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Fanconi anaemia (FA), a model syndrome of genome instability, is caused by a deficiency in DNA interstrand crosslink repair resulting in chromosome breakage1–3. The FA repair pathway protects against endogenous and exogenous carcinogenic aldehydes4–7. Individuals with FA are hundreds to thousands fold more likely to develop head and neck (HNSCC), oesophageal and anogenital squamous cell carcinomas8 (SCCs). Molecular studies of SCCs from individuals with FA (FA SCCs) are limited, and it is unclear how FA SCCs relate to sporadic HNSCCs primarily driven by tobacco and alcohol exposure or infection with human papillomavirus9 (HPV). Here, by sequencing genomes and exomes of FA SCCs, we demonstrate that the primary genomic signature of FA repair deficiency is the presence of high numbers of structural variants. Structural variants are enriched for small deletions, unbalanced translocations and fold-back inversions, and are often connected, thereby forming complex rearrangements. They arise in the context of TP53 loss, but not in the context of HPV infection, and lead to somatic copy-number alterations of HNSCC driver genes. We further show that FA pathway deficiency may lead to epithelial-to-mesenchymal transition and enhanced keratinocyte-intrinsic inflammatory signalling, which would contribute to the aggressive nature of FA SCCs. We propose that the genomic instability in sporadic HPV-negative HNSCC may arise as a result of the FA repair pathway being overwhelmed by DNA interstrand crosslink damage caused by alcohol and tobacco-derived aldehydes, making FA SCC a powerful model to study tumorigenesis resulting from DNA-crosslinking damage.

A novel severe acute respiratory syndrome (SARS)-like coronavirus (SARS-CoV-2) recently emerged and is rapidly spreading in humans, causing COVID-191,2. A key to tackling this pandemic is to understand the receptor recognition mechanism of the virus, which regulates its infectivity, pathogenesis and host range. SARS-CoV-2 and SARS-CoV recognize the same receptor—angiotensin-converting enzyme 2 (ACE2)—in humans3,4. Here we determined the crystal structure of the receptor-binding domain (RBD) of the spike protein of SARS-CoV-2 (engineered to facilitate crystallization) in complex with ACE2. In comparison with the SARS-CoV RBD, an ACE2-binding ridge in SARS-CoV-2 RBD has a more compact conformation; moreover, several residue changes in the SARS-CoV-2 RBD stabilize two virus-binding hotspots at the RBD–ACE2 interface. These structural features of SARS-CoV-2 RBD increase its ACE2-binding affinity. Additionally, we show that RaTG13, a bat coronavirus that is closely related to SARS-CoV-2, also uses human ACE2 as its receptor. The differences among SARS-CoV-2, SARS-CoV and RaTG13 in ACE2 recognition shed light on the potential animal-to-human transmission of SARS-CoV-2. This study provides guidance for intervention strategies that target receptor recognition by SARS-CoV-2. The crystal structure of the receptor-binding domain of the SARS-CoV-2 spike in complex with human ACE2, compared with the receptor-binding domain of SARS-CoV, sheds light on the structural features that increase its binding affinity to ACE2.

Catalysts are widely used to increase reaction rates. They function by stabilizing the transition state of the reaction at their active site, where the atomic arrangement ensures favourable interactions 1 . However, mechanistic understanding is often limited when catalysts possess multiple active sites-such as sites associated with either the step edges or the close-packed terraces of inorganic nanoparticles2-4-with distinct activities that cannot be measured simultaneously. An example is the oxidation of carbon monoxide over platinum surfaces, one of the oldest and best studied heterogeneous reactions. In 1824, this reaction was recognized to be crucial for the function of the Davy safety lamp, and today it is used to optimize combustion, hydrogen production and fuel-cell operation5,6. The carbon dioxide products are formed in a bimodal kinetic energy distribution7-13; however, despite extensive study 5 , it remains unclear whether this reflects the involvement of more than one reaction mechanism occurring at multiple active sites12,13. Here we show that the reaction rates at different active sites can be measured simultaneously, using molecular beams to controllably introduce reactants and slice ion imaging14,15 to map the velocity vectors of the product molecules, which reflect the symmetry and the orientation of the active site 16 . We use this velocity-resolved kinetics approach to map the oxidation rates of carbon monoxide at step edges and terrace sites on platinum surfaces, and find that the reaction proceeds through two distinct channels11-13: it is dominated at low temperatures by the more active step sites, and at high temperatures by the more abundant terrace sites. We expect our approach to be applicable to a wide range of heterogeneous reactions and to provide improved mechanistic understanding of the contribution of different active sites, which should be useful in the design of improved catalysts.

