Treatment of congenital Langerhans cell histiocytosis with cobimetinib.

We report a case of congenital multisystem Langerhans cell histiocytosis with cutaneous and hematopoietic involvement. After the failure of first-line (vinblastine and prednisolone) and second-line (vincristine and cytarabine) therapies, treatment with cobimetinib, a mitogen-activated protein kinase (MEK) inhibitor, led to the remission of disease and a sustained response after 11 months of ongoing treatment. Protein kinase inhibitors targeting BRAF or MEK could represent a promising future therapeutic option, also in children with LCH.

The ATM Ser49Cys Variant Effects ATM Function as a Regulator of Oncogene-Induced Senescence.

An apical component of the cell cycle checkpoint and DNA damage repair response is the ataxia-telangiectasia mutated (ATM) Ser/Thr protein kinase. A variant of ATM, Ser49Cys (rs1800054; minor allele frequency = 0.011), has been associated with an elevated risk of melanoma development; however, the functional consequence of this variant is not defined. ATM-dependent signalling in response to DNA damage has been assessed in a panel of patient-derived lymphoblastoid lines and primary human melanocytic cell strains heterozygous for the ATM Ser49Cys variant allele. The ATM Ser49Cys allele appears functional for acute p53-dependent signalling in response to DNA damage. Expression of the variant allele did reduce the efficacy of oncogene expression in inducing senescence. These findings demonstrate that the ATM 146C>G Ser49Cys allele has little discernible effect on the acute response to DNA damage but has reduced function observed in the chronic response to oncogene over-expression. Analysis of melanoma, naevus and skin colour genomics and GWAS analyses have demonstrated no association of this variant with any of these outcomes. The modest loss of function detected suggest that the variant may act as a modifier of other variants of ATM/p53-dependent signalling.

Germline ERBB3 mutation in familial non-small-cell lung carcinoma: expanding ErbB's role in oncogenesis.

Lung cancer is the commonest cause of cancer deaths worldwide. Although strongly associated with smoking, predisposition to lung cancer is also heritable, with multiple common risk variants identified. Rarely, dominantly inherited non-small-cell lung cancer (NSCLC) has been reported due to somatic mutations in EGFR/ErbB1 and ERBB2. Germline exome sequencing was performed in a multi-generation family with autosomal dominant NSCLC, including an affected child. Tumour samples were also sequenced. Full-length wild-type (wtErbB3) and mutant ERBB3 (mutErbB3) constructs were transfected into HeLa cells. Protein expression, stability, and subcellular localization were assessed, and cellular proliferation, pAkt/Akt and pERK levels determined. A novel germline variant in ERBB3 (c.1946 T > G: p.Iso649Arg), coding for receptor tyrosine-protein kinase erbB-3 (ErbB3), was identified, with appropriate segregation. There was no loss-of-heterozygosity in tumour samples. Both wtErbB3 and mutErbB3 were stably expressed. MutErbB3-transfected cells demonstrated an increased ratio of the 80 kDa form (which enhances proliferation) compared with the full-length (180 kDa) form. MutErbB3 and wtErbB3 had similar punctate cytoplasmic localization pre- and post-epidermal growth factor stimulation; however, epidermal growth factor receptor (EGFR) levels decreased faster post-stimulation in mutErbB3-transfected cells, suggesting more rapid processing of the mutErbB3/EGFR heterodimer. Cellular proliferation was increased in mutErbB3-transfected cells compared with wtErbB3 transfection. MutErbB3-transfected cells also showed decreased pAkt/tAkt ratios and increased pERK/tERK 30 min post-stimulation compared with wtErbB3 transfection, demonstrating altered signalling pathway activation. Cumulatively, these results support this mutation as tumorogenic. This is the first reported family with a germline ERBB3 mutation causing heritable NSCLC, furthering understanding of the ErbB family pathway in oncogenesis.

Proteome profiling of salivary small extracellular vesicles in glioblastoma patients.

