Nuclear-Import Receptors Reverse Aberrant Phase Transitions of RNA-Binding Proteins with Prion-like Domains.

RNA-binding proteins (RBPs) with prion-like domains (PrLDs) phase transition to functional liquids, which can mature into aberrant hydrogels composed of pathological fibrils that underpin fatal neurodegenerative disorders. Several nuclear RBPs with PrLDs, including TDP-43, FUS, hnRNPA1, and hnRNPA2, mislocalize to cytoplasmic inclusions in neurodegenerative disorders, and mutations in their PrLDs can accelerate fibrillization and cause disease. Here, we establish that nuclear-import receptors (NIRs) specifically chaperone and potently disaggregate wild-type and disease-linked RBPs bearing a NLS. Karyopherin-ß2 (also called Transportin-1) engages PY-NLSs to inhibit and reverse FUS, TAF15, EWSR1, hnRNPA1, and hnRNPA2 fibrillization, whereas Importin-α plus Karyopherin-ß1 prevent and reverse TDP-43 fibrillization. Remarkably, Karyopherin-ß2 dissolves phase-separated liquids and aberrant fibrillar hydrogels formed by FUS and hnRNPA1. In vivo, Karyopherin-ß2 prevents RBPs with PY-NLSs accumulating in stress granules, restores nuclear RBP localization and function, and rescues degeneration caused by disease-linked FUS and hnRNPA2. Thus, NIRs therapeutically restore RBP homeostasis and mitigate neurodegeneration.

Placental Evolution: Innovating how to Feed Babies.

The evolution of the placenta was transformative. It changed how offspring are fed during gestation from depositing all the resources into an egg to continually supplying resources throughout gestation. Placental evolution is infinitely complex, with many moving parts, but at the core it is driven by a conflict over resources between the mother and the baby, which sets up a Red Queen race, fueling rapid diversification of morphological, cellular, and genetic forms. Placentas from even closely related species are highly divergent in form and function, and many cellular processes are distinct. If we could extract the entirety of genomic information for placentas across all species, including the many hundreds that have evolved in fish and reptiles, we could find their shared commonality, and that would tell us which of the many pieces really matter. We do not have this information, but we do have clues. Convergent evolution mechanisms were repeatedly used in the placenta, including the intense selective pressure to co-opt an envelope protein to build a multinucleated syncytium, the use of the same hormones and structural proteins in placentas derived from separate embryonic origins that arose hundreds of millions of years apart, and the co-option of endogenous retroviruses to form capsids as a way of transport and as mutagens to form new enhancers. As a result, the placental genome is the Wild West of biology, set up to rapidly change, adapt, and innovate. This ability to adapt facilitated the evolution of big babies with big brains and will continue to support offspring and their mothers in our ever-changing global environment.

IL-17+ γδ T cells as kick-starters of inflammation.

Shortly after the discovery of interleukin 17 (IL-17)-producing CD4+ helper T cells (TH17 cells), it was found that γδ T cells can also secrete large amounts of this pro-inflammatory cytokine. A decade later, it is now known that IL-17+ γδ T cells (γδ17 T cells) are often the main providers of IL-17A in various models of inflammatory diseases, while they also contribute to protective immune responses to infectious organisms. Due to an intricate thymic program of differentiation, γδ17 T cells are able to respond faster than TH17 cells do and thus predominate in the early stages of inflammatory responses. Here we review the current knowledge of the development, activation and pathophysiological functions of γδ17 T cells, aiming to increase the awareness in the community of the therapeutic potential of this 'other side' of IL-17-mediated immune responses.

H3K4me3 breadth is linked to cell identity and transcriptional consistency.

