Expression Atlas update: insights from sequencing data at both bulk and single cell level.

Expression Atlas (www.ebi.ac.uk/gxa) and its newest counterpart the Single Cell Expression Atlas (www.ebi.ac.uk/gxa/sc) are EMBL-EBI's knowledgebases for gene and protein expression and localisation in bulk and at single cell level. These resources aim to allow users to investigate their expression in normal tissue (baseline) or in response to perturbations such as disease or changes to genotype (differential) across multiple species. Users are invited to search for genes or metadata terms across species or biological conditions in a standardised consistent interface. Alongside these data, new features in Single Cell Expression Atlas allow users to query metadata through our new cell type wheel search. At the experiment level data can be explored through two types of dimensionality reduction plots, t-distributed Stochastic Neighbor Embedding (tSNE) and Uniform Manifold Approximation and Projection (UMAP), overlaid with either clustering or metadata information to assist users' understanding. Data are also visualised as marker gene heatmaps identifying genes that help confer cluster identity. For some data, additional visualisations are available as interactive cell level anatomograms and cell type gene expression heatmaps.

Enhancing European capabilities for application of multi-omics studies in biology and biomedicine space research.

Following on from the NASA twins' study, there has been a tremendous interest in the use of omics techniques in spaceflight. Individual space agencies, NASA's GeneLab, JAXA's ibSLS, and the ESA-funded Space Omics Topical Team and the International Standards for Space Omics Processing (ISSOP) groups have established several initiatives to support this growth. Here, we present recommendations from the Space Omics Topical Team to promote standard application of space omics in Europe. We focus on four main themes: i) continued participation in and coordination with international omics endeavors, ii) strengthening of the European space omics infrastructure including workforce and facilities, iii) capitalizing on the emerging opportunities in the commercial space sector, and iv) capitalizing on the emerging opportunities in human subjects research.

Mannose metabolism inhibition sensitizes acute myeloid leukaemia cells to therapy by driving ferroptotic cell death.

Resistance to standard and novel therapies remains the main obstacle to cure in acute myeloid leukaemia (AML) and is often driven by metabolic adaptations which are therapeutically actionable. Here we identify inhibition of mannose-6-phosphate isomerase (MPI), the first enzyme in the mannose metabolism pathway, as a sensitizer to both cytarabine and FLT3 inhibitors across multiple AML models. Mechanistically, we identify a connection between mannose metabolism and fatty acid metabolism, that is mediated via preferential activation of the ATF6 arm of the unfolded protein response (UPR). This in turn leads to cellular accumulation of polyunsaturated fatty acids, lipid peroxidation and ferroptotic cell death in AML cells. Our findings provide further support to the role of rewired metabolism in AML therapy resistance, unveil a connection between two apparently independent metabolic pathways and support further efforts to achieve eradication of therapy-resistant AML cells by sensitizing them to ferroptotic cell death.

Matrix and analysis metadata standards (MAMS) to facilitate harmonization and reproducibility of single-cell data.

A large number of genomic and imaging datasets are being produced by consortia that seek to characterize healthy and disease tissues at single-cell resolution. While much effort has been devoted to capturing information related to biospecimen information and experimental procedures, the metadata standards that describe data matrices and the analysis workflows that produced them are relatively lacking. Detailed metadata schema related to data analysis are needed to facilitate sharing and interoperability across groups and to promote data provenance for reproducibility. To address this need, we developed the Matrix and Analysis Metadata Standards (MAMS) to serve as a resource for data coordinating centers and tool developers. We first curated several simple and complex "use cases" to characterize the types of feature-observation matrices (FOMs), annotations, and analysis metadata produced in different workflows. Based on these use cases, metadata fields were defined to describe the data contained within each matrix including those related to processing, modality, and subsets. Suggested terms were created for the majority of fields to aid in harmonization of metadata terms across groups. Additional provenance metadata fields were also defined to describe the software and workflows that produced each FOM. Finally, we developed a simple list-like schema that can be used to store MAMS information and implemented in multiple formats. Overall, MAMS can be used as a guide to harmonize analysis-related metadata which will ultimately facilitate integration of datasets across tools and consortia. MAMS specifications, use cases, and examples can be found at https://github.com/single-cell-mams/mams/.