BACKGROUND: Extracellular vesicles (EVs) play a critical role in intercellular communication under physiological and pathological conditions, including cancer. EVs cargo reflects their cell of origin, suggesting their utility as biomarkers. EVs are detected in several biofluids, and their ability to cross the blood-brain barrier has highlighted their potential as prognostic and diagnostic biomarkers in gliomas, including glioblastoma (GBM). Studies have demonstrated the potential clinical utility of plasma-derived EVs in glioma. However, little is known about the clinical utility of saliva-derived EVs in GBM. METHODS: Small EVs were isolated from whole mouth saliva of GBM patients pre- and postoperatively. Isolation was performed using differential centrifugation and/or ultracentrifugation. EVs were characterized by concentration, size, morphology, and EVs cell-surface protein markers. Protein cargo in EVs was profiled using mass spectrometry. RESULTS: There were no statistically significant differences in size and concentration of EVs derived from pre- and post GBM patients' saliva samples. A higher number of proteins were detected in preoperative samples compared to postoperative samples. The authors found four highly abundant proteins (aldolase A, 14-3-3 protein ε, enoyl CoA hydratase 1, and transmembrane protease serine 11B) in preoperative saliva samples from GBM patients with poor outcomes. Functional enrichment analysis of pre- and postoperative saliva samples showed significant enrichment of several pathways, including those related to the immune system, cell cycle and programmed cell death. CONCLUSIONS: This study, for the first time, demonstrates the feasibility of isolating and characterizing small EVs from pre- and postoperative saliva samples from GBM patients. Preliminary findings encourage further large cohort validation studies on salivary small EVs to evaluate prognosis in GBM.

Head and neck cancer patient-derived tumouroid cultures: opportunities and challenges.

Head and neck cancers (HNC) are the seventh most prevalent cancer type globally. Despite their common categorisation, HNCs are a heterogeneous group of malignancies arising in various anatomical sites within the head and neck region. These cancers exhibit different clinical and biological manifestations, and this heterogeneity also contributes to the high rates of treatment failure and mortality. To evaluate patients who will respond to a particular treatment, there is a need to develop in vitro model systems that replicate in vivo tumour status. Among the methods developed, patient-derived cancer organoids, also known as tumouroids, recapitulate in vivo tumour characteristics including tumour architecture. Tumouroids have been used for general disease modelling and genetic instability studies in pan-cancer research. However, a limited number of studies have thus far been conducted using tumouroid-based drug screening. Studies have concluded that tumouroids can play an essential role in bringing precision medicine for highly heterogenous cancer types such as HNC.

Genetically encoded Runx3 and CD4+ intestinal epithelial lymphocyte deficiencies link SKG mouse and human predisposition to spondyloarthropathy.

Disturbances in immune regulation, intestinal dysbiosis and inflammation characterize ankylosing spondylitis (AS), which is associated with RUNX3 loss-of-function variants. ZAP70W163C mutant (SKG) mice have reduced ZAP70 signaling, spondyloarthritis and ileitis. In small intestine, Foxp3+ regulatory T cells (Treg) and CD4+CD8αα+TCRαß+ intraepithelial lymphocytes (CD4-IEL) control inflammation. TGF-ß and retinoic acid (RA)-producing dendritic cells and MHC-class II+ intestinal epithelial cells (IEC) are required for Treg and CD4-IEL differentiation from CD4+ conventional or Treg precursors, with upregulation of Runx3 and suppression of ThPOK. We show in SKG mouse ileum, that ZAP70W163C or ZAP70 inhibition prevented CD4-IEL but not Treg differentiation, dysregulating Runx3 and ThPOK. TGF-ß/RA-mediated CD4-IEL development, T-cell IFN-γ production, MHC class-II+ IEC, tissue-resident memory T-cell and Runx3-regulated genes were reduced. In AS intestine, CD4-IEL were decreased, while in AS blood CD4+CD8+ T cells were reduced and Treg increased. Thus, genetically-encoded TCR signaling dysfunction links intestinal T-cell immunodeficiency in mouse and human spondyloarthropathy.

Circulating tumour DNA alterations: emerging biomarker in head and neck squamous cell carcinoma.

