Fear-related memory traces are encoded by sparse populations of hippocampal principal neurons that are recruited based on their inhibitory-excitatory balance during memory formation. Later, the reactivation of the same principal neurons can recall the memory. The details of this mechanism are still unclear. Here, we investigated whether disinhibition could play a major role in this process. Using optogenetic behavioral experiments, we found that when fear was associated with the inhibition of mouse hippocampal somatostatin positive interneurons, the re-inhibition of the same interneurons could recall fear memory. Pontine nucleus incertus neurons selectively inhibit hippocampal somatostatin cells. We also found that when fear was associated with the activity of these incertus neurons or fibers, the reactivation of the same incertus neurons or fibers could also recall fear memory. These incertus neurons showed correlated activity with hippocampal principal neurons during memory recall and were strongly innervated by memory-related neocortical centers, from which the inputs could also control hippocampal disinhibition in vivo. Nonselective inhibition of these mouse hippocampal somatostatin or incertus neurons impaired memory recall. Our data suggest a novel disinhibition-based memory mechanism in the hippocampus that is supported by local somatostatin interneurons and their pontine brainstem inputs.

Rab26 is known to regulate multiple membrane trafficking events, but its role in insulin secretion in pancreatic β cells remains unclear despite it was first identified in the pancreas. In this study, we generated Rab26-/- mice through CRISPR/Cas9 technique. Surprisingly, insulin levels in the blood of the Rab26-/- mice do not decrease upon glucose stimulation but conversely increase. Deficiency of Rab26 promotes insulin secretion, which was independently verified by Rab26 knockdown in pancreatic insulinoma cells. Conversely, overexpression of Rab26 suppresses insulin secretion in both insulinoma cell lines and isolated mouse islets. Islets overexpressing Rab26, upon transplantation, also failed to restore glucose homeostasis in type 1 diabetic mice. Immunofluorescence microscopy revealed that overexpression of Rab26 results in clustering of insulin granules. GST-pulldown experiments reveal that Rab26 interacts with synaptotagmin-1 (Syt1) through directly binding to its C2A domain, which interfering with the interaction between Syt1 and SNAP25, and consequently inhibiting the exocytosis of newcomer insulin granules revealed by TIRF microscopy. Our results suggest that Rab26 serves as a negative regulator of insulin secretion, via suppressing insulin granule fusion with plasma membrane through sequestering Syt1.

Dynamic reorganization of the cytoplasm is key to many core cellular processes, such as cell division, cell migration, and cell polarization. Cytoskeletal rearrangements are thought to constitute the main drivers of cytoplasmic flows and reorganization. In contrast, remarkably little is known about how dynamic changes in size and shape of cell organelles affect cytoplasmic organization. Here, we show that within the maturing zebrafish oocyte, the surface localization of exocytosis-competent cortical granules (Cgs) upon germinal vesicle breakdown (GVBD) is achieved by the combined activities of yolk granule (Yg) fusion and microtubule aster formation and translocation. We find that Cgs are moved towards the oocyte surface through radially outward cytoplasmic flows induced by Ygs fusing and compacting towards the oocyte center in response to GVBD. We further show that vesicles decorated with the small Rab GTPase Rab11, a master regulator of vesicular trafficking and exocytosis, accumulate together with Cgs at the oocyte surface. This accumulation is achieved by Rab11-positive vesicles being transported by acentrosomal microtubule asters, the formation of which is induced by the release of CyclinB/Cdk1 upon GVBD, and which display a net movement towards the oocyte surface by preferentially binding to the oocyte actin cortex. We finally demonstrate that the decoration of Cgs by Rab11 at the oocyte surface is needed for Cg exocytosis and subsequent chorion elevation, a process central in egg activation. Collectively, these findings unravel a yet unrecognized role of organelle fusion, functioning together with cytoskeletal rearrangements, in orchestrating cytoplasmic organization during oocyte maturation.

