Biological dinitrogen (N2) fixation is a key metabolic process exclusively performed by prokaryotes, some of which are symbiotic with eukaryotes. Species of the marine haptophyte algae Braarudosphaera bigelowii harbor the N2-fixing endosymbiotic cyanobacteria UCYN-A, which might be evolving organelle-like characteristics. We found that the size ratio between UCYN-A and their hosts is strikingly conserved across sublineages/species, which is consistent with the size relationships of organelles in this symbiosis and other species. Metabolic modeling showed that this size relationship maximizes the coordinated growth rate based on trade-offs between resource acquisition and exchange. Our findings show that the size relationships of N2-fixing endosymbionts and organelles in unicellular eukaryotes are constrained by predictable metabolic underpinnings and that UCYN-A is, in many regards, functioning like a hypothetical N2-fixing organelle (or nitroplast).

The repertoire of modifications to bile acids and related steroidal lipids by host and microbial metabolism remains incompletely characterized. To address this knowledge gap, we created a reusable resource of tandem mass spectrometry (MS/MS) spectra by filtering 1.2 billion publicly available MS/MS spectra for bile-acid-selective ion patterns. Thousands of modifications are distributed throughout animal and human bodies as well as microbial cultures. We employed this MS/MS library to identify polyamine bile amidates, prevalent in carnivores. They are present in humans, and their levels alter with a diet change from a Mediterranean to a typical American diet. This work highlights the existence of many more bile acid modifications than previously recognized and the value of leveraging public large-scale untargeted metabolomics data to discover metabolites. The availability of a modification-centric bile acid MS/MS library will inform future studies investigating bile acid roles in health and disease.

Cell cycle progression relies on coordinated changes in the composition and subcellular localization of the proteome. By applying two distinct convolutional neural networks on images of millions of live yeast cells, we resolved proteome-level dynamics in both concentration and localization during the cell cycle, with resolution of ∼20 subcellular localization classes. We show that a quarter of the proteome displays cell cycle periodicity, with proteins tending to be controlled either at the level of localization or concentration, but not both. Distinct levels of protein regulation are preferentially utilized for different aspects of the cell cycle, with changes in protein concentration being mostly involved in cell cycle control and changes in protein localization in the biophysical implementation of the cell cycle program. We present a resource for exploring global proteome dynamics during the cell cycle, which will aid in understanding a fundamental biological process at a systems level.

Neutrophils, the most abundant and efficient defenders against pathogens, exert opposing functions across cancer types. However, given their short half-life, it remains challenging to explore how neutrophils adopt specific fates in cancer. Here, we generated and integrated single-cell neutrophil transcriptomes from 17 cancer types (225 samples from 143 patients). Neutrophils exhibited extraordinary complexity, with 10 distinct states including inflammation, angiogenesis, and antigen presentation. Notably, the antigen-presenting program was associated with favorable survival in most cancers and could be evoked by leucine metabolism and subsequent histone H3K27ac modification. These neutrophils could further invoke both (neo)antigen-specific and antigen-independent T cell responses. Neutrophil delivery or a leucine diet fine-tuned the immune balance to enhance anti-PD-1 therapy in various murine cancer models. In summary, these data not only indicate the neutrophil divergence across cancers but also suggest therapeutic opportunities such as antigen-presenting neutrophil delivery.

Dorsal root ganglia (DRG) somatosensory neurons detect mechanical, thermal, and chemical stimuli acting on the body. Achieving a holistic view of how different DRG neuron subtypes relay neural signals from the periphery to the CNS has been challenging with existing tools. Here, we develop and curate a mouse genetic toolkit that allows for interrogating the properties and functions of distinct cutaneous targeting DRG neuron subtypes. These tools have enabled a broad morphological analysis, which revealed distinct cutaneous axon arborization areas and branching patterns of the transcriptionally distinct DRG neuron subtypes. Moreover, in vivo physiological analysis revealed that each subtype has a distinct threshold and range of responses to mechanical and/or thermal stimuli. These findings support a model in which morphologically and physiologically distinct cutaneous DRG sensory neuron subtypes tile mechanical and thermal stimulus space to collectively encode a wide range of natural stimuli.

The World Health Organization declared mpox a public health emergency of international concern in July 2022. To investigate global mpox transmission and population-level changes associated with controlling spread, we built phylogeographic and phylodynamic models to analyze MPXV genomes from five global regions together with air traffic and epidemiological data. Our models reveal community transmission prior to detection, changes in case reporting throughout the epidemic, and a large degree of transmission heterogeneity. We find that viral introductions played a limited role in prolonging spread after initial dissemination, suggesting that travel bans would have had only a minor impact. We find that mpox transmission in North America began declining before more than 10% of high-risk individuals in the USA had vaccine-induced immunity. Our findings highlight the importance of broader routine specimen screening surveillance for emerging infectious diseases and of joint integration of genomic and epidemiological information for early outbreak control.

