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Stable associations between partners over time are critical for the evolution of mutualism. Hosts employ a variety of mechanisms to maintain specificity with bacterial associates. Acromyrmex leaf-cutting ants farm a fungal cultivar as their primary nutrient source. These ants also carry a Pseudonocardia Actinobacteria exosymbiont on their bodies that produces antifungal compounds that help inhibit specialized parasites of the ants' fungal garden. Major workers emerge from their pupal cases (eclose) symbiont-free, but exhibit visible Actinobacterial coverage within 14 days post-eclosion. Using subcolony experiments, we investigate exosymbiont transmission within Acromyrmex colonies. We found successful transmission to newly eclosed major workers fostered by major workers with visible Actinobacteria in all cases (100% acquiring, n = 19). In contrast, newly eclosed major workers reared without exosymbiont-carrying major workers did not acquire visible Actinobacteria (0% acquiring, n = 73). We further show that the majority of ants exposed to major workers with exosymbionts within 2 hours of eclosion acquired bacteria (60.7% acquiring, n = 28), while normal acquisition did not occur when exposure occurred later than 2 hours post-eclosion (0% acquiring, n = 18). Our findings show that transmission of exosymbionts to newly eclosed major workers occurs through interactions with exosymbiont-covered workers within a narrow time window after eclosion. This mode of transmission likely helps ensure the defensive function within colonies, as well as specificity and partner fidelity in the ant-bacterium association.

Joseph Gleeson and colleagues identify CEP41 mutations as a cause of Joubert syndrome. Their functional studies suggest that CEP41 regulates ciliary entry of TTLL6, an enzyme required for tubulin glutamylation at the cilium. Tubulin glutamylation is a post-translational modification that occurs predominantly in the ciliary axoneme and has been suggested to be important for ciliary function 1 , 2 . However, its relationship to disorders of the primary cilium, termed ciliopathies, has not been explored. Here we mapped a new locus for Joubert syndrome (JBTS) 3 , which we have designated as JBTS15, and identified causative mutations in CEP41 , which encodes a 41-kDa centrosomal protein 4 . We show that CEP41 is localized to the basal body and primary cilia, and regulates ciliary entry of TTLL6, an evolutionarily conserved polyglutamylase enzyme 5 . Depletion of CEP41 causes ciliopathy-related phenotypes in zebrafish and mice and results in glutamylation defects in the ciliary axoneme. Our data identify CEP41 mutations as a cause of JBTS and implicate tubulin post-translational modification in the pathogenesis of human ciliary dysfunction.

Leaf-cutter ants are one of the most important herbivorous insects in the Neotropics, harvesting vast quantities of fresh leaf material. The ants use leaves to cultivate a fungus that serves as the colony's primary food source. This obligate ant-fungus mutualism is one of the few occurrences of farming by non-humans and likely facilitated the formation of their massive colonies. Mature leaf-cutter ant colonies contain millions of workers ranging in size from small garden tenders to large soldiers, resulting in one of the most complex polymorphic caste systems within ants. To begin uncovering the genomic underpinnings of this system, we sequenced the genome of Atta cephalotes using 454 pyrosequencing. One prediction from this ant's lifestyle is that it has undergone genetic modifications that reflect its obligate dependence on the fungus for nutrients. Analysis of this genome sequence is consistent with this hypothesis, as we find evidence for reductions in genes related to nutrient acquisition. These include extensive reductions in serine proteases (which are likely unnecessary because proteolysis is not a primary mechanism used to process nutrients obtained from the fungus), a loss of genes involved in arginine biosynthesis (suggesting that this amino acid is obtained from the fungus), and the absence of a hexamerin (which sequesters amino acids during larval development in other insects). Following recent reports of genome sequences from other insects that engage in symbioses with beneficial microbes, the A. cephalotes genome provides new insights into the symbiotic lifestyle of this ant and advances our understanding of host-microbe symbioses.

Leaf-cutter ants are one of the most important herbivorous insects in the Neotropics, harvesting vast quantities of fresh leaf material. The ants use leaves to cultivate a fungus that serves as the colony's primary food source. This obligate ant-fungus mutualism is one of the few occurrences of farming by non-humans and likely facilitated the formation of their massive colonies. Mature leaf-cutter ant colonies contain millions of workers ranging in size from small garden tenders to large soldiers, resulting in one of the most complex polymorphic caste systems within ants. To begin uncovering the genomic underpinnings of this system, we sequenced the genome of Atta cephalotes using 454 pyrosequencing. One prediction from this ant's lifestyle is that it has undergone genetic modifications that reflect its obligate dependence on the fungus for nutrients. Analysis of this genome sequence is consistent with this hypothesis, as we find evidence for reductions in genes related to nutrient acquisition. These include extensive reductions in serine proteases (which are likely unnecessary because proteolysis is not a primary mechanism used to process nutrients obtained from the fungus), a loss of genes involved in arginine biosynthesis (suggesting that this amino acid is obtained from the fungus), and the absence of a hexamerin (which sequesters amino acids during larval development in other insects). Following recent reports of genome sequences from other insects that engage in symbioses with beneficial microbes, the A. cephalotes genome provides new insights into the symbiotic lifestyle of this ant and advances our understanding of host-microbe symbioses.