Head and Neck cancers (HNC) are a heterogeneous group of upper aero-digestive tract cancer and account for 931,922 new cases and 467,125 deaths worldwide. About 90% of these cancers are of squamous cell origin (HNSCC). HNSCC is associated with excessive tobacco and alcohol consumption and infection with oncogenic viruses. Genotyping tumour tissue to guide clinical decision-making is becoming common practice in modern oncology, but in the management of patients with HNSCC, cytopathology or histopathology of tumour tissue remains the mainstream for diagnosis and treatment planning. Due to tumour heterogeneity and the lack of access to tumour due to its anatomical location, alternative methods to evaluate tumour activities are urgently needed. Liquid biopsy approaches can overcome issues such as tumour heterogeneity, which is associated with the analysis of small tissue biopsy. In addition, liquid biopsy offers repeat biopsy sampling, even for patients with tumours with access limitations. Liquid biopsy refers to biomarkers found in body fluids, traditionally blood, that can be sampled to provide clinically valuable information on both the patient and their underlying malignancy. To date, the majority of liquid biopsy research has focused on blood-based biomarkers, such as circulating tumour DNA (ctDNA), circulating tumour cells (CTCs), and circulating microRNA. In this review, we will focus on ctDNA as a biomarker in HNSCC because of its robustness, its presence in many body fluids, adaptability to existing clinical laboratory-based technology platforms, and ease of collection and transportation. We will discuss mechanisms of ctDNA release into circulation, technological advances in the analysis of ctDNA, ctDNA as a biomarker in HNSCC management, and some of the challenges associated with translating ctDNA into clinical and future perspectives. ctDNA provides a minimally invasive method for HNSCC prognosis and disease surveillance and will pave the way in the future for personalized medicine, thereby significantly improving outcomes and reducing healthcare costs.

Pulled Diversification Rates, Lineages-Through-Time Plots, and Modern Macroevolutionary Modeling.

Estimating time-dependent rates of speciation and extinction from dated phylogenetic trees of extant species (timetrees), and determining how and why they vary, is key to understanding how ecological and evolutionary processes shape biodiversity. Due to an increasing availability of phylogenetic trees, a growing number of process-based methods relying on the birth-death model have been developed in the last decade to address a variety of questions in macroevolution. However, this methodological progress has regularly been criticized such that one may wonder how reliable the estimations of speciation and extinction rates are. In particular, using lineages-through-time (LTT) plots, a recent study has shown that there are an infinite number of equally likely diversification scenarios that can generate any timetree. This has led to questioning whether or not diversification rates should be estimated at all. Here, we summarize, clarify, and highlight technical considerations on recent findings regarding the capacity of models to disentangle diversification histories. Using simulations, we illustrate the characteristics of newly proposed "pulled rates" and their utility. We recognize that the recent findings are a step forward in understanding the behavior of macroevolutionary modeling, but they in no way suggest we should abandon diversification modeling altogether. On the contrary, the study of macroevolution using phylogenetic trees has never been more exciting and promising than today. We still face important limitations in regard to data availability and methods, but by acknowledging them we can better target our joint efforts as a scientific community. [Birth-death models; extinction; phylogenetics; speciation.].

A sequential diversification with Miocene extinction and Pliocene speciation linked to mountain uplift explains the diversity of the African rain forest clade Monodoreae (Annonaceae).