Trimethylation of histone H3 at lysine 4 (H3K4me3) is a chromatin modification known to mark the transcription start sites of active genes. Here, we show that H3K4me3 domains that spread more broadly over genes in a given cell type preferentially mark genes that are essential for the identity and function of that cell type. Using the broadest H3K4me3 domains as a discovery tool in neural progenitor cells, we identify novel regulators of these cells. Machine learning models reveal that the broadest H3K4me3 domains represent a distinct entity, characterized by increased marks of elongation. The broadest H3K4me3 domains also have more paused polymerase at their promoters, suggesting a unique transcriptional output. Indeed, genes marked by the broadest H3K4me3 domains exhibit enhanced transcriptional consistency and [corrected] increased transcriptional levels, and perturbation of H3K4me3 breadth leads to changes in transcriptional consistency. Thus, H3K4me3 breadth contains information that could ensure transcriptional precision at key cell identity/function genes.

BTN3A3 evasion promotes the zoonotic potential of influenza A viruses.

Spillover events of avian influenza A viruses (IAVs) to humans could represent the first step in a future pandemic1. Several factors that limit the transmission and replication of avian IAVs in mammals have been identified. There are several gaps in our understanding to predict which virus lineages are more likely to cross the species barrier and cause disease in humans1. Here, we identified human BTN3A3 (butyrophilin subfamily 3 member A3)2 as a potent inhibitor of avian IAVs but not human IAVs. We determined that BTN3A3 is expressed in human airways and its antiviral activity evolved in primates. We show that BTN3A3 restriction acts primarily at the early stages of the virus life cycle by inhibiting avian IAV RNA replication. We identified residue 313 in the viral nucleoprotein (NP) as the genetic determinant of BTN3A3 sensitivity (313F or, rarely, 313L in avian viruses) or evasion (313Y or 313V in human viruses). However, avian IAV serotypes, such as H7 and H9, that spilled over into humans also evade BTN3A3 restriction. In these cases, BTN3A3 evasion is due to substitutions (N, H or Q) in NP residue 52 that is adjacent to residue 313 in the NP structure3. Thus, sensitivity or resistance to BTN3A3 is another factor to consider in the risk assessment of the zoonotic potential of avian influenza viruses.

TCR signal strength controls thymic differentiation of discrete proinflammatory γδ T cell subsets.

The mouse thymus produces discrete γδ T cell subsets that make either interferon-γ (IFN-γ) or interleukin 17 (IL-17), but the role of the T cell antigen receptor (TCR) in this developmental process remains controversial. Here we show that Cd3g(+/-) Cd3d(+/-) (CD3 double-haploinsufficient (CD3DH)) mice have reduced TCR expression and signaling strength on γδ T cells. CD3DH mice had normal numbers and phenotypes of αß thymocyte subsets, but impaired differentiation of fetal Vγ6(+) (but not Vγ4(+)) IL-17-producing γδ T cells and a marked depletion of IFN-γ-producing CD122(+) NK1.1(+) γδ T cells throughout ontogeny. Adult CD3DH mice showed reduced peripheral IFN-γ(+) γδ T cells and were resistant to experimental cerebral malaria. Thus, TCR signal strength within specific thymic developmental windows is a major determinant of the generation of proinflammatory γδ T cell subsets and their impact on pathophysiology.

The Role of Interferon-γ in Autoimmune Polyendocrine Syndrome Type 1.

