Explore the vast range of titles available.

In vertebrate hearts, the ventricular trabecular myocardium develops as a sponge-like network of cardiomyocytes that is critical for contraction and conduction, ventricular septation, papillary muscle formation and wall thickening through the process of compaction 1 . Defective trabeculation leads to embryonic lethality 2 – 4 or non-compaction cardiomyopathy (NCC) 5 . There are divergent views on when and how trabeculation is initiated in different species. In zebrafish, trabecular cardiomyocytes extrude from compact myocardium 6 , whereas in chicks, chamber wall thickening occurs before overt trabeculation 7 . In mice, the onset of trabeculation has not been described, but is proposed to begin at embryonic day 9.0, when cardiomyocytes form radially oriented ribs 2 . Endocardium–myocardium communication is essential for trabeculation, and numerous signalling pathways have been identified, including Notch 2 , 8 and Neuregulin (NRG) 4 . Late disruption of the Notch pathway causes NCC 5 . Whereas it has been shown that mutations in the extracellular matrix (ECM) genes Has2 and Vcan prevent the formation of trabeculae in mice 9 , 10 and the matrix metalloprotease ADAMTS1 promotes trabecular termination 3 , the pathways involved in ECM dynamics and the molecular regulation of trabeculation during its early phases remain unexplored. Here we present a model of trabeculation in mice that integrates dynamic endocardial and myocardial cell behaviours and ECM remodelling, and reveal new epistatic relationships between the involved signalling pathways. NOTCH1 signalling promotes ECM degradation during the formation of endocardial projections that are critical for individualization of trabecular units, whereas NRG1 promotes myocardial ECM synthesis, which is necessary for trabecular rearrangement and growth. These systems interconnect through NRG1 control of Vegfa , but act antagonistically to establish trabecular architecture. These insights enabled the prediction of persistent ECM and cardiomyocyte growth in a mouse NCC model, providing new insights into the pathophysiology of congenital heart disease. A new model of cardiac trabeculation in mice is presented in which NOTCH1 and NRG1 have opposing roles in extracellular matrix degradation and synthesis that are essential for defining trabecular architecture.

Advances in single-cell RNA sequencing (scRNA-seq) have allowed for comprehensive analysis of the immune system. In this Review, we briefly describe the available scRNA-seq technologies together with their corresponding strengths and weaknesses. We discuss in depth how scRNA-seq can be used to deconvolve immune system heterogeneity by identifying novel distinct immune cell subsets in health and disease, characterizing stochastic heterogeneity within a cell population and building developmental 'trajectories' for immune cells. Finally, we discuss future directions of the field and present integrated approaches to complement molecular information from a single cell with studies of the environment, epigenetic state and cell lineage.

Mycorrhizal fungi are mutualists that play crucial roles in nutrient acquisition in terrestrial ecosystems. Mycorrhizal symbioses arose repeatedly across multiple lineages of Mucoromycotina, Ascomycota, and Basidiomycota. Considerable variation exists in the capacity of mycorrhizal fungi to acquire carbon from soil organic matter. Here, we present a combined analysis of 135 fungal genomes from 73 saprotrophic, endophytic and pathogenic species, and 62 mycorrhizal species, including 29 new mycorrhizal genomes. This study samples ecologically dominant fungal guilds for which there were previously no symbiotic genomes available, including ectomycorrhizal Russulales, Thelephorales and Cantharellales. Our analyses show that transitions from saprotrophy to symbiosis involve (1) widespread losses of degrading enzymes acting on lignin and cellulose, (2) co-option of genes present in saprotrophic ancestors to fulfill new symbiotic functions, (3) diversification of novel, lineage-specific symbiosis-induced genes, (4) proliferation of transposable elements and (5) divergent genetic innovations underlying the convergent origins of the ectomycorrhizal guild.