The standard consolidation theory suggests that the hippocampus (HPC) is critically involved in acquiring new memory, while storage and recall gradually become independent of it. Converging studies have shown separate involvements of the perirhinal cortex (PRC) and parahippocampal cortex (PHC) in item and spatial processes, whereas HPC relates the item to a spatial context. These 2 strands of literature raise the following question; which brain region is involved in the recall process of item-location associative memory? To solve this question, this study applied an item-location associative (ILA) paradigm in a single-unit study of nonhuman primates. We trained 2 macaques to associate 4 visual item pairs with 4 locations on a background map in an allocentric manner before the recording sessions. In each trial, 1 visual item and the map image at a tilt (-90° to 90°) were sequentially presented as the item-cue and the context-cue, respectively. The macaques chose the item-cue location relative to the context-cue by positioning their gaze. Neurons in the PRC, PHC, and HPC, but not area TE, exhibited item-cue responses which signaled retrieval of item-location associative memory. This retrieval signal first appeared in the PRC, followed by the HPC and PHC. We examined whether neural representations of the retrieved locations were related to the external space that the macaques viewed. A positive representation similarity was found in the HPC and PHC, but not in the PRC, thus suggesting a contribution of the HPC to relate the retrieved location from the PRC with a first-person perspective of the subjects and provide the self-referenced retrieved location to the PHC. These results imply distinct but complementary contributions of the PRC and HPC to recall of item-location associative memory that can be used across multiple spatial contexts.

Meiotic drive loci distort the normally equal segregation of alleles, which benefits their own transmission even in the face of severe fitness costs to their host organism. However, relatively little is known about the molecular identity of meiotic drivers, their strategies of action, and mechanisms that can suppress their activity. Here, we present data from the fruitfly Drosophila simulans that address these questions. We show that a family of de novo, protamine-derived X-linked selfish genes (the Dox gene family) is silenced by a pair of newly emerged hairpin RNA (hpRNA) small interfering RNA (siRNA)-class loci, Nmy and Tmy. In the w[XD1] genetic background, knockout of nmy derepresses Dox and MDox in testes and depletes male progeny, whereas knockout of tmy causes misexpression of PDox genes and renders males sterile. Importantly, genetic interactions between nmy and tmy mutant alleles reveal that Tmy also specifically maintains male progeny for normal sex ratio. We show the Dox loci are functionally polymorphic within D. simulans, such that both nmy-associated sex ratio bias and tmy-associated sterility can be rescued by wild-type X chromosomes bearing natural deletions in different Dox family genes. Finally, using tagged transgenes of Dox and PDox2, we provide the first experimental evidence Dox family genes encode proteins that are strongly derepressed in cognate hpRNA mutants. Altogether, these studies support a model in which protamine-derived drivers and hpRNA suppressors drive repeated cycles of sex chromosome conflict and resolution that shape genome evolution and the genetic control of male gametogenesis.

In recent years, there has been a strong drive to improve the inclusion of animals of both sexes in the design of in vivo research studies, driven by a need to increase sex representation in fundamental biology and drug development. This has resulted in inclusion mandates by funding bodies and journals, alongside numerous published manuscripts highlighting the issue and providing guidance to scientists. However, progress is slow and barriers to the routine use of both sexes remain. A frequent, major concern is the perceived need for a higher overall sample size to achieve an equivalent level of statistical power, which would result in an increased ethical and resource burden. This perception arises from either the belief that sex inclusion will increase variability in the data (either through a baseline difference or a treatment effect that depends on sex), thus reducing the sensitivity of statistical tests, or from misapprehensions about the correct way to analyse the data, including disaggregation or pooling by sex. Here, we conduct an in-depth examination of the consequences of including both sexes on statistical power. We performed simulations by constructing artificial datasets that encompass a range of outcomes that may occur in studies studying a treatment effect in the context of both sexes. This includes both baseline sex differences and situations in which the size of the treatment effect depends on sex in both the same and opposite directions. The data were then analysed using either a factorial analysis approach, which is appropriate for the design, or a t test approach following pooling or disaggregation of the data, which are common but erroneous strategies. The results demonstrate that there is no loss of power to detect treatment effects when splitting the sample size across sexes in most scenarios, providing that the data are analysed using an appropriate factorial analysis method (e.g., two-way ANOVA). In the rare situations where power is lost, the benefit of understanding the role of sex outweighs the power considerations. Additionally, use of the inappropriate analysis pipelines results in a loss of statistical power. Therefore, we recommend analysing data collected from both sexes using factorial analysis and splitting the sample size across male and female mice as a standard strategy.