BACKGROUND: Autoimmune polyendocrine syndrome type 1 (APS-1) is a life-threatening, autosomal recessive syndrome caused by autoimmune regulator (AIRE) deficiency. In APS-1, self-reactive T cells escape thymic negative selection, infiltrate organs, and drive autoimmune injury. The effector mechanisms governing T-cell-mediated damage in APS-1 remain poorly understood. METHODS: We examined whether APS-1 could be classified as a disease mediated by interferon-γ. We first assessed patients with APS-1 who were participating in a prospective natural history study and evaluated mRNA and protein expression in blood and tissues. We then examined the pathogenic role of interferon-γ using Aire-/-Ifng-/- mice and Aire-/- mice treated with the Janus kinase (JAK) inhibitor ruxolitinib. On the basis of our findings, we used ruxolitinib to treat five patients with APS-1 and assessed clinical, immunologic, histologic, transcriptional, and autoantibody responses. RESULTS: Patients with APS-1 had enhanced interferon-γ responses in blood and in all examined autoimmunity-affected tissues. Aire-/- mice had selectively increased interferon-γ production by T cells and enhanced interferon-γ, phosphorylated signal transducer and activator of transcription 1 (pSTAT1), and CXCL9 signals in multiple organs. Ifng ablation or ruxolitinib-induced JAK-STAT blockade in Aire-/- mice normalized interferon-γ responses and averted T-cell infiltration and damage in organs. Ruxolitinib treatment of five patients with APS-1 led to decreased levels of T-cell-derived interferon-γ, normalized interferon-γ and CXCL9 levels, and remission of alopecia, oral candidiasis, nail dystrophy, gastritis, enteritis, arthritis, Sjögren's-like syndrome, urticaria, and thyroiditis. No serious adverse effects from ruxolitinib were identified in these patients. CONCLUSIONS: Our findings indicate that APS-1, which is caused by AIRE deficiency, is characterized by excessive, multiorgan interferon-γ-mediated responses. JAK inhibition with ruxolitinib in five patients showed promising results. (Funded by the National Institute of Allergy and Infectious Diseases and others.).

Circadian autophagy drives iTRF-mediated longevity.

Time-restricted feeding (TRF) has recently gained interest as a potential anti-ageing treatment for organisms from Drosophila to humans1-5. TRF restricts food intake to specific hours of the day. Because TRF controls the timing of feeding, rather than nutrient or caloric content, TRF has been hypothesized to depend on circadian-regulated functions; the underlying molecular mechanisms of its effects remain unclear. Here, to exploit the genetic tools and well-characterized ageing markers of Drosophila, we developed an intermittent TRF (iTRF) dietary regimen that robustly extended fly lifespan and delayed the onset of ageing markers in the muscles and gut. We found that iTRF enhanced circadian-regulated transcription and that iTRF-mediated lifespan extension required both circadian regulation and autophagy, a conserved longevity pathway. Night-specific induction of autophagy was both necessary and sufficient to extend lifespan on an ad libitum diet and also prevented further iTRF-mediated lifespan extension. By contrast, day-specific induction of autophagy did not extend lifespan. Thus, these results identify circadian-regulated autophagy as a critical contributor to iTRF-mediated health benefits in Drosophila. Because both circadian regulation and autophagy are highly conserved processes in human ageing, this work highlights the possibility that behavioural or pharmaceutical interventions that stimulate circadian-regulated autophagy might provide people with similar health benefits, such as delayed ageing and lifespan extension.

Global chemical effects of the microbiome include new bile-acid conjugations.

A mosaic of cross-phylum chemical interactions occurs between all metazoans and their microbiomes. A number of molecular families that are known to be produced by the microbiome have a marked effect on the balance between health and disease1-9. Considering the diversity of the human microbiome (which numbers over 40,000 operational taxonomic units10), the effect of the microbiome on the chemistry of an entire animal remains underexplored. Here we use mass spectrometry informatics and data visualization approaches11-13 to provide an assessment of the effects of the microbiome on the chemistry of an entire mammal by comparing metabolomics data from germ-free and specific-pathogen-free mice. We found that the microbiota affects the chemistry of all organs. This included the amino acid conjugations of host bile acids that were used to produce phenylalanocholic acid, tyrosocholic acid and leucocholic acid, which have not previously been characterized despite extensive research on bile-acid chemistry14. These bile-acid conjugates were also found in humans, and were enriched in patients with inflammatory bowel disease or cystic fibrosis. These compounds agonized the farnesoid X receptor in vitro, and mice gavaged with the compounds showed reduced expression of bile-acid synthesis genes in vivo. Further studies are required to confirm whether these compounds have a physiological role in the host, and whether they contribute to gut diseases that are associated with microbiome dysbiosis.

The multisensory regulation of unconventional T cell homeostasis.