Epigenetic and transcriptional regulations prime cell fate before division during human pluripotent stem cell differentiation.

Stem cells undergo cellular division during their differentiation to produce daughter cells with a new cellular identity. However, the epigenetic events and molecular mechanisms occurring between consecutive cell divisions have been insufficiently studied due to technical limitations. Here, using the FUCCI reporter we developed a cell-cycle synchronised human pluripotent stem cell (hPSC) differentiation system for uncovering epigenome and transcriptome dynamics during the first two divisions leading to definitive endoderm. We observed that transcription of key differentiation markers occurs before cell division, while chromatin accessibility analyses revealed the early inhibition of alternative cell fates. We found that Activator protein-1 members controlled by p38/MAPK signalling are necessary for inducing endoderm while blocking cell fate shifting toward mesoderm, and that enhancers are rapidly established and decommissioned between different cell divisions. Our study has practical biomedical utility for producing hPSC-derived patient-specific cell types since p38/MAPK induction increased the differentiation efficiency of insulin-producing pancreatic beta-cells.

Challenges and considerations for single-cell and spatially resolved transcriptomics sample collection during spaceflight.

Single-cell RNA sequencing (scRNA-seq) and spatially resolved transcriptomics (SRT) have experienced rapid development in recent years. The findings of spaceflight-based scRNA-seq and SRT investigations are likely to improve our understanding of life in space and our comprehension of gene expression in various cell systems and tissue dynamics. However, compared to their Earth-based counterparts, gene expression experiments conducted in spaceflight have not experienced the same pace of development. Out of the hundreds of spaceflight gene expression datasets available, only a few used scRNA-seq and SRT. In this perspective piece, we explore the growing importance of scRNA-seq and SRT in space biology and discuss the challenges and considerations relevant to robust experimental design to enable growth of these methods in the field.

The Maleth program: Malta's first space mission discoveries on the microbiome of diabetic foot ulcers.

The purpose of the Maleth Program, also known as Project Maleth, is Malta's first space program to evaluate human skin tissue microbiome changes in type 2 diabetes mellitus (T2DM) patients afflicted with diabetic foot ulcers (DFU). This was carried out in both ground-based models and spaceflight. The first mission (Maleth I) under this program was carried out to uncover the effects of spaceflight, microgravity and radiation on human skin tissue microbiome samples from six T2DM patients recruited into the study. Each patient human skin tissue sample was split in three, with one section processed immediately for genomic profiling by 16S typing and the rest were processed for longer term ground-control and spaceflight experiments. Ground-control and spaceflight human skin tissue samples were also processed for genomic profiling upon mission re-entry and completion. Maleth I's overall objective was achieved, as human skin tissue samples with their microbiomes travelled to space and back yielding positive results by both standard microbiology techniques and genetic typing using 16S rRNA amplicon sequencing. Preliminary findings of this mission are discussed in light of its innovative approach at DFU microbiome research, and the clinical implications that may emerge from this and other future similar studies.

Machine learning algorithm to characterize antimicrobial resistance associated with the International Space Station surface microbiome.