Humans can easily tune in to one talker in a multitalker environment while still picking up bits of background speech; however, it remains unclear how we perceive speech that is masked and to what degree non-target speech is processed. Some models suggest that perception can be achieved through glimpses, which are spectrotemporal regions where a talker has more energy than the background. Other models, however, require the recovery of the masked regions. To clarify this issue, we directly recorded from primary and non-primary auditory cortex (AC) in neurosurgical patients as they attended to one talker in multitalker speech and trained temporal response function models to predict high-gamma neural activity from glimpsed and masked stimulus features. We found that glimpsed speech is encoded at the level of phonetic features for target and non-target talkers, with enhanced encoding of target speech in non-primary AC. In contrast, encoding of masked phonetic features was found only for the target, with a greater response latency and distinct anatomical organization compared to glimpsed phonetic features. These findings suggest separate mechanisms for encoding glimpsed and masked speech and provide neural evidence for the glimpsing model of speech perception.

Orcinol glucoside (OG), mainly found in the rhizome of the traditional Chinese herb Curculigo orchioides Gaertn, is noted for its antidepressant effects. In this study, an efficient screening pipeline was established for identifying the highly active orcinol synthase (ORS) and UDP-dependent glycosyltransferase (UGT) involved in the biosynthesis of OG by combining transcriptome analysis, structure-based virtual screening, and in vitro enzyme activity assays. By enhancing the downstream pathway, metabolic engineering and fermentation optimization, the OG production in Yarrowia lipolytica was improved 100-fold, resulting in a final yield of 43.46 g/L (0.84 g/g DCW), which is almost 6,400-fold higher than the extraction yield from C. orchioides roots. This study provides a reference for rapid identification of functional genes and high-yield production of natural products.

Characterizing cellular diversity at different levels of biological organization and across data modalities is a prerequisite to understanding the function of cell types in the brain. Classification of neurons is also essential to manipulate cell types in controlled ways and to understand their variation and vulnerability in brain disorders. The BRAIN Initiative Cell Census Network (BICCN) is an integrated network of data-generating centers, data archives, and data standards developers, with the goal of systematic multimodal brain cell type profiling and characterization. Emphasis of the BICCN is on the whole mouse brain with demonstration of prototype feasibility for human and nonhuman primate (NHP) brains. Here, we provide a guide to the cellular and spatial approaches employed by the BICCN, and to accessing and using these data and extensive resources, including the BRAIN Cell Data Center (BCDC), which serves to manage and integrate data across the ecosystem. We illustrate the power of the BICCN data ecosystem through vignettes highlighting several BICCN analysis and visualization tools. Finally, we present emerging standards that have been developed or adopted toward Findable, Accessible, Interoperable, and Reusable (FAIR) neuroscience. The combined BICCN ecosystem provides a comprehensive resource for the exploration and analysis of cell types in the brain.

In the past 2 decades, structural biology has transformed from a single technique used on single proteins to a multimodal integrative approach. Recently, protein structure prediction algorithms have opened new avenues to address challenging biological questions.

The primate brain has unique anatomical characteristics, which translate into advanced cognitive, sensory, and motor abilities. Thus, it is important that we gain insight on its structure to provide a solid basis for models that will clarify function. Here, we report on the implementation and features of the Brain/MINDS Marmoset Connectivity Resource (BMCR), a new open-access platform that provides access to high-resolution anterograde neuronal tracer data in the marmoset brain, integrated to retrograde tracer and tractography data. Unlike other existing image explorers, the BMCR allows visualization of data from different individuals and modalities in a common reference space. This feature, allied to an unprecedented high resolution, enables analyses of features such as reciprocity, directionality, and spatial segregation of connections. The present release of the BMCR focuses on the prefrontal cortex (PFC), a uniquely developed region of the primate brain that is linked to advanced cognition, including the results of 52 anterograde and 164 retrograde tracer injections in the cortex of the marmoset. Moreover, the inclusion of tractography data from diffusion MRI allows systematic analyses of this noninvasive modality against gold-standard cellular connectivity data, enabling detection of false positives and negatives, which provide a basis for future development of tractography. This paper introduces the BMCR image preprocessing pipeline and resources, which include new tools for exploring and reviewing the data.