Unconventional T cells typically group γδ T cells, invariant Natural Killer T cells (NKT) and Mucosal Associated Invariant T (MAIT) cells. With their pre-activated status and biased tropism for non-lymphoid organs, they provide a rapid (innate-like) and efficient first line of defense against pathogens at strategical barrier sites, while they can also trigger chronic inflammation, and unexpectedly contribute to steady state physiology. Thus, a tight control of their homeostasis is critical to maintain tissue integrity. In this review, we discuss the recent advances of our understanding of the factors, from neuroimmune to inflammatory regulators, shaping the size and functional properties of unconventional T cell subsets in non-lymphoid organs. We present a general overview of the mechanisms common to these populations, while also acknowledging specific aspects of their diversity. We mainly focus on their maintenance at steady state and upon inflammation, highlighting some key unresolved issues and raising upcoming technical, fundamental and translational challenges.

Metabotropic NMDAR Signaling Contributes to Sex Differences in Synaptic Plasticity and Episodic Memory.

NMDA receptor (NMDAR) - mediated calcium influx triggers the induction and initial expression of Long-Term Potentiation (LTP). Here we report that in male rodents ion flux-independent (metabotropic) NMDAR signaling is critical for a third step in the production of enduring LTP, i.e., cytoskeletal changes that stabilize the activity-induced synaptic modifications. Surprisingly, females rely upon estrogen receptor alpha (ERα) for the metabotropic NMDAR operations used by males. Blocking NMDAR channels with MK-801 eliminated LTP expression in hippocampal field CA1 of both sexes but left intact theta burst stimulation (TBS)-induced actin polymerization within dendritic spines. A selective antagonist (Ro25-6981) of the NMDAR GluN2B subunit had minimal effects on synaptic responses but blocked actin polymerization and LTP consolidation in males only. Conversely, an ERα antagonist thoroughly disrupted TBS-induced actin polymerization and LTP in females while having no evident effect in males. In an episodic memory paradigm, Ro25-6981 prevented acquisition of spatial locations by males but not females whereas an ERα antagonist blocked acquisition in females but not males. Sex differences in LTP consolidation were accompanied by pronounced differences in episodic memory in tasks involving a minimal (for learning) cue-sampling. Males did better on acquisition of spatial information whereas females had much higher scores than males on tests for acquisition of the identity of cues (episodic 'what') and the order in which the cues were sampled (episodic 'when'). We propose that sex differences in synaptic processes used to stabilize LTP result in differential encoding of the basic elements of episodic memory.Significance Statement Calcium influx through NMDARs has long been recognized as the initiating event for LTP. Results of the present studies call for a substantial revision to this fundamental observation about learning-related synaptic plasticity. Specifically, we show cytoskeletal mechanisms that consolidate field CA1 LTP and episodic memory are triggered not by NMDAR-mediated calcium but by ion flux-independent (metabotropic) signaling. Males used metabotropic functions of the NMDARs for this purpose whereas females relied upon synaptic estrogen receptors. This unprecedented instance of sex differences in synaptic function was accompanied by surprisingly large male/female differences in the acquisition of the three basic elements of episodic memory.

The Joubert-Meckel-Nephronophthisis Spectrum of Ciliopathies.

The Joubert syndrome (JS), Meckel syndrome (MKS), and nephronophthisis (NPH) ciliopathy spectrum could be the poster child for advances and challenges in Mendelian human genetics over the past half century. Progress in understanding these conditions illustrates many core concepts of human genetics. The JS phenotype alone is caused by pathogenic variants in more than 40 genes; remarkably, all of the associated proteins function in and around the primary cilium. Primary cilia are near-ubiquitous, microtubule-based organelles that play crucial roles in development and homeostasis. Protruding from the cell, these cellular antennae sense diverse signals and mediate Hedgehog and other critical signaling pathways. Ciliary dysfunction causes many human conditions termed ciliopathies, which range from multiple congenital malformations to adult-onset single-organ failure. Research on the genetics of the JS-MKS-NPH spectrum has spurred extensive functional work exploring the broadly important role of primary cilia in health and disease. This functional work promises to illuminate the mechanisms underlying JS-MKS-NPH in humans, identify therapeutic targets across genetic causes, and generate future precision treatments.