BACKGROUND: Antimicrobial resistance (AMR) has a detrimental impact on human health on Earth and it is equally concerning in other environments such as space habitat due to microgravity, radiation and confinement, especially for long-distance space travel. The International Space Station (ISS) is ideal for investigating microbial diversity and virulence associated with spaceflight. The shotgun metagenomics data of the ISS generated during the Microbial Tracking-1 (MT-1) project and resulting metagenome-assembled genomes (MAGs) across three flights in eight different locations during 12 months were used in this study. The objective of this study was to identify the AMR genes associated with whole genomes of 226 cultivable strains, 21 shotgun metagenome sequences, and 24 MAGs retrieved from the ISS environmental samples that were treated with propidium monoazide (PMA; viable microbes). RESULTS: We have analyzed the data using a deep learning model, allowing us to go beyond traditional cut-offs based only on high DNA sequence similarity and extending the catalog of AMR genes. Our results in PMA treated samples revealed AMR dominance in the last flight for Kalamiella piersonii, a bacteria related to urinary tract infection in humans. The analysis of 226 pure strains isolated from the MT-1 project revealed hundreds of antibiotic resistance genes from many isolates, including two top-ranking species that corresponded to strains of Enterobacter bugandensis and Bacillus cereus. Computational predictions were experimentally validated by antibiotic resistance profiles in these two species, showing a high degree of concordance. Specifically, disc assay data confirmed the high resistance of these two pathogens to various beta-lactam antibiotics. CONCLUSION: Overall, our computational predictions and validation analyses demonstrate the advantages of machine learning to uncover concealed AMR determinants in metagenomics datasets, expanding the understanding of the ISS environmental microbiomes and their pathogenic potential in humans. Video Abstract.

Mutational synergy during leukemia induction remodels chromatin accessibility, histone modifications and three-dimensional DNA topology to alter gene expression.

Altered transcription is a cardinal feature of acute myeloid leukemia (AML); however, exactly how mutations synergize to remodel the epigenetic landscape and rewire three-dimensional DNA topology is unknown. Here, we apply an integrated genomic approach to a murine allelic series that models the two most common mutations in AML: Flt3-ITD and Npm1c. We then deconvolute the contribution of each mutation to alterations of the epigenetic landscape and genome organization, and infer how mutations synergize in the induction of AML. Our studies demonstrate that Flt3-ITD signals to chromatin to alter the epigenetic environment and synergizes with mutations in Npm1c to alter gene expression and drive leukemia induction. These analyses also allow the identification of long-range cis-regulatory circuits, including a previously unknown superenhancer of Hoxa locus, as well as larger and more detailed gene-regulatory networks, driven by transcription factors including PU.1 and IRF8, whose importance we demonstrate through perturbation of network members.

Modeling HNF1B-associated monogenic diabetes using human iPSCs reveals an early stage impairment of the pancreatic developmental program.

Heterozygous mutations in HNF1B in humans result in a multisystem disorder, including pancreatic hypoplasia and diabetes mellitus. Here we used a well-controlled human induced pluripotent stem cell pancreatic differentiation model to elucidate the molecular mechanisms underlying HNF1B-associated diabetes. Our results show that lack of HNF1B blocks specification of pancreatic fate from the foregut progenitor (FP) stage, but HNF1B haploinsufficiency allows differentiation of multipotent pancreatic progenitor cells (MPCs) and insulin-secreting ß-like cells. We show that HNF1B haploinsufficiency impairs cell proliferation in FPs and MPCs. This could be attributed to impaired induction of key pancreatic developmental genes, including SOX11, ROBO2, and additional TEAD1 target genes whose function is associated with MPC self-renewal. In this work we uncover an exhaustive list of potential HNF1B gene targets during human pancreas organogenesis whose downregulation might underlie HNF1B-associated diabetes onset in humans, thus providing an important resource to understand the pathogenesis of this disease.

MULTIMODAL IMAGING IN A CASE OF CENTRAL RETINAL ARTERY OCCLUSION IN THE PRESENCE OF TYPE 2 MACULAR TELANGIECTASIA.