The transcriptome is the readout of the genome. Identifying common features in it across distant species can reveal fundamental principles. To this end, the ENCODE and modENCODE consortia have generated large amounts of matched RNA-sequencing data for human, worm and fly. Uniform processing and comprehensive annotation of these data allow comparison across metazoan phyla, extending beyond earlier within-phylum transcriptome comparisons and revealing ancient, conserved features1, 2, 3, 4, 5, 6. Specifically, we discover co-expression modules shared across animals, many of which are enriched in developmental genes. Moreover, we use expression patterns to align the stages in worm and fly development and find a novel pairing between worm embryo and fly pupae, in addition to the embryo-to-embryo and larvae-to-larvae pairings. Furthermore, we find that the extent of non-canonical, non-coding transcription is similar in each organism, per base pair. Finally, we find in all three organisms that the gene-expression levels, both coding and non-coding, can be quantitatively predicted from chromatin features at the promoter using a ‘universal model’ based on a single set of organism-independent parameters. In particular, this work was funded by a contract from the National Human Genome Research Institute modENCODE Project, contract U01 HG004271 and U54 HG006944, to S.E.C. (principal investigator) and P.C., T.R.G., R.A.H. and B.R.G. (co-principal investigators) with additional support from R01 GM076655 (S.E.C.) both under Department of Energy contract no. DE-AC02-05CH11231, and U54 HG007005 to B.R.G. J.B.B.’s work was supported by NHGRI K99 HG006698 and DOE DE-AC02-05CH11231. Work in P.J.B.’s group was supported by the modENCODE DAC sub award 5710003102, 1U01HG007031-01 and the ENCODE DAC 5U01HG004695-04. Work in M.B.G.’s group was supported by NIH grants HG007000 and HG007355. Work in Bloomington was supported in part by the Indiana METACyt Initiative of Indiana University, funded by an award from the Lilly Endowment, Inc. Work in E.C.L.’s group was supported by U01-HG004261 and RC2-HG005639. P.J.P. acknowledges support from the National Institutes of Health (grant no. U01HG004258). We thank the HAVANA team for providing annotation of the human reference genome, whose work is supported by National Institutes of Health (grant no. 5U54HG004555), the Wellcome Trust (grant no. WT098051). R.G. acknowledges support from the Spanish Ministry of Education (grant BIO2011-26205). We also acknowledge use of the Yale University Biomedical High Performance Computing Center. R.W.'s lab was supported by grant no. U01 HG 004263.

At central excitatory synapses, N-methyl-D-aspartate (NMDA) receptors, which have a high affinity for glutamate, produce a slowly rising synaptic current in response to a single transmitter pulse and an additional current after a second, closely timed stimulus. Here we show, by examining the kinetics of transmitter binding and channel gating in single-channel currents from recombinant NR1/NR2A receptors, that the synaptic response to trains of impulses is determined by the molecular reaction mechanism of the receptor. The rate constants estimated for the activation reaction predict that, after binding neurotransmitter, receptors hesitate for approximately 4 ms in a closed high-affinity conformation before they either proceed towards opening or release neurotransmitter, with about equal probabilities. Because only about half of the initially fully occupied receptors become active, repetitive stimulation elicits currents with distinct waveforms depending on pulse frequency. This high-affinity/low-efficiency activation mechanism might serve as a link between stimulation frequency and the directionality of the ensuing synaptic plasticity.