The basement membrane (BM) is a specialized extracellular matrix (ECM), which underlies or encases developing tissues. Mechanical properties of encasing BMs have been shown to profoundly influence the shaping of associated tissues. Here, we use the migration of the border cells (BCs) of the Drosophila egg chamber to unravel a new role of encasing BMs in cell migration. BCs move between a group of cells, the nurse cells (NCs), that are enclosed by a monolayer of follicle cells (FCs), which is, in turn, surrounded by a BM, the follicle BM. We show that increasing or reducing the stiffness of the follicle BM, by altering laminins or type IV collagen levels, conversely affects BC migration speed and alters migration mode and dynamics. Follicle BM stiffness also controls pairwise NC and FC cortical tension. We propose that constraints imposed by the follicle BM influence NC and FC cortical tension, which, in turn, regulate BC migration. Encasing BMs emerge as key players in the regulation of collective cell migration during morphogenesis.

The active state of centromeres is epigenetically defined by the presence of CENP-A interspersed with histone H3 nucleosomes. While the importance of dimethylation of H3K4 for centromeric transcription has been highlighted in various studies, the identity of the enzyme(s) depositing these marks on the centromere is still unknown. The MLL (KMT2) family plays a crucial role in RNA polymerase II (Pol II)-mediated gene regulation by methylating H3K4. Here, we report that MLL methyltransferases regulate transcription of human centromeres. CRISPR-mediated down-regulation of MLL causes loss of H3K4me2, resulting in an altered epigenetic chromatin state of the centromeres. Intriguingly, our results reveal that loss of MLL, but not SETD1A, increases co-transcriptional R-loop formation, and Pol II accumulation at the centromeres. Finally, we report that the presence of MLL and SETD1A is crucial for kinetochore maintenance. Altogether, our data reveal a novel molecular framework where both the H3K4 methylation mark and the methyltransferases regulate stability and identity of the centromere.

A defining property of circadian clocks is temperature compensation, characterized by the resilience of their near 24-hour free-running periods against changes in environmental temperature within the physiological range. While temperature compensation is evolutionary conserved across different taxa of life and has been studied within many model organisms, its molecular underpinnings remain elusive. Posttranscriptional regulations such as temperature-sensitive alternative splicing or phosphorylation have been described as underlying reactions. Here, we show that knockdown of cleavage and polyadenylation specificity factor subunit 6 (CPSF6), a key regulator of 3'-end cleavage and polyadenylation, significantly alters circadian temperature compensation in human U-2 OS cells. We apply a combination of 3'-end-RNA-seq and mass spectrometry-based proteomics to globally quantify changes in 3' UTR length as well as gene and protein expression between wild-type and CPSF6 knockdown cells and their dependency on temperature. Since changes in temperature compensation behavior should be reflected in alterations of temperature responses within one or all of the 3 regulatory layers, we statistically assess differential responses upon changes in ambient temperature between wild-type and CPSF6 knockdown cells. By this means, we reveal candidate genes underlying circadian temperature compensation, including eukaryotic translation initiation factor 2 subunit 1 (EIF2S1).

T-cell acute lymphoblastic leukemia (T-ALL) protein 1 (TAL1) is a central transcription factor in hematopoiesis. The timing and level of TAL1 expression orchestrate the differentiation to specialized blood cells and its overexpression is a common cause of T-ALL. Here, we studied the 2 protein isoforms of TAL1, short and long, which are generated by the use of alternative promoters as well as by alternative splicing. We analyzed the expression of each isoform by deleting an enhancer or insulator, or by opening chromatin at the enhancer location. Our results show that each enhancer promotes expression from a specific TAL1 promoter. Expression from a specific promoter gives rise to a unique 5' UTR with differential regulation of translation. Moreover, our study suggests that the enhancers regulate TAL1 exon 3 alternative splicing by inducing changes in the chromatin at the splice site, which we demonstrate is mediated by KMT2B. Furthermore, our results indicate that TAL1-short binds more strongly to TAL1 E-protein partners and functions as a stronger transcription factor than TAL1-long. Specifically TAL1-short has a unique transcription signature promoting apoptosis. Finally, when we expressed both isoforms in mice bone marrow, we found that while overexpression of both isoforms prevents lymphoid differentiation, expression of TAL1-short alone leads to hematopoietic stem cell exhaustion. Furthermore, we found that TAL1-short promoted erythropoiesis and reduced cell survival in the CML cell line K562. While TAL1 and its partners are considered promising therapeutic targets in the treatment of T-ALL, our results show that TAL1-short could act as a tumor suppressor and suggest that altering TAL1 isoform's ratio could be a preferred therapeutic approach.