We sequenced and assembled using multiple long-read sequencing technologies the genomes of chimpanzee, bonobo, gorilla, orangutan, gibbon, macaque, owl monkey, and marmoset. We identified 1,338,997 lineage-specific fixed structural variants (SVs) disrupting 1,561 protein-coding genes and 136,932 regulatory elements, including the most complete set of human-specific fixed differences. We estimate that 819.47 Mbp or ∼27% of the genome has been affected by SVs across primate evolution. We identify 1,607 structurally divergent regions wherein recurrent structural variation contributes to creating SV hotspots where genes are recurrently lost (e.g., CARD, C4, and OLAH gene families) and additional lineage-specific genes are generated (e.g., CKAP2, VPS36, ACBD7, and NEK5 paralogs), becoming targets of rapid chromosomal diversification and positive selection (e.g., RGPD gene family). High-fidelity long-read sequencing has made these dynamic regions of the genome accessible for sequence-level analyses within and between primate species.

Apelin is a key hormone in cardiovascular homeostasis that activates the apelin receptor (APLNR), which is regarded as a promising therapeutic target for cardiovascular disease. However, adverse effects through the β-arrestin pathway limit its pharmacological use. Here, we report cryoelectron microscopy (cryo-EM) structures of APLNR-Gi1 complexes bound to three agonists with divergent signaling profiles. Combined with functional assays, we have identified "twin hotspots" in APLNR as key determinants for signaling bias, guiding the rational design of two exclusive G-protein-biased agonists WN353 and WN561. Cryo-EM structures of WN353- and WN561-stimulated APLNR-G protein complexes further confirm that the designed ligands adopt the desired poses. Pathophysiological experiments have provided evidence that WN561 demonstrates superior therapeutic effects against cardiac hypertrophy and reduced adverse effects compared with the established APLNR agonists. In summary, our designed APLNR modulator may facilitate the development of next-generation cardiovascular medications.

Neonates are highly susceptible to inflammation and infection. Here, we investigate how late fetal liver (FL) mouse hematopoietic stem and progenitor cells (HSPCs) respond to inflammation, testing the hypothesis that deficits in the engagement of emergency myelopoiesis (EM) pathways limit neutrophil output and contribute to perinatal neutropenia. We show that fetal HSPCs have limited production of myeloid cells at steady state and fail to activate a classical adult-like EM transcriptional program. Moreover, we find that fetal HSPCs can respond to EM-inducing inflammatory stimuli in vitro but are restricted by maternal anti-inflammatory factors, primarily interleukin-10 (IL-10), from activating EM pathways in utero. Accordingly, we demonstrate that the loss of maternal IL-10 restores EM activation in fetal HSPCs but at the cost of fetal demise. These results reveal the evolutionary trade-off inherent in maternal anti-inflammatory responses that maintain pregnancy but render the fetus unresponsive to EM activation signals and susceptible to infection.

Ribosome production is essential for cell growth. Approximately 200 assembly factors drive this complicated pathway that starts in the nucleolus and ends in the cytoplasm. A large number of structural snapshots of the pre-60S pathway have revealed the principles behind large subunit synthesis. To view this SnapShot, open or download the PDF.

Most membrane proteins are modified by covalent addition of complex sugars through N- and O-glycosylation. Unlike proteins, glycans do not typically adopt specific secondary structures and remain very mobile, shielding potentially large fractions of protein surface. High glycan conformational freedom hinders complete structural elucidation of glycoproteins. Computer simulations may be used to model glycosylated proteins but require hundreds of thousands of computing hours on supercomputers, thus limiting routine use. Here, we describe GlycoSHIELD, a reductionist method that can be implemented on personal computers to graft realistic ensembles of glycan conformers onto static protein structures in minutes. Using molecular dynamics simulation, small-angle X-ray scattering, cryoelectron microscopy, and mass spectrometry, we show that this open-access toolkit provides enhanced models of glycoprotein structures. Focusing on N-cadherin, human coronavirus spike proteins, and gamma-aminobutyric acid receptors, we show that GlycoSHIELD can shed light on the impact of glycans on the conformation and activity of complex glycoproteins.

While CD4+ T cell depletion is key to disease progression in people living with HIV and SIV-infected macaques, the mechanisms underlying this depletion remain incompletely understood, with most cell death involving uninfected cells. In contrast, SIV infection of "natural" hosts such as sooty mangabeys does not cause CD4+ depletion and AIDS despite high-level viremia. Here, we report that the CARD8 inflammasome is activated immediately after HIV entry by the viral protease encapsulated in incoming virions. Sensing of HIV protease activity by CARD8 leads to rapid pyroptosis of quiescent cells without productive infection, while T cell activation abolishes CARD8 function and increases permissiveness to infection. In humanized mice reconstituted with CARD8-deficient cells, CD4+ depletion is delayed despite high viremia. Finally, we discovered loss-of-function mutations in CARD8 from "natural hosts," which may explain the peculiarly non-pathogenic nature of these infections. Our study suggests that CARD8 drives CD4+ T cell depletion during pathogenic HIV/SIV infections.