PURPOSE: To report a case of central retinal artery occlusion in a patient with macular telangiectasia Type 2 using multimodal imaging. METHODS: Observational case report. RESULTS: A 58-year-old woman who presented with acute painless unilateral vision loss was diagnosed with central retinal artery occlusion in her right eye with macular telangiectasia Type 2 findings in both eyes. Fundus examination revealed retinal whitening with unusual cherry-red spot plus small crystalline deposits in the temporal macula. Surprisingly, spectral domain optical coherence tomography of the contralateral eye showed characteristically intraretinal hyporreflective spaces, whereas optical coherence tomography angiography exhibited microvascular abnormalities in the deep capillary plexus. At follow-up, a pseudolamellar macular hole was noticed in the affected eye with no recovery of best-corrected visual acuity. CONCLUSION: This case describes an onset of central artery occlusion in a patient with underlying macular telangiectasia Type 2 and suggests that it had a possible role in the acceleration of its natural course. The utility of multimodal imaging lies on better accuracy in diagnosis as well as prognosis, management, and monitoring of the disease.

Functional Divergence of the Arabidopsis Florigen-Interacting bZIP Transcription Factors FD and FDP.

Flowering of many plant species depends on interactions between basic leucine zipper (bZIP) transcription factors and systemically transported florigen proteins. Members of the genus Arabidopsis contain two of these bZIPs, FD and FDP, which we show have largely complementary expression patterns in shoot apices before and during flowering. CRISPR-Cas9-induced null mutants for FDP flower slightly earlier than wild-type, whereas fd mutants are late flowering. Identical G-box sequences are enriched at FD and FDP binding sites, but only FD binds to genes involved in flowering and only fd alters their transcription. However, both proteins bind to genes involved in responses to the phytohormone abscisic acid (ABA), which controls developmental and stress responses. Many of these genes are differentially expressed in both fd and fdp mutant seedlings, which also show reduced ABA sensitivity. Thus, florigen-interacting bZIPs have distinct functions in flowering dependent on their expression patterns and, at earlier stages in development, play common roles in phytohormone signaling.

Publisher Correction: Single-cell RNA-sequencing of differentiating iPS cells reveals dynamic genetic effects on gene expression.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Single-cell RNA-sequencing of differentiating iPS cells reveals dynamic genetic effects on gene expression.

Recent developments in stem cell biology have enabled the study of cell fate decisions in early human development that are impossible to study in vivo. However, understanding how development varies across individuals and, in particular, the influence of common genetic variants during this process has not been characterised. Here, we exploit human iPS cell lines from 125 donors, a pooled experimental design, and single-cell RNA-sequencing to study population variation of endoderm differentiation. We identify molecular markers that are predictive of differentiation efficiency of individual lines, and utilise heterogeneity in the genetic background across individuals to map hundreds of expression quantitative trait loci that influence expression dynamically during differentiation and across cellular contexts.

GATA6 Cooperates with EOMES/SMAD2/3 to Deploy the Gene Regulatory Network Governing Human Definitive Endoderm and Pancreas Formation.

Heterozygous de novo mutations in GATA6 are the most frequent cause of pancreatic agenesis in humans. In mice, however, a similar phenotype requires the biallelic loss of Gata6 and its paralog Gata4. To elaborate the human-specific requirements for GATA6, we chose to model GATA6 loss in vitro by combining both gene-edited and patient-derived pluripotent stem cells (hPSCs) and directed differentiation toward ß-like cells. We find that GATA6 heterozygous hPSCs show a modest reduction in definitive endoderm (DE) formation, while GATA6-null hPSCs fail to enter the DE lineage. Consistent with these results, genome-wide studies show that GATA6 binds and cooperates with EOMES/SMAD2/3 to regulate the expression of cardinal endoderm genes. The early deficit in DE is accompanied by a significant reduction in PDX1+ pancreatic progenitors and C-PEPTIDE+ ß-like cells. Taken together, our data position GATA6 as a gatekeeper to early human, but not murine, pancreatic ontogeny.

Functional data analysis for computational biology.

SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Efficacy and safety of pop-titrated versus fixed-energy trans-scleral diode laser cyclophotocoagulation for refractory glaucoma.