BACKGROUND AND AIMS: Throughout the Cenozoic, Africa underwent several climatic and geological changes impacting the evolution of tropical rain forests (TRF). African TRF are thought to have extended from East to West in a 'pan-African' TRF, followed by several events of fragmentation during drier climate periods. During the Miocene, climate cooling and mountain uplift led to the aridification of tropical Africa and open habitats expanded at the expense of TRF, which likely experienced local extinctions. However, in plants, these drivers were previously inferred using limited taxonomic and molecular data. Here, we tested the impact of climate and geological changes on diversification within the diverse clade Monodoreae (Annonaceae) composed of 90 tree species restricted to African TRF. METHODS: We reconstructed a near complete phylogenetic tree, based on 32 nuclear genes, and dated using relaxed clocks and fossil calibrations in a Bayesian framework. We inferred the biogeographic history and the diversification dynamics of the clade using multiple birth-death models. KEY RESULTS: Monodoreae originated in East African TRF ca. 25 million years ago (Ma) and expanded toward Central Africa during the Miocene. We inferred range contractions during the middle Miocene and document important connections between East and West African TRF after 15-13 Ma. Our results indicated a sudden extinction event during the late Miocene, followed by an increase in speciation rates. Birth-death models suggested that African elevation change (orogeny) is positively linked to speciation in this clade. CONCLUSION: East Africa is inferred as an important source of Monodoreae species, and possibly for African plant diversity in general. Our results support a "sequential scenario of diversification" where increased aridification triggered extinction of TRF species in Monodoreae. This was quickly followed by rain forests fragmentation, subsequently enhancing lagged speciation resulting from vicariance and improved climate conditions. In contrast to previous ideas, the uplift of East Africa is shown to have played a positive role in Monodoreae diversification.

Transmission Dynamics of Large Coronavirus Disease Outbreak in Homeless Shelter, Chicago, Illinois, USA, 2020.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has the potential for rapid transmission in congregate settings. We describe the multidisciplinary response to an outbreak of coronavirus disease (COVID-19) in a large homeless shelter in Chicago, Illinois, USA. The response to the outbreak included 4 rounds of mass PCR testing of all staff and residents and subsequent isolation of persons who tested positive for SARS-CoV-2. We further describe the dynamics of the shelter outbreak by fitting a modified susceptible-exposed-infectious-recovered compartmental model incorporating the widespread SARS-CoV-2 testing and isolation measures implemented in this shelter. Our model demonstrates that rapid transmission of COVID-19 in the shelter occurred before the outbreak was detected; rates of transmission declined after widespread testing and isolation measures were put in place. Overall, we demonstrate the feasibility of mass PCR testing and isolation in congregate settings and suggest the necessity of prompt response to suspected COVID-19 outbreaks in homeless shelters.

Heterozygous loss of WBP11 function causes multiple congenital defects in humans and mice.

The genetic causes of multiple congenital anomalies are incompletely understood. Here, we report novel heterozygous predicted loss-of-function (LoF) and predicted damaging missense variants in the WW domain binding protein 11 (WBP11) gene in seven unrelated families with a variety of overlapping congenital malformations, including cardiac, vertebral, tracheo-esophageal, renal and limb defects. WBP11 encodes a component of the spliceosome with the ability to activate pre-messenger RNA splicing. We generated a Wbp11 null allele in mouse using CRISPR-Cas9 targeting. Wbp11 homozygous null embryos die prior to E8.5, indicating that Wbp11 is essential for development. Fewer Wbp11 heterozygous null mice are found than expected due to embryonic and postnatal death. Importantly, Wbp11 heterozygous null mice are small and exhibit defects in axial skeleton, kidneys and esophagus, similar to the affected individuals, supporting the role of WBP11 haploinsufficiency in the development of congenital malformations in humans. LoF WBP11 variants should be considered as a possible cause of VACTERL association as well as isolated Klippel-Feil syndrome, renal agenesis or esophageal atresia.

Analysis of human leukocyte antigen associations in human papillomavirus-positive and -negative head and neck cancer: Comparison with cervical cancer.