To respond to the world around them, animals rely on the input of a network of sensory organs distributed throughout the body. Distinct classes of sensory organs are specialized for the detection of specific stimuli such as strain, pressure, or taste. The features that underlie this specialization relate both to the neurons that innervate sensory organs and the accessory cells they comprise. To understand the genetic basis of this diversity of cell types, both within and between sensory organs, we performed single-cell RNA sequencing on the first tarsal segment of the male Drosophila melanogaster foreleg during pupal development. This tissue displays a wide variety of functionally and structurally distinct sensory organs, including campaniform sensilla, mechanosensory bristles, and chemosensory taste bristles, as well as the sex comb, a recently evolved male-specific structure. In this study, we characterize the cellular landscape in which the sensory organs reside, identify a novel cell type that contributes to the construction of the neural lamella, and resolve the transcriptomic differences among support cells within and between sensory organs. We identify the genes that distinguish between mechanosensory and chemosensory neurons, resolve a combinatorial transcription factor code that defines 4 distinct classes of gustatory neurons and several types of mechanosensory neurons, and match the expression of sensory receptor genes to specific neuron classes. Collectively, our work identifies core genetic features of a variety of sensory organs and provides a rich, annotated resource for studying their development and function.

Technological advancements in biology and microscopy have empowered a transition from bioimaging as an observational method to a quantitative one. However, as biologists are adopting quantitative bioimaging and these experiments become more complex, researchers need additional expertise to carry out this work in a rigorous and reproducible manner. This Essay provides a navigational guide for experimental biologists to aid understanding of quantitative bioimaging from sample preparation through to image acquisition, image analysis, and data interpretation. We discuss the interconnectedness of these steps, and for each, we provide general recommendations, key questions to consider, and links to high-quality open-access resources for further learning. This synthesis of information will empower biologists to plan and execute rigorous quantitative bioimaging experiments efficiently.

Intrinsically generated neural activity propagates through the developing auditory system to promote maturation and refinement of sound processing circuits prior to hearing onset. This early patterned activity is induced by non-sensory supporting cells in the organ of Corti, which are highly interconnected through gap junctions containing connexin 26 (Gjb2). Although loss of function mutations in Gjb2 impair cochlear development and are the most common cause of congenital deafness, it is not known if these variants disrupt spontaneous activity and the developmental trajectory of sound processing circuits in the brain. Here, we show in a new mouse model of Gjb2-mediated congenital deafness that cochlear supporting cells adjacent to inner hair cells (IHCs) unexpectedly retain intercellular coupling and the capacity to generate spontaneous activity, exhibiting only modest deficits prior to hearing onset. Supporting cells lacking Gjb2 elicited coordinated activation of IHCs, leading to coincident bursts of activity in central auditory neurons that will later process similar frequencies of sound. Despite alterations in the structure of the sensory epithelium, hair cells within the cochlea of Gjb2-deficient mice were intact and central auditory neurons could be activated within appropriate tonotopic domains by loud sounds at hearing onset, indicating that early maturation and refinement of auditory circuits was preserved. Only after cessation of spontaneous activity following hearing onset did progressive hair cell degeneration and enhanced auditory neuron excitability manifest. This preservation of cochlear spontaneous neural activity in the absence of connexin 26 may increase the effectiveness of early therapeutic interventions to restore hearing.

