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A hexanucleotide-repeat expansion in C9ORF72 is the most common genetic variant that contributes to amyotrophic lateral sclerosis and frontotemporal dementia 1 , 2 . The C9ORF72 mutation acts through gain- and loss-of-function mechanisms to induce pathways that are implicated in neural degeneration 3 – 9 . The expansion is transcribed into a long repetitive RNA, which negatively sequesters RNA-binding proteins 5 before its non-canonical translation into neural-toxic dipeptide proteins 3 , 4 . The failure of RNA polymerase to read through the mutation also reduces the abundance of the endogenous C9ORF72 gene product, which functions in endolysosomal pathways and suppresses systemic and neural inflammation 6 – 9 . Notably, the effects of the repeat expansion act with incomplete penetrance in families with a high prevalence of amyotrophic lateral sclerosis or frontotemporal dementia, indicating that either genetic or environmental factors modify the risk of disease for each individual. Identifying disease modifiers is of considerable translational interest, as it could suggest strategies to diminish the risk of developing amyotrophic lateral sclerosis or frontotemporal dementia, or to slow progression. Here we report that an environment with reduced abundance of immune-stimulating bacteria 10 , 11 protects C9orf72 -mutant mice from premature mortality and significantly ameliorates their underlying systemic inflammation and autoimmunity. Consistent with C9orf72 functioning to prevent microbiota from inducing a pathological inflammatory response, we found that reducing the microbial burden in mutant mice with broad spectrum antibiotics—as well as transplanting gut microflora from a protective environment—attenuated inflammatory phenotypes, even after their onset. Our studies provide further evidence that the microbial composition of our gut has an important role in brain health and can interact in surprising ways with well-known genetic risk factors for disorders of the nervous system. Reduced abundance of immune-stimulating gut bacteria ameliorated the inflammatory and autoimmune phenotypes of mice with mutations in C9orf72 , which in the human orthologue are linked to amyotrophic lateral sclerosis and frontotemporal dementia.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by a progressive loss of motor function linked to degenerating extratelencephalic neurons/Betz cells (ETNs). The reasons why these neurons are selectively affected remain unclear. Here, to understand the unique molecular properties that may sensitize ETNs to ALS, we performed RNA sequencing of 79,169 single nuclei from cortices of patients and controls. In both patients and unaffected individuals, we found significantly higher expression of ALS risk genes in THY1 + ETNs, regardless of diagnosis. In patients, this was accompanied by the induction of genes involved in protein homeostasis and stress responses that were significantly induced in a wide collection of ETNs. Examination of oligodendroglial and microglial nuclei revealed patient-specific downregulation of myelinating genes in oligodendrocytes and upregulation of an endolysosomal reactive state in microglia. Our findings suggest that selective vulnerability of extratelencephalic neurons is partly connected to their intrinsic molecular properties sensitizing them to genetics and mechanisms of degeneration.

Amyotrophic Lateral Sclerosis (ALS) is a fatal neurodegenerative disorder characterised by a progressive loss of motor function. The eponymous spinal sclerosis observed at autopsy is the result of the degeneration of extratelencephalic neurons, Betz cells (ETNs, Cortico-Spinal Motor Neuron). It remains unclear why this neuronal subtype is selectively affected. To understand the unique molecular properties that sensitise these cells to ALS, we performed RNA sequencing of 79,169 single nuclei from cortices of patients and controls. In unaffected individuals, we found that expression of ALS risk genes was significantly enriched in THY1+-ETNs and not in other cell types. In patients, these genetic risk factors, as well as genes involved in protein homeostasis and stress responses, were significantly induced in a wide collection of ETNs, but not in neurons with more superficial identities. Examination of oligodendroglial and microglial nuclei revealed patient-specific changes that were at least in part a response to alterations in neurons: downregulation of myelinating genes in oligodendrocytes and upregulation of a reactive state connected to endo-lysosomal pathways in microglia. Our findings suggest that the selective vulnerability of extratelencephalic neurons is partly connected to their intrinsic molecular properties sensitising them to genetics and mechanisms of degeneration.Competing Interest StatementThe authors have declared no competing interest.Footnotes* New figures and text

Human pluripotent stem cells (hPSCs) are a powerful tool for disease modelling and drug discovery, especially when access to primary tissue is limited, such as in the brain. Current neuronal differentiation approaches use either small molecules for directed differentiation or transcription-factor-mediated programming. In this study we coupled the overexpression of the neuralising transcription factor Neurogenin2 (Ngn2) with small molecule patterning to differentiate hPSCs into lower induced Motor Neurons (liMoNes). We showed that this approach induced activation of the motor neuron (MN) specific transcription factor Hb9/MNX1, using anHb9::GFP-reporter line, with up to 95% of cells becoming Hb9::GFP+. These cells acquired and maintained expression of canonical early and mature MN markers. Molecular and functional profiling revealed that liMoNes resembled bona fide hPSC-derived MN differentiated by conventional small molecule patterning. liMoNes exhibited spontaneous electrical activity, expressed synaptic markers and formed contacts with muscle cells in vitro. Pooled, multiplex single-cell RNA sequencing on 50 cell lines revealed multiple anatomically distinct MN subtypes of cervical and brachial, limb-innervating MNs in reproducible quantities. We conclude that combining small molecule patterning with Ngn2 can facilitate the high-yield, robust and reproducible production of multiple disease-relevant MN subtypes, which is fundamental in the path to propel forward our knowledge of motoneuron biology and its disruption in disease.