Parkinson's disease is a widespread condition caused by the loss of midbrain neurons that synthesize the neurotransmitter dopamine. Cells derived from the fetal midbrain can modify the course of the disease, but they are an inadequate source of dopamine-synthesizing neurons because their ability to generate these neurons is unstable. In contrast, embryonic stem (ES) cells proliferate extensively and can generate dopamine neurons. If ES cells are to become the basis for cell therapies, we must develop methods of enriching for the cell of interest and demonstrate that these cells show functions that will assist in treating the disease. Here we show that a highly enriched population of midbrain neural stem cells can be derived from mouse ES cells. The dopamine neurons generated by these stem cells show electrophysiological and behavioural properties expected of neurons from the midbrain. Our results encourage the use of ES cells in cell-replacement therapy for Parkinson's disease.

Allosteric transitions allow fast regulation of protein function in living systems. Even though the end points of such conformational changes are known for many proteins, the characteristics of the paths connecting these states remain largely unexplored. Rate-equilibrium linear free-energy relationships (LFERs) provide information about such pathways by relating changes in the free energy of the transition state to those of the ground states upon systematic perturbation of the system. Here we present an LFER analysis of the gating reaction pathway of the muscle acetylcholine receptor. We studied the closed open conformational change at the single-molecule level following perturbation by series of single-site mutations, agonists and membrane voltages. This method provided a snapshot of several regions of the receptor at the transition state in terms of their approximate positions along the reaction coordinate, on a scale from 0 (closed-like) to 1 (open-like). The resulting map reveals a spatial gradient of positional values, which suggests that the conformational change proceeds in a wave-like manner, with the low-to-high affinity change at the transmitter-binding sites preceding the complete opening of the pore.

Studies and models of trace-gas flux in the Arctic consider temperature and moisture to be the dominant controls over land–atmosphere exchange1,2, with little attention having been paid to the effects of different substrates. Likewise, current Arctic vegetation maps for models of vegetation change recognize one or two tundra types3,4 and do not portray the extensive regions with different soils within the Arctic. Here we show that rapid changes to ecosystem processes (such as photosynthesis and respiration) that are related to changes in climate and land usage will be superimposed upon and modulated by differences in substrate pH. A sharp soil pH boundary along the northern front of the Arctic Foothills in Alaska separates non-acidic (pH > 6.5) ecosystems to the north from predominantly acidic (pH < 5.5) ecosystems to the south. Moist non-acidic tundra has greater heat flux, deeper summer thaw (active layer), is less of a carbon sink, and is a smaller source of methane than moist acidic tundra.

Gaining insight into the nature and dynamics of the transition state is the essence of mechanistic investigations of chemical reactions1, yet the fleeting configuration when existing chemical bonds dissociate while new ones form is extremely difficult to examine directly2. Adiabatic potential-energy surfaces—usually derived using quantum chemical methods3 that assume mutually independent nuclear and electronic motion4—quantify the fundamental forces between atoms involved in reaction and thus provide accurate descriptions of a reacting system as it moves through its transition state5,6. This approach, widely tested for gas-phase reactions7, is now also commonly applied to chemical reactions at metal surfaces8. There is, however, some evidence calling into question the correctness of this theoretical approach for surface reactions: electronic excitation upon highly exothermic chemisorption has been observed9, and indirect evidence suggests that large-amplitude vibrations of reactant molecules can excite electrons at metal surfaces10,11. Here we report the detection of ‘hot’ electrons leaving a metal surface as vibrationally highly excited NO molecules collide with it. Electron emission only occurs once the vibrational energy exceeds the surface work function, and is at least 10,000 times more efficient than the emissions seen in similar systems where large-amplitude vibrations were not involved12,13,14,15,16,17,18. These observations unambiguously demonstrate the direct conversion of vibrational to electronic excitation, thus questioning one of the basic assumptions currently used in theoretical approaches to describing bond-dissociation at metal surfaces.