Lipoic acid is an essential biomolecule found in all domains of life and is involved in central carbon metabolism and dissimilatory sulfur oxidation. The machineries for lipoate assembly in mitochondria and chloroplasts of higher eukaryotes, as well as in the apicoplasts of some protozoa, are all of prokaryotic origin. Here, we provide experimental evidence for a novel lipoate assembly pathway in bacteria based on a sLpl(AB) lipoate:protein ligase, which attaches octanoate or lipoate to apo-proteins, and 2 radical SAM proteins, LipS1 and LipS2, which work together as lipoyl synthase and insert 2 sulfur atoms. Extensive homology searches combined with genomic context analyses allowed us to precisely distinguish between the new and established pathways and map them on the tree of life. This not only revealed a much wider distribution of lipoate biogenesis systems than expected, in particular, the novel sLpl(AB)-LipS1/S2 pathway, and indicated a highly modular nature of the enzymes involved, with unforeseen combinations, but also provided a new framework for the evolution of lipoate assembly. Our results show that dedicated machineries for both de novo lipoate biogenesis and scavenging from the environment were implemented early in evolution and that their distribution in the 2 prokaryotic domains was shaped by a complex network of horizontal gene transfers, acquisition of additional genes, fusions, and losses. Our large-scale phylogenetic analyses identify the bipartite archaeal LplAB ligase as the ancestor of the bacterial sLpl(AB) proteins, which were obtained by horizontal gene transfer. LipS1/S2 have a more complex evolutionary history with multiple of such events but probably also originated in the domain archaea.

Mas-related G-protein-coupled receptors X1-X4 (MRGPRX1-X4) are 4 primate-specific receptors that are recently reported to be responsible for many biological processes, including itch sensation, pain transmission, and inflammatory reactions. MRGPRX1 is the first identified human MRGPR, and its expression is restricted to primary sensory neurons. Due to its dual roles in itch and pain signaling pathways, MRGPRX1 has been regarded as a promising target for itch remission and pain inhibition. Here, we reported a cryo-electron microscopy (cryo-EM) structure of Gq-coupled MRGPRX1 in complex with a synthetic agonist compound 16 in an active conformation at an overall resolution of 3.0 Å via a NanoBiT tethering strategy. Compound 16 is a new pain-relieving compound with high potency and selectivity to MRGPRX1 over other MRGPRXs and opioid receptor. MRGPRX1 was revealed to share common structural features of the Gq-mediated receptor activation mechanism of MRGPRX family members, but the variable residues in orthosteric pocket of MRGPRX1 exhibit the unique agonist recognition pattern, potentially facilitating to design MRGPRX1-specific modulators. Together with receptor activation and itch behavior evaluation assays, our study provides a structural snapshot to modify therapeutic molecules for itch relieving and analgesia targeting MRGPRX1.

Numerous, diverse plant viruses encode movement proteins (MPs) that aid the virus movement through plasmodesmata, the plant intercellular channels. MPs are essential for virus spread and propagation in distal tissues, and several unrelated MPs have been identified. The 30K superfamily of MPs (named after the molecular mass of tobacco mosaic virus MP, the classical model of plant virology) is the largest and most diverse MP variety, represented in 16 virus families, but its evolutionary origin remained obscure. Here, we show that the core structural domain of the 30K MPs is homologous to the jelly-roll domain of the capsid proteins (CPs) of small RNA and DNA viruses, in particular, those infecting plants. The closest similarity was observed between the 30K MPs and the CPs of the viruses in the families Bromoviridae and Geminiviridae. We hypothesize that the MPs evolved via duplication or horizontal acquisition of the CP gene in a virus that infected an ancestor of vascular plants, followed by neofunctionalization of one of the paralogous CPs, potentially through the acquisition of unique N- and C-terminal regions. During the subsequent coevolution of viruses with diversifying vascular plants, the 30K MP genes underwent explosive horizontal spread among emergent RNA and DNA viruses, likely permitting viruses of insects and fungi that coinfected plants to expand their host ranges, molding the contemporary plant virome.