PURPOSE: To compare the efficacy and safety of pop-titrated versus fixed-energy diode laser trans-scleral cyclophotocoagulation (DLTSC) for refractory glaucoma. METHODS: This is a prospective, interventional, longitudinal, and comparative case-control study. Patients with refractory glaucoma treated with pop-titrated DLTSC were compared to a fixed-energy DLTSC control group. Variables analyzed included: age, gender, diagnosis, pre- and post-treatment intraocular pressure (IOP). Success rate, anti-glaucoma medications reduction, and complications were analyzed at day 90 post-treatment. Primary success criterion consisted of eyes with a postoperative IOP ≤ 22 mmHg or a 30% reduction of pre-treatment IOP and managed with topical anti-glaucoma medications only. RESULTS: A total of 68 eyes from 67 patients were included for analysis: 30 in the pop-titrated group and 38 in the fixed-energy group. Therapeutic success was achieved in 56-72% of the pop-titrated group versus 47-52% in the fixed-energy group considering the 3 different criteria analyzed (p = 0.23-0.4). There was a 22% (from 4.1 to 3.2 drugs) reduction of anti-glaucoma medications in the pop-titrated group, compared to 32% (from 3.5 to 2.4 drugs) in the fixed-energy group (p = 0.42). Five eyes (13.1%) developed hypotony, all of which belonged to the fixed-energy group (p = 0.048). CONCLUSIONS: Pop-titrated DLTSC represents an effective and safe option for the management of refractory glaucoma. We found no statistically significant difference in success rates among both groups. However, there was a significantly higher risk of hypotony in eyes treated with the fixed-energy protocol.

hiPSC hepatocyte model demonstrates the role of unfolded protein response and inflammatory networks in α1-antitrypsin deficiency.

BACKGROUND & AIMS: α1-Antitrypsin deficiency (A1ATD) is an autosomal recessive disorder caused by mutations in the SERPINA1 gene. Individuals with the Z variant (Gly342Lys) retain polymerised protein in the endoplasmic reticulum (ER) of their hepatocytes, predisposing them to liver disease. The concomitant lack of circulating A1AT also causes lung emphysema. Greater insight into the mechanisms that link protein misfolding to liver injury will facilitate the design of novel therapies. METHODS: Human-induced pluripotent stem cell (hiPSC)-derived hepatocytes provide a novel approach to interrogate the molecular mechanisms of A1ATD because of their patient-specific genetic architecture and reflection of human physiology. To that end, we utilised patient-specific hiPSC hepatocyte-like cells (ZZ-HLCs) derived from an A1ATD (ZZ) patient, which faithfully recapitulated key aspects of the disease at the molecular and cellular level. Subsequent functional and "omics" comparisons of these cells with their genetically corrected isogenic-line (RR-HLCs) and primary hepatocytes/human tissue enabled identification of new molecular markers and disease signatures. RESULTS: Our studies showed that abnormal A1AT polymer processing (immobilised ER components, reduced luminal protein mobility and disrupted ER cisternae) occurred heterogeneously within hepatocyte populations and was associated with disrupted mitochondrial structure, presence of the oncogenic protein AKR1B10 and two upregulated molecular clusters centred on members of inflammatory (IL-18 and Caspase-4) and unfolded protein response (Calnexin and Calreticulin) pathways. These results were validated in a second patient-specific hiPSC line. CONCLUSIONS: Our data identified novel pathways that potentially link the expression of Z A1AT polymers to liver disease. These findings could help pave the way towards identification of new therapeutic targets for the treatment of A1ATD. LAY SUMMARY: This study compared the gene expression and protein profiles of healthy liver cells and those affected by the inherited disease α1-antitrypsin deficiency. This approach identified specific factors primarily present in diseased samples which could provide new targets for drug development. This study also demonstrates the interest of using hepatic cells generated from human-induced pluripotent stem cells to model liver disease in vitro for uncovering new mechanisms with clinical relevance.