THE outer layer of the blastocyst, or trophectoderm, is the first cell lineage to differentiate in the mouse embryo1,2, but little is known about the genetic control of its development. Lineage-specific transcription factors may be important in lineage specification, and the product of the Mash-2 gene3,4 fulfils the criteria for such a factor. Mash-2 is a mammalian member of the achaete-scute family5–7 which encodes basic-helix–loop–helix transcription factors8 and is strongly expressed in the extraembryonic tropho-blast lineage. Mash-2 transcripts are found in the female germ line and in the embryo throughout preimplantation development, but are highly expressed later only in the ectoplacental cone, the chor-ion and their derivatives in the placenta. Mash-2 transcripts are not found in primary and secondary giant cells, yolk sac or allantois at any post-implantation stage, and are present only transiently and at low levels in the embryo during gastrulation. To analyse the role of Mash-2 in development, we have used gene targeting to generate mice having no Mash-2 function. We report here that Mash-2-/- embryos die from placental failure at 10 days post-coitum. In mutant placentas, spongiotrophoblast cells and their precursors are absent and chorionic ectoderm is reduced. We have rescued this placental mutant phenotype by constructing chimaeras with tetraploid wild-type embryos which contribute almost exclusively to extraembryonic tissues9,10. Mash-2-/- embryos developed normally and adult Mash-2-/- mice were viable, demonstrating that Mash-2 has no major role in the embryo itself. Mash-2 is the first transcription factor shown to play a critical part in the development of the mammalian trophoblast lineage.

Tuberous sclerosis complex and fragile X syndrome are genetic diseases characterized by intellectual disability and autism. Because both syndromes are caused by mutations in genes that regulate protein synthesis in neurons, it has been hypothesized that excessive protein synthesis is one core pathophysiological mechanism of intellectual disability and autism. Using electrophysiological and biochemical assays of neuronal protein synthesis in the hippocampus of Tsc2(+/-) and Fmr1(-/y) mice, here we show that synaptic dysfunction caused by these mutations actually falls at opposite ends of a physiological spectrum. Synaptic, biochemical and cognitive defects in these mutants are corrected by treatments that modulate metabotropic glutamate receptor 5 in opposite directions, and deficits in the mutants disappear when the mice are bred to carry both mutations. Thus, normal synaptic plasticity and cognition occur within an optimal range of metabotropic glutamate-receptor-mediated protein synthesis, and deviations in either direction can lead to shared behavioural impairments.

Muscle contraction is triggered by the opening of acetylcholine receptors at the vertebrate nerve-muscle synapse. The M2 helix of this allosteric membrane protein lines the channel, and contains a 'gate' that regulates the flow of ions through the pore. We used single-molecule kinetic analysis to probe the transition state of the gating conformational change and estimate the relative timing of M2 motions in the alpha-subunit of the murine acetylcholine receptor. This analysis produces a 'Phi-value' for a given residue that reflects its open-like versus closed-like character at the transition state. Here we show that most of the residues throughout the length of M2 have a Phi-value of approximately 0.64 but that some near the middle have lower Phi-values of 0.52 or 0.31, suggesting that alphaM2 moves in three discrete steps. The core of the channel serves both as a gate that regulates ion flow and as a hub that directs the propagation of the gating isomerization through the membrane domain of the acetylcholine receptor.

A noteworthy feature of the high-temperature superconductors is the unconventional symmetry of the superconducting order parameter. Several experiments1,2,3 have established that the order parameter has a four-fold symmetry under rotation of the lattice (the order parameter of conventional superconductors is, in contrast, isotropic). An intriguing and much debated possibility is that, in certain cases, an additional imaginary component might be present, having an isotropic s-wave4,5,6 or dxy symmetry7,8,9,10. A consequence of a complex order parameter of the form + idxy is that it would break both reflection (parity, P) symmetry and time-reversal (T) symmetry, a clear signature of which would be the spontaneous appearance of a macroscopic magnetization at the superconducting transition temperature. Broken T symmetry has been reported5,11, but searches for the effects of combined P and T symmetry breaking have so far yielded null results12,13,14,15. Here we report the observation of a weak (∼10-5 gauss) magnetic field that appears spontaneously at the superconducting transition temperature of epitaxial thin films of YBa2Cu3O7-δ. The magnetic signal originates near the edges of the samples. One interpretation for this observation is that the order parameter carries an intrinsic angular momentum, related to the breaking of P and T symmetries, but other possibilities cannot yet be excluded.