The immune response to Mycobacterium tuberculosis infection determines tuberculosis disease outcomes, yet we have an incomplete understanding of what immune factors contribute to a protective immune response. Neutrophilic inflammation has been associated with poor disease prognosis in humans and in animal models during M. tuberculosis infection and, therefore, must be tightly regulated. ATG5 is an essential autophagy protein that is required in innate immune cells to control neutrophil-dominated inflammation and promote survival during M. tuberculosis infection; however, the mechanistic basis for how ATG5 regulates neutrophil recruitment is unknown. To interrogate what innate immune cells require ATG5 to control neutrophil recruitment during M. tuberculosis infection, we used different mouse strains that conditionally delete Atg5 in specific cell types. We found that ATG5 is required in CD11c+ cells (lung macrophages and dendritic cells) to control the production of proinflammatory cytokines and chemokines during M. tuberculosis infection, which would otherwise promote neutrophil recruitment. This role for ATG5 is autophagy dependent, but independent of mitophagy, LC3-associated phagocytosis, and inflammasome activation, which are the most well-characterized ways that autophagy proteins regulate inflammation. In addition to the increased proinflammatory cytokine production from macrophages during M. tuberculosis infection, loss of ATG5 in innate immune cells also results in an early induction of TH17 responses. Despite prior published in vitro cell culture experiments supporting a role for autophagy in controlling M. tuberculosis replication in macrophages, the effects of autophagy on inflammatory responses occur without changes in M. tuberculosis burden in macrophages. These findings reveal new roles for autophagy proteins in lung resident macrophages and dendritic cells that are required to suppress inflammatory responses that are associated with poor control of M. tuberculosis infection.

Bak is a critical executor of apoptosis belonging to the Bcl-2 protein family. Bak contains a hydrophobic groove where the BH3 domain of proapoptotic Bcl-2 family members can be accommodated, which initiates its activation. Once activated, Bak undergoes a conformational change to oligomerize, which leads to mitochondrial destabilization and the release of cytochrome c into the cytosol and eventual apoptotic cell death. In this study, we investigated the molecular aspects and functional consequences of the interaction between Bak and peroxisomal testis-specific 1 (Pxt1), a noncanonical BH3-only protein exclusively expressed in the testis. Together with various biochemical approaches, this interaction was verified and analyzed at the atomic level by determining the crystal structure of the Bak-Pxt1 BH3 complex. In-depth biochemical and cellular analyses demonstrated that Pxt1 functions as a Bak-activating proapoptotic factor, and its BH3 domain, which mediates direct intermolecular interaction with Bak, plays a critical role in triggering apoptosis. Therefore, this study provides a molecular basis for the Pxt1-mediated novel pathway for the activation of apoptosis and expands our understanding of the cell death signaling coordinated by diverse BH3 domain-containing proteins.

Pluripotency defines the unlimited potential of individual cells of vertebrate embryos, from which all adult somatic cells and germ cells are derived. Understanding how the programming of pluripotency evolved has been obscured in part by a lack of data from lower vertebrates; in model systems such as frogs and zebrafish, the function of the pluripotency genes NANOG and POU5F1 have diverged. Here, we investigated how the axolotl ortholog of NANOG programs pluripotency during development. Axolotl NANOG is absolutely required for gastrulation and germ-layer commitment. We show that in axolotl primitive ectoderm (animal caps; ACs) NANOG and NODAL activity, as well as the epigenetic modifying enzyme DPY30, are required for the mass deposition of H3K4me3 in pluripotent chromatin. We also demonstrate that all 3 protein activities are required for ACs to establish the competency to differentiate toward mesoderm. Our results suggest the ancient function of NANOG may be establishing the competence for lineage differentiation in early cells. These observations provide insights into embryonic development in the tetrapod ancestor from which terrestrial vertebrates evolved.

Auxin response factors (ARFs) are a family of transcription factors that are responsible for regulating gene expression in response to changes in auxin level. The analysis of ARF sequence and activity indicates that there are 2 major groups: activators and repressors. One clade of ARFs, clade-D, is sister to clade-A activating ARFs, but are unique in that they lack a DNA-binding domain. Clade-D ARFs are present in lycophytes and bryophytes but absent in other plant lineages. The transcriptional activity of clade-D ARFs, as well as how they regulate gene expression, is not well understood. Here, we report that clade-D ARFs are transcriptional activators in the model bryophyte Physcomitrium patens and have a major role in the development of this species. Δarfddub protonemata exhibit a delay in filament branching, as well as a delay in the chloronema to caulonema transition. Additionally, leafy gametophore development in Δarfddub lines lags behind wild type. We present evidence that ARFd1 interacts with activating ARFs via their PB1 domains, but not with repressing ARFs. Based on these results, we propose a model in which clade-D ARFs enhance gene expression by interacting with DNA bound clade-A ARFs. Further, we show that ARFd1 must form oligomers for full activity.