A key aspect of nearly all single-cell sequencing experiments is dissociation of intact tissues into single-cell suspensions. While many protocols have been optimized for optimal cell yield, they have often overlooked the effects that dissociation can have on ex vivo gene expression. Here, we demonstrate that use of enzymatic dissociation on brain tissue induces an aberrant ex vivo gene expression signature, most prominently in microglia, which is prevalent in published literature and can substantially confound downstream analyses. To address this issue, we present a rigorously validated protocol that preserves both in vivo transcriptional profiles and cell-type diversity and yield across tissue types and species. We also identify a similar signature in postmortem human brain single-nucleus RNA-sequencing datasets, and show that this signature is induced in freshly isolated human tissue by exposure to elevated temperatures ex vivo. Together, our results provide a methodological solution for preventing artifactual gene expression changes during fresh tissue digestion and a reference for future deeper analysis of the potential confounding states present in postmortem human samples.Marsh et al. demonstrate that enzymatic dissociation induces an aberrant ex vivo gene expression signature, most prominently in microglia, which when not addressed can substantially confound downstream analyses. They also identify a similar signature in postmortem human brain in snRNA-seq.

Microglia, the macrophages of the brain parenchyma, are key players in neurodegenerative diseases such as Alzheimer’s disease. These cells adopt distinct transcriptional subtypes known as states. Understanding state function, especially in human microglia, has been elusive owing to a lack of tools to model and manipulate these cells. Here, we developed a platform for modeling human microglia transcriptional states in vitro. We found that exposure of human stem-cell-differentiated microglia to synaptosomes, myelin debris, apoptotic neurons or synthetic amyloid-beta fibrils generated transcriptional diversity that mapped to gene signatures identified in human brain microglia, including disease-associated microglia, a state enriched in neurodegenerative diseases. Using a new lentiviral approach, we demonstrated that the transcription factor MITF drives a disease-associated transcriptional signature and a highly phagocytic state. Together, these tools enable the manipulation and functional interrogation of human microglial states in both homeostatic and disease-relevant contexts. Stevens and colleagues show that human stem-cell-differentiated microglia can be used to model the extensive transcriptional diversity of human brain microglia.

A knowledge of protein structure greatly enhances our understanding of protein function. In many cases, function depends on oligomerization. Ahnert et al. used mass spectrometry data together with a large-scale analysis of structures of protein complexes to examine the fundamental steps of protein assembly. Systematically combining assembly steps revealed a large set of quaternary topologies that were organized into a periodic table. Based on this table, the authors accurately predicted the expected frequencies of quaternary structure topologies. Science , this issue p. 10.1126/science.aaa2245 Understanding the organizing principles allows exhaustive enumeration of possible protein quaternary structure topologies. Structural insights into protein complexes have had a broad impact on our understanding of biological function and evolution. In this work, we sought a comprehensive understanding of the general principles underlying quaternary structure organization in protein complexes. We first examined the fundamental steps by which protein complexes can assemble, using experimental and structure-based characterization of assembly pathways. Most assembly transitions can be classified into three basic types, which can then be used to exhaustively enumerate a large set of possible quaternary structure topologies. These topologies, which include the vast majority of observed protein complex structures, enable a natural organization of protein complexes into a periodic table. On the basis of this table, we can accurately predict the expected frequencies of quaternary structure topologies, including those not yet observed. These results have important implications for quaternary structure prediction, modeling, and engineering.

Clonal hematopoiesis of indeterminate potential (CHIP) increases rapidly in prevalence beyond age 60 and has been associated with increased risk for malignancy, heart disease and ischemic stroke. CHIP is driven by somatic mutations in hematopoietic stem and progenitor cells (HSPCs). Because mutations in HSPCs often drive leukemia, we hypothesized that HSPC fitness substantially contributes to transformation from CHIP to leukemia. HSPC fitness is defined as the proliferative advantage over cells carrying no or only neutral mutations. If mutations in different genes lead to distinct fitness advantages, this could enable patient stratification. We quantified the fitness effects of mutations over 12 years in older age using longitudinal sequencing and developed a filtering method that considers individual mutational context alongside mutation co-occurrence to quantify the growth potential of variants within individuals. We found that gene-specific fitness differences can outweigh inter-individual variation and, therefore, could form the basis for personalized clinical management. An analysis of blood samples from longitudinal cohorts reveals insights on the dynamics of clonal hematopoiesis of indeterminate potential and proposes a model that could be used for individualized clone monitoring over time.

InterFeron Inducible TransMembrane proteins 1-3 (IFITM1, IFITM2 and IFITM3) are a family of proteins capable of inhibiting the cellular entry of numerous human and animal viruses. IFITM1-3 are unique amongst the currently described viral restriction factors in their apparent ability to block viral entry. This restrictive property is dependant on the localisation of the proteins to plasma and endosomal membranes, which constitute the main portals of viral entry into cells. The topology of the IFITM proteins within cell membranes is an unresolved aspect of their biology. Here we present data from immunofluorescence microscopy, protease cleavage, biotin-labelling and immuno-electron microscopy assays, showing that human IFITM1 has a membrane topology in which the N-terminal domain resides in the cytoplasm, and the C-terminal domain is extracellular. Furthermore, we provide evidence that this topology is conserved for all of the human interferon-induced IFITM proteins. This model is consistent with that recently proposed for murine IFITM3, but differs from that proposed for murine IFITM1.