The simple aggregation of pluripotent ES cells with morulae-stage embryos can result in viable chimaeras at a similar frequency to that of blastocyst injection. The savings in time and equipment inherent in the aggregation techniques, however, make this approach well worth considering.

An experiment is described in which tracer quantities of Cs/sup 137/ were used in a relatively large-scale field investigation. A forest ecosystem tagged with Cs/sup 137/ was used: to determine cesium cycles and rates of cycling in a mesic forest ecosystem; to measure cesium transfer between components of an ecosystem and apply these results to a theoretical model of element circulation; and to derive information useful for predicting the fate of problem radionuclides in those ecosystems potentially having a large capacity for storage in biological materials. The site chosen was a sinkhole habitat forested by Liriodendron tulipifera. Methods used for dose determination, inoculation of trees, avoiding undesirable contamination, and sampling of the forest are described. Seasonal variations in isotope distribution are discussed. Preliminary results emphasize the importance of rain as an agent in radiocesium transfer. Approximately 20 percent of the total litter burden was derived from rain leaching. Seven percent of the original radiocesium input to Liriodendron was transferred to the forest floor during the first growing season. (H.M.G.)

The biological hazards of radiation from radioactive fall-out are discussed. Authoritative statements on hazards from man-made radiation are reviewed. Dosage estimates and genetic consequences are considered. (C.H.)

FANCONI'S anaemia (FA) is one of the genetic syndromes in which chromosomal instability has been associated with an increased predisposition to cancer1. An increased risk of malignancy has also been reported for presumptive carriers of the FA gene2,3, although no evidence of chromosome abnormalities has been found in cultured lymphocytes or fibroblasts from the FA heterozygote. The nature of the genetic defect in FA is still unclear, although there is evidence for defective DNA repair4–6.Lymphocytes from FA homozygotes have an increased incidence of chromosome aberrations after exposure to ionising radiation7 or alkylating agents8,9. We reported recently that baseline breakage rates for FA cell strains varied from 0.20 to 0.36 breaks per cell, and that after carcinogen treatment of the type described below, these rates were increased three to fivefold10. The same carcinogen treatment had no clastogenic effect on normal, xero-derma pigmentosum and trisomy 18 fibroblasts. There is at present no way to distinguish FA heterozygous individuals from normal. The only distinction that has been reported has been an increased susceptibility to in vitro transformation by the onco-genic virus SV40 in both homozygous and heterozygous FA fibroblasts11, but similar results have been obtained with cell strains from patients with several other syndromes. As the ability to identify carriers of the FA gene would be of great benefit because of their increased cancer risk, we report here the results of experiments in which we were able to distinguish FA heterozygous cell strains from normals, after exposing the cells to the difunctional alkylating agent diepoxybutane (DEB), an active mutagen and carcinogen 12–15.

The source of stem cells within the early mammalian embryo has not been identified. The yolk sac, once thought to provide the early stem cells in birds, is now seen more as a primitive effector organ capable of allorecognition, natural cytotoxic reactions and elaboration of cytokines. In lower vertebrates the source appears to be within a region delineated by the anterior limbs, foregut and mesonephros. That region defines the boundaries of the developing omentum, a fold in the peritoneum. As the omentum is known to develop lymphoid cells postnatally we have now examined the omental rudiment for the presence of lymphoid cell precursors. Our experiments provide evidence that the presumptive omentum of the 13-day mouse embryo is capable of histiotypic differentiation into a reticular organ containing a significant number of Thy 1+ lymphocytes, which suggests that the omentum may represent a new primary lymphoid organ in the mouse.