Clotting factor genes are associated with preeclampsia in high-altitude pregnant women in the Peruvian Andes.

Preeclampsia is a multi-organ complication of pregnancy characterized by sudden hypertension and proteinuria that is among the leading causes of preterm delivery and maternal morbidity and mortality worldwide. The heterogeneity of preeclampsia poses a challenge for understanding its etiology and molecular basis. Intriguingly, risk for the condition increases in high-altitude regions such as the Peruvian Andes. To investigate the genetic basis of preeclampsia in a population living at high altitude, we characterized genome-wide variation in a cohort of preeclamptic and healthy Andean families (n = 883) from Puno, Peru, a city located above 3,800 meters of altitude. Our study collected genomic DNA and medical records from case-control trios and duos in local hospital settings. We generated genotype data for 439,314 SNPs, determined global ancestry patterns, and mapped associations between genetic variants and preeclampsia phenotypes. A transmission disequilibrium test (TDT) revealed variants near genes of biological importance for placental and blood vessel function. The top candidate region was found on chromosome 13 of the fetal genome and contains clotting factor genes PROZ, F7, and F10. These findings provide supporting evidence that common genetic variants within coagulation genes play an important role in preeclampsia. A selection scan revealed a potential adaptive signal around the ADAM12 locus on chromosome 10, implicated in pregnancy disorders. Our discovery of an association in a functional pathway relevant to pregnancy physiology in an understudied population of Native American origin demonstrates the increased power of family-based study design and underscores the importance of conducting genetic research in diverse populations.

Key actors in neuropathophysiology: The role of γδ T cells.

The neuroimmune axis has been the focus of many studies, with special emphasis on the interactions between the central nervous system and the different immune cell subsets. T cells are namely recognized to play a critical role due to their interaction with nerves, by secreting cytokines and neurotrophins, which regulate the development, function, and survival of neurons. In this context, γδ T cells are particularly relevant, as they colonize specific tissues, namely the meninges, and have a wide variety of complex functions that balance physiological systems. Notably, γδ T cells are not only key components for maintaining brain homeostasis but are also responsible for triggering or preventing inflammatory responses in various pathologies, including neurodegenerative diseases as well as neuropsychiatric and developmental disorders. Here, we provide an overview of the current state of the art on the contribution of γδ T cells in neuropathophysiology and delve into the molecular mechanisms behind it. We aim to shed light on γδ T cell functions in the central nervous system while highlighting upcoming challenges in the field and providing new clues for potential therapeutic strategies.

The effect of Huntington's disease on cognitive and physical motivation.

Apathy is one of the most common neuropsychiatric features of Huntington's disease. A hallmark of apathy is diminished goal-directed behaviour, which is characterized by a lower motivation to engage in cognitively or physically effortful actions. However, it remains unclear whether this reduction in goal-directed behaviour is driven primarily by a motivational deficit and/or is secondary to the progressive cognitive and physical deficits that accompany more advanced disease. We addressed this question by testing 17 individuals with manifest Huntington's disease and 22 age-matched controls on an effort-based decision-making paradigm. Participants were first trained on separate cognitively and physically effortful tasks and provided explicit feedback about their performance. Next, they chose on separate trials how much effort they were willing to exert in each domain in return for varying reward. At the conclusion of the experiment, participants were asked to rate their subjective perception of task load. In the cognitive task, the Huntington's disease group were more averse to cognitive effort than controls. Although the Huntington's disease group were more impaired than controls on the task itself, their greater aversion to cognitive effort persisted even after controlling for task performance. This suggests that the lower levels of cognitive motivation in the Huntington's disease group relative to controls was most likely driven by a primary motivational deficit. In contrast, both groups expressed a similar preference for physical effort. Importantly, the similar levels of physical motivation across both groups occurred even though participants with Huntington's disease performed objectively worse than controls on the physical effort task, and were aware of their performance through explicit feedback on each trial. This indicates that the seemingly preserved level of physical motivation in Huntington's disease was driven by a willingness to engage in physically effortful actions despite a reduced capacity to do so. Finally, the Huntington's disease group provided higher ratings of subjective task demand than controls for the cognitive (but not physical) effort task and when assessing the mental (but not the physical) load of each task. Together, these results revealed a dissociation in cognitive and physical motivation deficits between Huntington's disease and controls, which were accompanied by differences in how effort was subjectively perceived by the two groups. This highlights that motivation is the final manifestation of a complex set of mechanisms involved in effort processing, which are separable across different domains of behaviour. These findings have important clinical implications for the day-to-day management of apathy in Huntington's disease.