Plasmids are extrachromosomal genetic elements that often encode fitness-enhancing features. However, many bacteria carry "cryptic" plasmids that do not confer clear beneficial functions. We identified one such cryptic plasmid, pBI143, which is ubiquitous across industrialized gut microbiomes and is 14 times as numerous as crAssphage, currently established as the most abundant extrachromosomal genetic element in the human gut. The majority of mutations in pBI143 accumulate in specific positions across thousands of metagenomes, indicating strong purifying selection. pBI143 is monoclonal in most individuals, likely due to the priority effect of the version first acquired, often from one's mother. pBI143 can transfer between Bacteroidales, and although it does not appear to impact bacterial host fitness in vivo, it can transiently acquire additional genetic content. We identified important practical applications of pBI143, including its use in identifying human fecal contamination and its potential as an alternative approach to track human colonic inflammatory states.

Chloroplast genes encoding photosynthesis-associated proteins are predominantly transcribed by the plastid-encoded RNA polymerase (PEP). PEP is a multi-subunit complex composed of plastid-encoded subunits similar to bacterial RNA polymerases (RNAPs) stably bound to a set of nuclear-encoded PEP-associated proteins (PAPs). PAPs are essential to PEP activity and chloroplast biogenesis, but their roles are poorly defined. Here, we present cryoelectron microscopy (cryo-EM) structures of native 21-subunit PEP and a PEP transcription elongation complex from white mustard (Sinapis alba). We identify that PAPs encase the core polymerase, forming extensive interactions that likely promote complex assembly and stability. During elongation, PAPs interact with DNA downstream of the transcription bubble and with the nascent mRNA. The models reveal details of the superoxide dismutase, lysine methyltransferase, thioredoxin, and amino acid ligase enzymes that are subunits of PEP. Collectively, these data provide a foundation for the mechanistic understanding of chloroplast transcription and its role in plant growth and adaptation.

Chloroplasts are green plastids in the cytoplasm of eukaryotic algae and plants responsible for photosynthesis. The plastid-encoded RNA polymerase (PEP) plays an essential role during chloroplast biogenesis from proplastids and functions as the predominant RNA polymerase in mature chloroplasts. The PEP-centered transcription apparatus comprises a bacterial-origin PEP core and more than a dozen eukaryotic-origin PEP-associated proteins (PAPs) encoded in the nucleus. Here, we determined the cryo-EM structures of Nicotiana tabacum (tobacco) PEP-PAP apoenzyme and PEP-PAP transcription elongation complexes at near-atomic resolutions. Our data show the PEP core adopts a typical fold as bacterial RNAP. Fifteen PAPs bind at the periphery of the PEP core, facilitate assembling the PEP-PAP supercomplex, protect the complex from oxidation damage, and likely couple gene transcription with RNA processing. Our results report the high-resolution architecture of the chloroplast transcription apparatus and provide the structural basis for the mechanistic and functional study of transcription regulation in chloroplasts.

RNA polymerases (RNAPs) control the first step of gene expression in all forms of life by transferring genetic information from DNA to RNA, a process known as transcription. In this issue of Cell, Webster et al. and Wu et al. report three-dimensional structures of RNAP complexes from chloroplasts.

Genome editing has been a transformative force in the life sciences and human medicine, offering unprecedented opportunities to dissect complex biological processes and treat the underlying causes of many genetic diseases. CRISPR-based technologies, with their remarkable efficiency and easy programmability, stand at the forefront of this revolution. In this Review, we discuss the current state of CRISPR gene editing technologies in both research and therapy, highlighting limitations that constrain them and the technological innovations that have been developed in recent years to address them. Additionally, we examine and summarize the current landscape of gene editing applications in the context of human health and therapeutics. Finally, we outline potential future developments that could shape gene editing technologies and their applications in the coming years.

Human genetics has emerged as one of the most dynamic areas of biology, with a broadening societal impact. In this review, we discuss recent achievements, ongoing efforts, and future challenges in the field. Advances in technology, statistical methods, and the growing scale of research efforts have all provided many insights into the processes that have given rise to the current patterns of genetic variation. Vast maps of genetic associations with human traits and diseases have allowed characterization of their genetic architecture. Finally, studies of molecular and cellular effects of genetic variants have provided insights into biological processes underlying disease. Many outstanding questions remain, but the field is well poised for groundbreaking discoveries as it increases the use of genetic data to understand both the history of our species and its applications to improve human health.

Growing evidence from archaic and early modern human genomes brings new insights to the emergence of modern humans. We recount recent information collected from ancient DNA studies that inform us about the evolutionary pathway to modern humanity. These findings point to both individual- and population-level advantages underlying modern human expansion.

Genomic approaches have the potential to play a pivotal role in conservation, both to detect threats to species and populations and to restore biodiversity through actions. We here separate these approaches into two subdisciplines, vulnerability and restoration genomics, and discuss current applications, outstanding questions, and future potential.