WHEN tumour cells are inoculated into syngeneic mice their establishment and growth is subject to regulation by the host animal. We have shown this most effectively for the inoculation of a broad range of tumours into anterior compared with posterior portions of the trunk1. For example, when mammary tumour cells were injected intradermally into thoracic compared with lumbar regions of adult histocompatible mice, tumour growth anteriorly became detectable by more rapid palpation, and tumour growth was three to four times as rapid anteriorly as posteriorly. This difference between anterior and posterior injection sites also became manifest after subcutaneous inoculation; it was apparent after inoculation of a wide variety of tumour types (mastocytoma, melanoma, lymphoma, sarcoma, mammary adenocarcinoma); it was independent of the sex of the host animal; it was found in a variety of mouse strains; and it did not seem to be related to the immunocompetence of the host or to the immunogenicity of the tumour1,7. We have now determined that there are antero-posterior and dorso-ventral differences in the growth of tumour cells inoculated intradermally into adult histocompatible mice. Although we have no explanation for our results we suggest that the observed differences may reflect the existence of morphogenetic gradients reminiscent of those invoked to explain patterns of differentiation during ontogeny.

THREE decades of intensive study of nuclear phenomena have not brought a detailed understanding of the forces between the nucleons which comprise the atomic nucleus. Even if the forces were known, mathematical techniques are at present inadequate to solve the Schrodinger equation for nuclei containing more than a few nucleons.

INDIVIDUALS with certain genetic syndromes associated with chromosome damage show markedly increased incidence of cancer1. In one such syndrome, Fanconi's anaemia (FA), chromosomal breakage and rearrangement has been found in lymphocytes and fibroblasts2,3 years before the development of malignant tumours. There is some evidence that cells from FA patients are especially sensitive to oncogenic agents. FA fibroblasts are abnormally susceptible to SV40 transformation4, and lymphocytes from patients show increased chromosome aberrations after exposure to ionising radiation5, or to alkylating agents6,7. In these reports, however, evaluation was based on cells which entered metaphase during brief and possibly toxic exposures to chemicals. We report here experiments in which FA fibroblasts were exposed to a direct-acting mutagen or carcinogen for a period of 6 d, ensuring chronic exposure of the cells during one or more cell cycles, until increased cell density inhibited further cell division. After subculture we could then assay chromosome damage in cells capable of entering a new cycle of cell division after removal from the chemical. We found that viable FA fibroblasts showed increases in chromosome aberrations after exposure to the mutagen or carcinogen at concentrations that had no effect in other cell strains tested. Since fibroblasts can be maintained in serial passage, their use in this protocol makes it possible to examine residual and possibly lasting effects of the treatment.

Phosphorylation of ion channels has been suggested as one molecular mechanism responsible for learning-produced long-term changes in neuronal excitability. Persistent training-produced changes in two distinct K+ currents (IA (ref. 2), IK-Ca (refs 3,4)) and a voltage-dependent calcium current (ICa; refs 3,4) have previously been shown to occur in type B photoreceptors of Hermissenda, as a result of associative learning. But the identity of the phosphorylation pathway(s) responsible for these changes has not as yet been determined. Injections of cyclic AMP-dependent protein kinase reduce a K+ current (IK) in B cells which is different from those changed by training, but fails to reduce IA and IK-Ca. Phosphorylase b kinase (an exogenous calcium/calmodulin-dependent kinase) reduces IA, but whether IK-Ca and ICa are changed in the manner of associative training is not yet known. Another protein kinase present in high concentrations in both mammalian brain and molluscan nervous systems is protein kinase C, which is both calcium- and phospholipid-sensitive. We now present evidence that activation of protein kinase C by the tumour promoter phorbol ester (PDB) and intracellular injection of the enzyme induce conductance changes similar to those caused by associative training in Hermissenda B cells (that is a reduction of IA and IK-Ca, and enhancement of ICa). These results represent the first direct demonstration that protein kinase C affects membrane K+ ion conductance mechanisms.