Local probes of the electronic ground state are essential for understanding hydrogen bonding in aqueous environments. When tuned to the dissociative core-excited state at the O1 s pre-edge of water, resonant inelastic X-ray scattering back to the electronic ground state exhibits a long vibrational progression due to ultrafast nuclear dynamics. We show how the coherent evolution of the OH bonds around the core-excited oxygen provides access to high vibrational levels in liquid water. The OH bonds stretch into the long-range part of the potential energy curve, which makes the X-ray probe more sensitive than infra-red spectroscopy to the local environment. We exploit this property to effectively probe hydrogen bond strength via the distribution of intramolecular OH potentials derived from measurements. In contrast, the dynamical splitting in the spectral feature of the lowest valence-excited state arises from the short-range part of the OH potential curve and is rather insensitive to hydrogen bonding. Understanding how nuclear motions affect vibrational motions in molecular liquids remains challenging in modern condensed matter physics. Here the authors study the vibrational quantum effects in liquid water and show the sensitivity on the coherent evolution of OH bonds in core-excited states.

Diversity within or between tumours and metastases (known as intra-patient tumour heterogeneity) that develops during disease progression is a serious hurdle for therapy 1 – 3 . Metastasis is the fatal hallmark of cancer and the mechanisms of colonization, the most complex step in the metastatic cascade 4 , remain poorly defined. A clearer understanding of the cellular and molecular processes that underlie both intra-patient tumour heterogeneity and metastasis is crucial for the success of personalized cancer therapy. Here, using transcriptional profiling of tumours and matched metastases in patient-derived xenograft models in mice, we show cancer-site-specific phenotypes and increased glucocorticoid receptor activity in distant metastases. The glucocorticoid receptor mediates the effects of stress hormones, and of synthetic derivatives of these hormones that are used widely in the clinic as anti-inflammatory and immunosuppressive agents. We show that the increase in stress hormones during breast cancer progression results in the activation of the glucocorticoid receptor at distant metastatic sites, increased colonization and reduced survival. Our transcriptomics, proteomics and phospho-proteomics studies implicate the glucocorticoid receptor in the activation of multiple processes in metastasis and in the increased expression of kinase ROR1, both of which correlate with reduced survival. The ablation of ROR1 reduced metastatic outgrowth and prolonged survival in preclinical models. Our results indicate that the activation of the glucocorticoid receptor increases heterogeneity and metastasis, which suggests that caution is needed when using glucocorticoids to treat patients with breast cancer who have developed cancer-related complications. In patient-derived xenograft models of breast cancer in mice, an increase in stress hormones during progression or treatment with their synthetic derivatives activates the glucocorticoid receptor, and results in increased metastatic colonization and reduced survival.

The dynamics of fragmentation and vibration of molecular systems with a large number of coupled degrees of freedom are key aspects for understanding chemical reactivity and properties. Here we present a resonant inelastic X-ray scattering (RIXS) study to show how it is possible to break down such a complex multidimensional problem into elementary components. Local multimode nuclear wave packets created by X-ray excitation to different core-excited potential energy surfaces (PESs) will act as spatial gates to selectively probe the particular ground-state vibrational modes and, hence, the PES along these modes. We demonstrate this principle by combining ultra-high resolution RIXS measurements for gas-phase water with state-of-the-art simulations. Investigating dynamics of polyatomic molecules is difficult as their potential energy surfaces are multidimensional due to coupled degrees of freedom. Here the authors demonstrate a spatial selective gating technique to probe the different vibrational modes of water upon core-level excitation with X-rays.

Pancreatic ductal adenocarcinoma has a dismal prognosis. A comprehensive analysis of single-cell multi-omic data from matched tumour-infiltrated CD45+ cells and peripheral blood in 12 patients, and two published datasets, reveals a complex immune infiltrate. Patients have either a myeloid-enriched or adaptive-enriched tumour microenvironment. Adaptive immune cell-enriched is intrinsically linked with highly distinct B and T cell clonal selection, diversification, and differentiation. Using TCR data, we see the largest clonal expansions in CD8 effector memory, senescent cells, and highly activated regulatory T cells which are induced within the tumour from naïve cells. We identify pathways that potentially lead to a suppressive microenvironment, including investigational targets TIGIT/PVR and SIRPA/CD47. Analysis of patients from the APACT clinical trial shows that myeloid enrichment had a shorter overall survival compared to those with adaptive cell enrichment. Strategies for rationale therapeutic development in this disease include boosting of B cell responses, targeting immunosuppressive macrophages, and specific Treg cell depletion approaches. Pancreatic ductal adenocarcinoma (PDAC) has a poor prognosis involving evasion of immune control. Here, the authors perform a comprehensive analysis of single-cell multi-omic data revealing either a myeloid-enriched or adaptive-enriched tumour microenvironment, linked to distinct B and T cell clonal selection and differentiation, distinct overall survival, and potential therapeutic approaches.

The timely release of SARS-CoV-2 first genomic sequences allowed the identification of the etiologic agent and development of diagnostic protocols. Genomic sequencing was a crucial step in generating data for driving laboratory response and detections of SARS-CoV-2 since the start of the COVID-19 pandemic. Because of all the progression and achievements that timely release of genetic sequence data represents in the public health response, the Pan American Health Organization (PAHO) in collaboration with countries’ public health laboratories, started implementation of a network for strengthening the Latin America and Caribbean (LAC) region on timely generation of SARS-CoV-2 genomic data. Here we describe the implementation of the COVID-19 Genomic Surveillance Regional Network in the Americas region during the beginning of the pandemic. The establishment of this network has strengthened laboratory response capacity at the country level, as well as facilitated timely release of SARS-CoV-2 genomic information to be used to complement the multiple response strategies for COVID-19 pandemic mitigation. As genomic epidemiology is useful for guiding public health decisions on outbreak and response, we also analysed the first SARS-CoV-2 genomic sequence data from countries of the Latin America and Caribbean Region.