The mammalian cerebral cortex is a complex laminar structure with a variety of neuronal and non-neuronal cell types that develop in a finely orchestrated and stereotypic manner. Final laminar position and synaptic specificity of most cortical cell types are well described. Strikingly, any alteration in the developmental unfolding of one of these processes, even for a single cell type among tens, can be sufficient to generate neurodevelopmental disorders. However, how the establishment of this precise cellular architecture is regulated at the molecular level remains largely unknown. Several lines of evidence suggest a role of cell-cell communications via ligand-receptor (LR) interactions. Using a single cell RNA-seq (scRNA-seq) approach in mice, we have generated a bioinformatic atlas that infers LR based cellular communications across all cell types over somatosensory (SS) cortex development. Querying our atlas for known LR interactions has demonstrated its validity, but new LR-mediated cell-cell interactions remain to be discovered to interrogate its power as a hypothesis generator. In parallel, a technique called Multiplexed-Error Robust Fluorescence In Situ Hybridization (MERFISH) has been recently developed and implemented by us, which images single cell transcriptomes in situ and thereby adds precious information about spatial expression. Here, we will: (i) test some LR interactions predicted by our scRNA-seq atlas for a role in SS microcircuit development, (ii) build on the MERFISH technique to complement our atlas with spatial information and to characterize SS cortex cellular development with unprecedented resolution and (iii) use MERFISH to interrogate altered developmental processes in the SS cortex of a mouse model of neurodevelopmental disorder, the Neurod2 KO mouse.

Copper ions are essential for life but posses a redox-activity which makes them potentially toxic, and their cellular availability is highly regulated by an intricate network of intracellular chaperones, transcription factors and membrane transporters. Copper homeostatic imbalance is connected to several major neurological diseases. The detailed mechanisms of copper movement across membranes remain unknown due to the difficulty to characterize at atomic level the different proteins involved, which are mainly integral membrane systems. In humans, high-affinity copper uptake is modulated by hCTR1, a trimeric membrane transporter which has so far fled from high-resolution x-ray or cryo-EM investigations and is extremely challenging to produce and recover in workable amounts for structural studies. The central objective of the present project is to develop and apply a solid-state Magic-Angle Spinning (MAS) NMR approach to allow complete characterization of the structure and mechanism of lipid-bound hCTR1. Building on a decade of continuous advances of the NMR community, the recent development of very fast (up to 100 kHz) MAS probes has revolutionised this field, with developments that speed up the analysis of proteins of considerable size and open the way to complex biological solids available in limited amounts. We propose to leverage the unique expertise and equipment available in the consortium, and achieve the objectives above through a combination of innovative strategies for isotopic sample preparation, advanced spectroscopic tools to obtain NMR signatures of the structure and dynamics, and new instrumentation capable of even faster MAS rates. The project will provide breakthrough data for understanding structure-activity relationships in a challenging integral membrane protein, and will allow the addition of solid-state NMR to the method portfolio for the characterization of medically relevant targets.

We propose to decipher the mechanisms of action of neuroendothelial N-methyl-D-aspartate receptors (NMDAR). In addition to neurons, where they drive glutamatergic neurotransmission, NMDAR are expressed in a variety of cell types. In particular, brain endothelial cells express NMDAR, which could be involved in blood-brain barrier maintenance and alteration. In a recent paper, we identified an unexpected population of NMDAR in endothelial cells, expressed at the luminal side of microvessels and located at the vicinity of blood/spinal cord barrier-forming tight junctions. We developed a monoclonal antibody (Glunomab®), directed against a specific site of NMDAR (aminoacids 176-180), which blocked the entry of leukocytes to the inflamed spinal cord, thus providing therapeutic effects in experimental autoimmune encephalomyelitis (EAE), a mouse model of multiple sclerosis (MS). Nevertheless, the downstream targets which link endothelial NMDAR function to leukocyte migration across the blood/brain and blood/spinal cord barriers are not fully understood yet. Interestingly, our current studies show that these receptors have an unconventional composition, including the presence of the rare GluN3 subunit, which provides response to glycine (in addition to glutamate) and metabotropic signaling (in addition to ionotropic function). Interestingly, naturally occurring circulating auto-antibodies against NMDAR are present in ~10% of human subjects and are overexpressed in neuropsychiatric and neurological diseases. Beyond these quantitative data, qualitative information are needed concerning the regions of NMDAR recognized by these antibodies. In fact, circulating autoantibodies against NMDAR could provide either beneficial or deleterious effects, depending on the region that they target. In line with this, we postulate that identifying the regions targeted by NMDAR auto-antibodies could have prognosis value in neurological diseases. Given our recent work concerning NMDAR in animal models of MS, we believe that investigating circulating antibodies would be particularly relevant for prognosis of MS. Thus the goals of this project are i) To characterize the signaling pathways and downstream targets triggered by NMDAR activation in endothelial cells, in relation to leukocyte penetration towards the spinal cord, ii) To identify the repertoire of NMDAR antibodies in MS patients (based on their target regions on NMDAR) and to determine whether different clusters of antibodies are associated with different outcomes, iii) To investigate the effects of these different clusters on the function of endothelial NMDAR and in animal models of MS, and iv) To bridge experimental data and clinical observations.