The heterogeneity of the tumour immune microenvironment (TIME), organized by various immune and stromal cells, is a major contributing factor of tumour metastasis, relapse and drug resistance 1 – 3 , but how different TIME subtypes are connected to the clinical relevance in liver cancer remains unclear. Here we performed single-cell RNA-sequencing (scRNA-seq) analysis of 189 samples collected from 124 patients and 8 mice with liver cancer. With more than 1 million cells analysed, we stratified patients into five TIME subtypes, including immune activation, immune suppression mediated by myeloid or stromal cells, immune exclusion and immune residence phenotypes. Different TIME subtypes were spatially organized and associated with chemokine networks and genomic features. Notably, tumour-associated neutrophil (TAN) populations enriched in the myeloid-cell-enriched subtype were associated with an unfavourable prognosis. Through in vitro induction of TANs and ex vivo analyses of patient TANs, we showed that CCL4 + TANs can recruit macrophages and that PD-L1 + TANs can suppress T cell cytotoxicity. Furthermore, scRNA-seq analysis of mouse neutrophil subsets revealed that they are largely conserved with those of humans. In vivo neutrophil depletion in mouse models attenuated tumour progression, confirming the pro-tumour phenotypes of TANs. With this detailed cellular heterogeneity landscape of liver cancer, our study illustrates diverse TIME subtypes, highlights immunosuppressive functions of TANs and sheds light on potential immunotherapies targeting TANs. Tumour-associated neutrophil populations enriched in the myeloid-cell-enriched tumour immune microenvironment subtype are associated with unfavourable prognosis in humans and mice with liver cancer.

Single-cell and spatial technologies that profile gene expression across a whole tissue are revolutionizing the resolution of molecular states in clinical samples. Current commercially available technologies provide whole transcriptome single-cell, whole transcriptome spatial, or targeted in situ gene expression analysis. Here, we combine these technologies to explore tissue heterogeneity in large, FFPE human breast cancer sections. This integrative approach allowed us to explore molecular differences that exist between distinct tumor regions and to identify biomarkers involved in the progression towards invasive carcinoma. Further, we study cell neighborhoods and identify rare boundary cells that sit at the critical myoepithelial border confining the spread of malignant cells. Here, we demonstrate that each technology alone provides information about molecular signatures relevant to understanding cancer heterogeneity; however, it is the integration of these technologies that leads to deeper insights, ushering in discoveries that will progress oncology research and the development of diagnostics and therapeutics. The integration of single-cell and spatial data can provide a more comprehensive picture of the network of cells within the tumour microenvironment. Here the authors use a combination of single-cell and spatial technologies including 10x Xenium to characterise serial formalin-fixed, paraffin-embedded human breast cancer sections.

Therapeutic mRNAs and vaccines are being developed for a broad range of human diseases, including COVID-19. However, their optimization is hindered by mRNA instability and inefficient protein expression. Here, we describe design principles that overcome these barriers. We develop an RNA sequencing-based platform called PERSIST-seq to systematically delineate in-cell mRNA stability, ribosome load, as well as in-solution stability of a library of diverse mRNAs. We find that, surprisingly, in-cell stability is a greater driver of protein output than high ribosome load. We further introduce a method called In-line-seq, applied to thousands of diverse RNAs, that reveals sequence and structure-based rules for mitigating hydrolytic degradation. Our findings show that highly structured “superfolder” mRNAs can be designed to improve both stability and expression with further enhancement through pseudouridine nucleoside modification. Together, our study demonstrates simultaneous improvement of mRNA stability and protein expression and provides a computational-experimental platform for the enhancement of mRNA medicines. The authors develop an RNA sequencing-based platform, PERSIST-seq, to simultaneously delineate in-cell mRNA stability, ribosome load, and in-solution stability of a diverse mRNA library to derive design principles for improved mRNA therapeutics.