Innovative, combinatorial and high-throughput approaches to degrader synthesis.

Targeted protein degraders such as PROTACs and molecular glues are a rapidly emerging therapeutic modality within industry and academia. Degraders possess unique mechanisms of action that lead to the removal of specific proteins by co-opting the cell's natural degradation mechanisms via induced proximity. Their optimisation thus far has often been largely empirical, requiring the synthesis and screening of a large number of analogues. In addition, the synthesis and development of degraders is often challenging, leading to lengthy optimisation campaigns to deliver candidate-quality compounds. This review highlights how the synthesis of degraders has evolved in recent years, in particular focusing on means of applying high-throughput chemistry and screening approaches to expedite these timelines, which we anticipate to be valuable in shaping the future of degrader optimisation campaigns.

Generative ß-hairpin design using a residue-based physicochemical property landscape.

De novo peptide design is a new frontier that has broad application potential in the biological and biomedical fields. Most existing models for de novo peptide design are largely based on sequence homology that can be restricted based on evolutionarily derived protein sequences and lack the physicochemical context essential in protein folding. Generative machine learning for de novo peptide design is a promising way to synthesize theoretical data that are based on, but unique from, the observable universe. In this study, we created and tested a custom peptide generative adversarial network intended to design peptide sequences that can fold into the ß-hairpin secondary structure. This deep neural network model is designed to establish a preliminary foundation of the generative approach based on physicochemical and conformational properties of 20 canonical amino acids, for example, hydrophobicity and residue volume, using extant structure-specific sequence data from the PDB. The beta generative adversarial network model robustly distinguishes secondary structures of ß hairpin from α helix and intrinsically disordered peptides with an accuracy of up to 96% and generates artificial ß-hairpin peptide sequences with minimum sequence identities around 31% and 50% when compared against the current NCBI PDB and nonredundant databases, respectively. These results highlight the potential of generative models specifically anchored by physicochemical and conformational property features of amino acids to expand the sequence-to-structure landscape of proteins beyond evolutionary limits.

MHC-Fine: Fine-tuned AlphaFold for precise MHC-peptide complex prediction.

The precise prediction of major histocompatibility complex (MHC)-peptide complex structures is pivotal for understanding cellular immune responses and advancing vaccine design. In this study, we enhanced AlphaFold's capabilities by fine-tuning it with a specialized dataset consisting of exclusively high-resolution class I MHC-peptide crystal structures. This tailored approach aimed to address the generalist nature of AlphaFold's original training, which, while broad-ranging, lacked the granularity necessary for the high-precision demands of class I MHC-peptide interaction prediction. A comparative analysis was conducted against the homology-modeling-based method Pandora as well as the AlphaFold multimer model. Our results demonstrate that our fine-tuned model outperforms others in terms of root-mean-square deviation (median value for Cα atoms for peptides is 0.66 Å) and also provides enhanced predicted local distance difference test scores, offering a more reliable assessment of the predicted structures. These advances have substantial implications for computational immunology, potentially accelerating the development of novel therapeutics and vaccines by providing a more precise computational lens through which to view MHC-peptide interactions.