BACKGROUND: Although the majority of human papillomavirus (HPV) infections are cleared by the immune system, a small percentage of them progress to develop HPV-driven cancers. Cervical cancer studies highlight that HPV persistence and cancer risk are associated with genetic factors, especially at the human leukocyte antigen (HLA) genes. This study was conducted to investigate such associations in head and neck cancer (HNC). METHODS: In all, 192 patients with HNC and 384 controls were genotyped with the Infinium Global Screening Array (Illumina, Inc). HLA variants were imputed with SNP2HLA, and an association analysis was performed by logistic regression. RESULTS: HPV-positive HNCs were significantly associated with single-nucleotide polymorphisms (SNPs) at DRB1_32660090 (P = 1.728 × 10-6 ) and DRB1_32660116 (P = 1.728 × 10-6 ) and with the amino acid variant DRB1_11_32660115 (P = 1.728 × 10-6 ). None of these associations were observed in the HPV-negative cohort, and this suggested their specificity to convey risk for HPV-associated HNCs. In general, associations observed for HPV-negative HNC were relatively weak, and variants in the HLA-DPA1 region were the strongest among them (P = 4.531 × 10-4 ). Several lead signals reported by previous HNC genome-wide association studies, including SNPs rs3135001 (P = .012), rs1049055 (P = .012), and rs34518860 (P = .029) and allele HLA-DQB1*06 (P = .009), were replicated in the current study. However, these associations were limited to the HPV-positive HNC group. Several cervical cancer-associated HLA variants, including SNPs rs9272143 (P = .002) and rs9271858 (P = .002) and alleles HLA-B-1501 (P = .009) and HLA-B-15 (P = .015), were also exclusively associated with HPV-positive HNC. CONCLUSIONS: HPV-positive HNC risk is associated with distinct HLA variants, and some of them are shared by both cervical cancer and HPV-positive HNC. Human papillomavirus (HPV)-positive head and neck cancer (HNC) risk is associated with distinct human leukocyte antigen variants, and some of them are shared by both cervical cancer and HPV-positive HNC. LAY SUMMARY: Cervical cancer studies highlight that human papillomavirus (HPV)-driven cancer risk is linked with human leukocyte antigen (HLA) polymorphism. Hence, the current study was designed to investigate the HLA associations in HPV-positive and HPV-negative head and neck cancer (HNC) and compare these associations with cervical cancer. Several lead signals reported by previous HNC and cervical genome-wide association studies were replicated in the current study. However, these associations were limited to the HPV-positive HNC group, and this suggests that HPV-positive HNC risk is associated with distinct HLA variants, and some of them are shared by both cervical cancer and HPV-positive HNC.

Genome-wide association study in Turkish and Iranian populations identify rare familial Mediterranean fever gene (MEFV) polymorphisms associated with ankylosing spondylitis.

Ankylosing spondylitis (AS) is a highly heritable immune-mediated arthritis common in Turkish and Iranian populations. Familial Mediterranean Fever (FMF) is an autosomal recessive autoinflammatory disease most common in people of Mediterranean origin. MEFV, an FMF-associated gene, is also a candidate gene for AS. We aimed to identify AS susceptibility loci and also examine the association between MEFV and AS in Turkish and Iranian cohorts. We performed genome-wide association studies in 1001 Turkish AS patients and 1011 Turkish controls, and 479 Iranian AS patients and 830 Iranian controls. Serum IL-1ß, IL-17 and IL-23 cytokine levels were quantified in Turkish samples. An association of major effect was observed with a novel rare coding variant in MEFV in the Turkish cohort (rs61752717, M694V, OR = 5.3, P = 7.63×10(-12)), Iranian cohort (OR = 2.9, P = 0.042), and combined dataset (OR = 5.1, P = 1.65×10(-13)). 99.6% of Turkish AS cases, and 96% of those carrying MEFV rs61752717 variants, did not have FMF. In Turkish subjects, the association of rs61752717 was particularly strong in HLA-B27-negative cases (OR = 7.8, P = 8.93×10(-15)), but also positive in HLA-B27-positive cases (OR = 4.3, P = 7.69×10(-8)). Serum IL-1ß, IL-17 and IL-23 levels were higher in AS cases than controls. Among AS cases, serum IL-1ß and IL-23 levels were increased in MEFV 694V carriers compared with non-carriers. Our data suggest that FMF and AS have overlapping aetiopathogenic mechanisms. Functionally important MEFV mutations, such as M694V, lead to dysregulated inflammasome function and excessive IL-1ß function. As IL-1 inhibition is effective in FMF, AS cases carrying FMF-associated MEFV variants may benefit from such therapy.

Polygenic Risk Scores have high diagnostic capacity in ankylosing spondylitis.