Chromosome structure in mammals is thought to regulate transcription by modulating three-dimensional interactions between enhancers and promoters, notably through CTCF-mediated loops and topologically associating domains (TADs) 1 – 4 . However, how chromosome interactions are actually translated into transcriptional outputs remains unclear. Here, to address this question, we use an assay to position an enhancer at large numbers of densely spaced chromosomal locations relative to a fixed promoter, and measure promoter output and interactions within a genomic region with minimal regulatory and structural complexity. A quantitative analysis of hundreds of cell lines reveals that the transcriptional effect of an enhancer depends on its contact probabilities with the promoter through a nonlinear relationship. Mathematical modelling suggests that nonlinearity might arise from transient enhancer–promoter interactions being translated into slower promoter bursting dynamics in individual cells, therefore uncoupling the temporal dynamics of interactions from those of transcription. This uncovers a potential mechanism of how distal enhancers act from large genomic distances, and of how topologically associating domain boundaries block distal enhancers. Finally, we show that enhancer strength also determines absolute transcription levels as well as the sensitivity of a promoter to CTCF-mediated transcriptional insulation. Our measurements establish general principles for the context-dependent role of chromosome structure in long-range transcriptional regulation. The transcriptional effect of an enhancer depends on its contact probabilities with the promoter through a nonlinear relationship, and enhancer strength determines absolute transcription levels as well as the sensitivity of a promoter to CTCF-mediated transcriptional insulation.

N 6 -methyladenosine (m 6 A), the most prevalent internal RNA modification on mammalian messenger RNAs, regulates the fates and functions of modified transcripts through m 6 A-specific binding proteins 1 – 5 . In the nervous system, m 6 A is abundant and modulates various neural functions 6 – 11 . Whereas m 6 A marks groups of mRNAs for coordinated degradation in various physiological processes 12 – 15 , the relevance of m 6 A for mRNA translation in vivo remains largely unknown. Here we show that, through its binding protein YTHDF1, m 6 A promotes protein translation of target transcripts in response to neuronal stimuli in the adult mouse hippocampus, thereby facilitating learning and memory. Mice with genetic deletion of Ythdf1 show learning and memory defects as well as impaired hippocampal synaptic transmission and long-term potentiation. Re-expression of YTHDF1 in the hippocampus of adult Ythdf1 -knockout mice rescues the behavioural and synaptic defects, whereas hippocampus-specific acute knockdown of Ythdf1 or Mettl3 , which encodes the catalytic component of the m 6 A methyltransferase complex, recapitulates the hippocampal deficiency. Transcriptome-wide mapping of YTHDF1-binding sites and m 6 A sites on hippocampal mRNAs identified key neuronal genes. Nascent protein labelling and tether reporter assays in hippocampal neurons showed that YTHDF1 enhances protein synthesis in a neuronal-stimulus-dependent manner. In summary, YTHDF1 facilitates translation of m 6 A-methylated neuronal mRNAs in response to neuronal stimulation, and this process contributes to learning and memory. Neuronal stimulation induces protein translation of m 6 A-methylated neuronal mRNAs facilitated by YTHDF1, and this process contributes to learning and memory.

The growing understanding of RNA functions and their crucial roles in diseases promotes the application of various RNAs to selectively function on hitherto “undruggable” proteins, transcripts and genes, thus potentially broadening the therapeutic targets. Several RNA-based medications have been approved for clinical use, while others are still under investigation or preclinical trials. Various techniques have been explored to promote RNA intracellular trafficking and metabolic stability, despite significant challenges in developing RNA-based therapeutics. In this review, the mechanisms of action, challenges, solutions, and clinical application of RNA-based therapeutics have been comprehensively summarized.

Some circular RNAs (circRNAs) were found to be translated through IRES-driven mechanism, however the scope and functions of circRNA translation are unclear because endogenous IRESs are rare. To determine the prevalence and mechanism of circRNA translation, we develop a cell-based system to screen random sequences and identify 97 overrepresented hexamers that drive cap-independent circRNA translation. These IRES-like short elements are significantly enriched in endogenous circRNAs and sufficient to drive circRNA translation. We further identify multiple trans -acting factors that bind these IRES-like elements to initiate translation. Using mass-spectrometry data, hundreds of circRNA-coded peptides are identified, most of which have low abundance due to rapid degradation. As judged by mass-spectrometry, 50% of translatable endogenous circRNAs undergo rolling circle translation, several of which are experimentally validated. Consistently, mutations of the IRES-like element in one circRNA reduce its translation. Collectively, our findings suggest a pervasive translation of circRNAs, providing profound implications in translation control. Unbiased screen of random sequences identified many short IRES-like elements to drive circular RNA translation and hundreds of rolling circle translation events, suggesting a pervasive cap-independent translation in human transcriptome.