deficits of ASD patients and produces synaptic abnormalities, but the impact of the NLGN3 R451C-mutation on human neurons has not been investigated. Here, we generated human knock-in neurons with the NLGN3 R451C-mutation. Strikingly, analyses of NLGN3 R451C-mutant neurons revealed that the R451C-mutation decreased NLGN3 protein levels but enhanced the strength of excitatory synapses without affecting inhibitory synapses. No significant cell death and endoplasmic reticulum stress were detected. Importantly, the augmentation of excitatory transmission was confirmed in vivo with human neurons transplanted into mouse forebrain. Using single-cell RNA-seq experiments with co-cultured excitatory and inhibitory NLGN3 R451C-mutant neurons, we identified differentially expressed genes in relatively mature human neurons that corresponded to synaptic gene expression networks. Moreover, gene ontology and enrichment analyses revealed convergent gene networks associated with ASDs and other mental disorders. Our findings suggest that the NLGN3 R451C-mutation induces a gain-of-function enhancement in excitatory synaptic transmission that may contribute to the pathophysiology of ASDs.

neuroscience↗

bioRxiv2019

CNTNAP2 ectodomain, detected in neuronal and CSF sheddomes, modulates Ca2+ dynamics and network synchrony

SUMMARYWhile many neuronal membrane-anchored proteins undergo proteolytic cleavage, little is known about the biological significance of neuronal ectodomain shedding. Using mass spectrometry (MS)-based proteomics, we showed that the neuronal sheddome mirrors human cerebrospinal fluid (hCSF). Among shed synaptic proteins in hCSF was the ectodomain of CNTNAP2 (CNTNAP2-ecto), a risk factor for neurodevelopmental disorders (NDD). Using structured-illumination microscopy (SIM), we mapped the spatial organization of neuronal CNTNAP2-ecto shedding. Using affinity chromatography followed by MS, we identified the ATP2B/PMCA Ca2+ extrusion pumps as novel CNTNAP2-ecto binding partners. CNTNAP2-ecto coimmunoprecipitates with PMCA2, a known autism risk factor, and enhances its activity, thereby modulating neuronal Ca2+ levels. Finally, we showed that CNTNAP2-ecto regulates neuronal network synchrony in primary cultures and brain slices. These data provide new insights into the biology of synaptic ectodomain shedding and reveal a novel mechanism of regulation of Ca2+ homeostasis and neuronal network synchrony.

neuroscience↗