OBJECTIVE: We sought to test the hypothesis that Polygenic Risk Scores (PRSs) have strong capacity to discriminate cases of ankylosing spondylitis (AS) from healthy controls and individuals in the community with chronic back pain. METHODS: PRSs were developed and validated in individuals of European and East Asian ethnicity, using data from genome-wide association studies in 15 585 AS cases and 20 452 controls. The discriminatory values of PRSs in these populations were compared with other widely used diagnostic tests, including C-reactive protein (CRP), HLA-B27 and sacroiliac MRI. RESULTS: In people of European descent, PRS had high discriminatory capacity with area under the curve (AUC) in receiver operator characteristic analysis of 0.924. This was significantly better than for HLA-B27 testing alone (AUC=0.869), MRI (AUC=0.885) or C-reactive protein (AUC=0.700). PRS developed and validated in individuals of East Asian descent performed similarly (AUC=0.948). Assuming a prior probability of AS of 10% such as in patients with chronic back pain under 45 years of age, compared with HLA-B27 testing alone, PRS provides higher positive values for 35% of patients and negative predictive values for 67.5% of patients. For PRS, in people of European descent, the maximum positive predictive value was 78.2% and negative predictive value was 100%, whereas for HLA-B27, these values were 51.9% and 97.9%, respectively. CONCLUSIONS: PRS have higher discriminatory capacity for AS than CRP, sacroiliac MRI or HLA-B27 status alone. For optimal performance, PRS should be developed for use in the specific ethnic groups to which they are to be applied.

Whole-genome sequencing identifies EN1 as a determinant of bone density and fracture.

The extent to which low-frequency (minor allele frequency (MAF) between 1-5%) and rare (MAF ≤ 1%) variants contribute to complex traits and disease in the general population is mainly unknown. Bone mineral density (BMD) is highly heritable, a major predictor of osteoporotic fractures, and has been previously associated with common genetic variants, as well as rare, population-specific, coding variants. Here we identify novel non-coding genetic variants with large effects on BMD (ntotal = 53,236) and fracture (ntotal = 508,253) in individuals of European ancestry from the general population. Associations for BMD were derived from whole-genome sequencing (n = 2,882 from UK10K (ref. 10); a population-based genome sequencing consortium), whole-exome sequencing (n = 3,549), deep imputation of genotyped samples using a combined UK10K/1000 Genomes reference panel (n = 26,534), and de novo replication genotyping (n = 20,271). We identified a low-frequency non-coding variant near a novel locus, EN1, with an effect size fourfold larger than the mean of previously reported common variants for lumbar spine BMD (rs11692564(T), MAF = 1.6%, replication effect size = +0.20 s.d., Pmeta = 2 × 10(-14)), which was also associated with a decreased risk of fracture (odds ratio = 0.85; P = 2 × 10(-11); ncases = 98,742 and ncontrols = 409,511). Using an En1(cre/flox) mouse model, we observed that conditional loss of En1 results in low bone mass, probably as a consequence of high bone turnover. We also identified a novel low-frequency non-coding variant with large effects on BMD near WNT16 (rs148771817(T), MAF = 1.2%, replication effect size = +0.41 s.d., Pmeta = 1 × 10(-11)). In general, there was an excess of association signals arising from deleterious coding and conserved non-coding variants. These findings provide evidence that low-frequency non-coding variants have large effects on BMD and fracture, thereby providing rationale for whole-genome sequencing and improved imputation reference panels to study the genetic architecture of complex traits and disease in the general population.

Granulomatous dermatitis following measles, mumps, and rubella vaccination.

Granulomatous dermatitis following the administration of various vaccines has previously been reported. However, cases of cutaneous granulomatosis following the measles, mumps, and rubella (MMR) vaccine have not yet been reported. We report the case of a 3-year-old boy with a granuloma annulare-like reaction following MMR vaccination.

Correction: Defining the genetic susceptibility to cervical neoplasia-A genome-wide association study.

[This corrects the article DOI: 10.1371/journal.pgen.1006866.].

Near real-time detection of low-frequency baleen whale calls from an autonomous surface vehicle: Implementation, evaluation, and remaining challenges.