More than two decades ago has emerged a new technology exploiting electron spins to convey information within an electric circuitry. This technology, known as spintronics, translates in advantages such as nonvolatile storage technology, fast-data processing speed and low-power consumption. The working principle of a spintronic device is to generate a non-equilibrium spin population and to detect it. However, creation and detection occurs in different regions of the device. During the transfer process from one region to the other, the spin population tends to relax towards its non spin-polarized equilibrium state weakening then the efficiency of the device. One of the central research areas in spintronics therefore aims at perfecting this transfer process. Present efforts involve improving existing technology or finding novel radical ways of manipulating spin-polarized electrons. The SPINCOMM project is in line with this second approach and falls in the context of molecular spintronics. The purpose of project SPINCOMM is to carry out the first fundamental investigation of spin transport across a single organometallic wire. To achieve this ambitious goal, a pioneering bottom-up approach will be implemented through four innovating strategies: 1) SIMPLIFICATION: The wires will have a multi-decker architecture where single transition-metal atoms alternate with cyclopentadienyl rings (C5H5). Strikingly, these wires have been predicted to display a 100% spin-filtering efficiency over a wide bias range. 2) CONTROL: Transport measurements will be carried out with a low-temperature scanning tunneling microscope (STM) operated in ultrahigh vacuum. The molecules will be deposited onto a well-calibrated surface and then contacted by the STM tip. Junction formation with a single multi-decker molecule will be greatly facilitated by the upstanding adsorption geometry onto the surface. Precise information about the binding properties of the multi-decker molecule to the electrodes will be available. XMCD measurements will be carried out independently to carefully characterize the magnetic status of the molecules. 3) CUSTOMIZATION: The chemical composition of the molecule and its length will be modified directly in the STM junction to optimize spin transport. Moreover, the material of tip and surface will be changed in order to tackle different aspects of spin transport. These essentially consist in the Kondo effect (non-magnetic tip and surface) and its interplay with spin-polarized electrons (ferromagnetic tip and a non-magnetic surface), as well as a transport across a single-molecule spin-valve (ferromagnetic tip and surface). 4) SIMULATION: Given the unprecedented microscopic control exerted over the junction and the simplified molecular architecture employed, the experimental data will be highly amenable to first-principle calculations. State-of-the-art density functional theory and transport calculations will be used to unravel the key mechanisms governing spin transport, along with non-equilibrium and correlated calculations to treat the Kondo problem. With the know-how acquired, the mono-decker architecture of the molecule will be exploited for developing a new spin-sensitive microscopy. A molecular tip comprising a mono-decker molecule will be used to record “contact images” of the surface. Surfaces with opposite magnetizations are expected to produce a higher contrast than the one accessible to SP-STM due to the nearly ideal spin-filtering effect of the mono-decker molecule. With spin-polarized contact microscopy it will be possible to map the spin-polarized properties of surfaces and nanostructures with atomic-scale spatial resolution and to assess the impact of defects, surface impurities, and electronic inhomogeneities on spin transport. We expect this technique to develop quickly and to have a success similar to one of SP-STM in these last ten years.

This project focuses on three iron-base alloys that have growing potential for high-temperature, high-strength and strong- magnet applications: Fe-Cr, Fe-Mn and Fe-Co. Because of the key role of magnetism, an innovative materials design based on advanced modeling approaches is necessary to control key properties of these materials. Such a design strategy requires the combination of (i) highly accurate methods to determine atomic features with (ii) efficient coarse-graining techniques to access target physical properties and to perform the screening of materials compositions. For the former, density functional theory (DFT) has for many materials classes already proven to be a highly successful tool. For Fe-based alloys, however, a critical bottleneck is the role that magnetic ordering, excitations and transitions have on thermodynamic, defect and kinetic properties. Therefore, a complete and accurate modeling of magnetism is urgently needed to address the materials-design challenges: the resistance to radiation damage related to the chemical decomposition in Fe-Cr, the grain-boundary embrittlement in ferritic Fe-Mn and the high-strength of austenitic Fe-Mn, and the phase ordering and the relative stability of a and ? phases in Fe-Co cannot be fully understood without properly accounting for the magnetic effects. The novelty of the current approach is twofold: First, on the DFT-side, we will make use of the recent important progress in treating magnetism in pure idealized Fe lattices, in order to go towards an accurate modeling of magnetic multi-component systems with point/extended defects, and beyond the standard collinear approximation. Second, we will develop new methods that allow us to bridge the gap between (i) highly accurate electronic calculations and (ii) large-scale atomistic thermodynamic and kinetic simulations for iron based alloys by – and this is decisive – fully taking into account the impact of magnetism on defect properties, diffusion and microstructural evolution. For the latter, lattice-based effective interaction models (EIMs) and tight-binding (TB) models will be developed based on data from DFT, including magnetic configurations, excitations and transitions. This will allow us to provide a coherent description of the role of magnetism on various properties of Fe-based alloys at different length scales and at finite temperature. It will further give us the ability to perform the optimization of key parameters controlling the relevant properties like phase decomposition in Fe-Cr, phase ordering in Fe-Co or decohesion of grain boundaries in Fe-Mn. Dedicated experiments in bulk alloys and along intergranular / interphase boundaries grown on demand will be performed in the project, which are essential for verifying the robustness of the theoretical predictions. The three chosen alloys exhibit a large variety of magnetic behavior. The methods developed and applied in this proposal are therefore expected to be transferrable to the modeling of other magnetic materials. The results of our simulations will lead to the improvement of thermo¬dynamic and diffusion databases and tools (such as DICTRA) that are nowadays routinely used in industrial R&D but that at present have difficulties in accounting for magnetism. In this way a better and more systematic understanding of the role of magnetism in Fe-based alloys will help to improve significantly the predictive power of the simulations and thus contribute to a more efficient and accurate development of new steel grades. Once fully implemented, the availability of such computational tools is expected to boost the efficiency, change the strategy in designing new steel grades and to form an important contribution for the future competitiveness of steelmakers.