"Lifelong Learning with Friends" provides diversity training to college students by having them learn science alongside adults with intellectual and developmental disabilities (IDDs). Volunteers showed increased interest in IDD-focused research, social interaction, and advocacy.

Identifying host genes essential for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has the potential to reveal novel drug targets and further our understanding of Coronavirus Disease 2019 (COVID-19). We previously performed a genome-wide CRISPR/Cas9 screen to identify proviral host factors for highly pathogenic human coronaviruses. Few host factors were required by diverse coronaviruses across multiple cell types, but DYRK1A was one such exception. Although its role in coronavirus infection was previously undescribed, DYRK1A encodes Dual Specificity Tyrosine Phosphorylation Regulated Kinase 1A and is known to regulate cell proliferation and neuronal development. Here, we demonstrate that DYRK1A regulates ACE2 and DPP4 transcription independent of its catalytic kinase function to support SARS-CoV, SARS-CoV-2, and Middle East Respiratory Syndrome Coronavirus (MERS-CoV) entry. We show that DYRK1A promotes DNA accessibility at the ACE2 promoter and a putative distal enhancer, facilitating transcription and gene expression. Finally, we validate that the proviral activity of DYRK1A is conserved across species using cells of nonhuman primate and human origin. In summary, we report that DYRK1A is a novel regulator of ACE2 and DPP4 expression that may dictate susceptibility to multiple highly pathogenic human coronaviruses.

Sensory perception modulates aging, yet we know little about how. An understanding of the neuronal mechanisms through which animals orchestrate biological responses to relevant sensory inputs would provide insight into the control systems that may be important for modulating lifespan. Here, we provide new awareness into how the perception of dead conspecifics, or death perception, which elicits behavioral and physiological effects in many different species, affects lifespan in the fruit fly, Drosophila melanogaster. Previous work demonstrated that cohousing Drosophila with dead conspecifics decreases fat stores, reduces starvation resistance, and accelerates aging in a manner that requires both sight and the serotonin receptor 5-HT2A. In this manuscript, we demonstrate that a discrete, 5-HT2A-expressing neural population in the ellipsoid body (EB) of the Drosophila central complex, identified as R2/R4 neurons, acts as a rheostat and plays an important role in transducing sensory information about the presence of dead individuals to modulate lifespan. Expression of the insulin-responsive transcription factor foxo in R2/R4 neurons and insulin-like peptides dilp3 and dilp5, but not dilp2, are required, with the latter likely altered in median neurosecretory cells (MNCs) after R2/R4 neuronal activation. These data generate new insights into the neural underpinnings of how perceptive events may impact aging and physiology across taxa.

Recent evidence suggests that at least some insect species might plausibly feel pain. These findings should prompt researchers to think about the welfare implications of insect experiments.

Adapting actions to changing goals and environments is central to intelligent behavior. There is evidence that the basal ganglia play a crucial role in reinforcing or adapting actions depending on their outcome. However, the corresponding electrophysiological correlates in the basal ganglia and the extent to which these causally contribute to action adaptation in humans is unclear. Here, we recorded electrophysiological activity and applied bursts of electrical stimulation to the subthalamic nucleus, a core area of the basal ganglia, in 16 patients with Parkinson's disease (PD) on medication using temporarily externalized deep brain stimulation (DBS) electrodes. Patients as well as 16 age- and gender-matched healthy participants attempted to produce forces as close as possible to a target force to collect a maximum number of points. The target force changed over trials without being explicitly shown on the screen so that participants had to infer target force based on the feedback they received after each movement. Patients and healthy participants were able to adapt their force according to the feedback they received (P < 0.001). At the neural level, decreases in subthalamic beta (13 to 30 Hz) activity reflected poorer outcomes and stronger action adaptation in 2 distinct time windows (Pcluster-corrected < 0.05). Stimulation of the subthalamic nucleus reduced beta activity and led to stronger action adaptation if applied within the time windows when subthalamic activity reflected action outcomes and adaptation (Pcluster-corrected < 0.05). The more the stimulation volume was connected to motor cortex, the stronger was this behavioral effect (Pcorrected = 0.037). These results suggest that dynamic modulation of the subthalamic nucleus and interconnected cortical areas facilitates adaptive behavior.