Mitigation of threats posed to marine mammals by human activities can be greatly improved with a better understanding of animal occurrence in real time. Recent advancements have enabled low-power passive acoustic systems to be integrated into long-endurance autonomous platforms for persistent near real-time monitoring of marine mammals via the sounds they produce. Here, the integration of a passive acoustic instrument capable of real-time detection and classification of low-frequency (LF) tonal sounds with a Liquid Robotics wave glider is reported. The goal of the integration was to enable monitoring of LF calls produced by baleen whales over periods of several months. Mechanical noises produced by the platform were significantly reduced by lubricating moving parts with polytetrafluoroethylene, incorporating rubber and springs to decelerate moving parts and shock mounting hydrophones. Flow noise was reduced with the development of a 21-element hydrophone array. Surface noise produced by breaking waves was not mitigated despite experimentation with baffles. Compared to a well-characterized moored passive acoustic monitoring buoy, the system greatly underestimated the occurrence of sei, fin, and North Atlantic right whales during a 37-d deployment, and therefore is not suitable in its current configuration for use in scientific or management applications for these species at this time.

NAD Deficiency, Congenital Malformations, and Niacin Supplementation.

BACKGROUND: Congenital malformations can be manifested as combinations of phenotypes that co-occur more often than expected by chance. In many such cases, it has proved difficult to identify a genetic cause. We sought the genetic cause of cardiac, vertebral, and renal defects, among others, in unrelated patients. METHODS: We used genomic sequencing to identify potentially pathogenic gene variants in families in which a person had multiple congenital malformations. We tested the function of the variant by using assays of in vitro enzyme activity and by quantifying metabolites in patient plasma. We engineered mouse models with similar variants using the CRISPR (clustered regularly interspaced short palindromic repeats)-Cas9 system. RESULTS: Variants were identified in two genes that encode enzymes of the kynurenine pathway, 3-hydroxyanthranilic acid 3,4-dioxygenase (HAAO) and kynureninase (KYNU). Three patients carried homozygous variants predicting loss-of-function changes in the HAAO or KYNU proteins (HAAO p.D162*, HAAO p.W186*, or KYNU p.V57Efs*21). Another patient carried heterozygous KYNU variants (p.Y156* and p.F349Kfs*4). The mutant enzymes had greatly reduced activity in vitro. Nicotinamide adenine dinucleotide (NAD) is synthesized de novo from tryptophan through the kynurenine pathway. The patients had reduced levels of circulating NAD. Defects similar to those in the patients developed in the embryos of Haao-null or Kynu-null mice owing to NAD deficiency. In null mice, the prevention of NAD deficiency during gestation averted defects. CONCLUSIONS: Disruption of NAD synthesis caused a deficiency of NAD and congenital malformations in humans and mice. Niacin supplementation during gestation prevented the malformations in mice. (Funded by the National Health and Medical Research Council of Australia and others.).

Cradles and museums of generic plant diversity across tropical Africa.

Determining where species diversify (cradles) and persist (museums) over evolutionary time is fundamental to understanding the distribution of biodiversity and for conservation prioritization. Here, we identify cradles and museums of angiosperm generic diversity across tropical Africa, one of the most biodiverse regions on Earth. Regions containing nonrandom concentrations of young (neo-) and old (paleo-) endemic taxa were identified using distribution data of 1719 genera combined with a newly generated time-calibrated mega-phylogenetic tree. We then compared the identified regions with the current network of African protected areas (PAs). At the generic level, phylogenetic diversity and endemism are mainly concentrated in the biogeographically complex region of Eastern Africa. We show that mountainous areas are centres of both neo- and paleo-endemism. By contrast, the Guineo-Congolian lowland rain forest region is characterized by widespread and old lineages. We found that the overlap between centres of phylogenetic endemism and PAs is high (> 85%). We show the vital role played by mountains acting simultaneously as cradles and museums of tropical African plant biodiversity. By contrast, lowland rainforests act mainly as museums for generic diversity. Our study shows that incorporating large-scale taxonomically verified distribution datasets and mega-phylogenies lead to an improved understanding of tropical plant biodiversity evolution.