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Perception of pathogenic effectors in plants often relies on nucleotide-binding domain (NBS) and leucine-rich-repeat-containing (NLR) proteins. Some NLRs contain additional domains that function as integrated decoys for pathogen effector targets and activation of immune signalling. Wheat stripe rust is one of the most devastating diseases of crop plants. Here, we report the cloning of YrU1 , a stripe rust resistance gene from the diploid wheat Triticum urartu , the progenitor of the A genome of hexaploid wheat. YrU1 encodes a coiled-coil-NBS-leucine-rich repeat protein with N-terminal ankyrin-repeat and C-terminal WRKY domains, representing a unique NLR structure in plants. Database searches identify similar architecture only in wheat relatives. Transient expression of YrU1 in Nicotiana benthamiana does not induce cell death in the absence of pathogens. The ankyrin-repeat and coiled-coil domains of YrU1 self-associate, suggesting that homodimerisation is critical for YrU1 function. The identification and cloning of this disease resistance gene sheds light on NLR protein function and may facilitate breeding to control the devastating wheat stripe rust disease. Wheat stripe rust is a major disease of wheat caused by a fungal pathogen. Here the authors report the map-based cloning of YrU1 , a stripe rust resistance gene from Triticum urartu , a diploid progenitor of common wheat, and show it encodes a NLR protein with unusual domain architecture

Giant ankyrin-B (ankB) is a neurospecific alternatively spliced variant of ANK2, a high-confidence autism spectrum disorder (ASD) gene. We report that a mouse model for human ASD mutation of giant ankB exhibits increased axonal branching in cultured neurons with ectopic CNS axon connectivity, as well as with a transient increase in excitatory synapses during postnatal development. We elucidate a mechanism normally limiting axon branching, whereby giant ankB localizes to periodic axonal plasma membrane domains through L1 cell-adhesion molecule protein, where it couples microtubules to the plasma membrane and prevents microtubule entry into nascent axon branches. Giant ankB mutation or deficiency results in a dominantly inherited impairment in selected communicative and social behaviors combined with superior executive function. Thus, gain of axon branching due to giant ankB-deficiency/mutation is a candidate cellular mechanism to explain aberrant structural connectivity and penetrant behavioral consequences in mice as well as humans bearing ASD-related ANK2 mutations.

Despite being a major health concern, little is known about the pathophysiological changes that underly concussion. Nonetheless, emerging evidence suggests that selective damage to white matter axons, or diffuse axonal injury (DAI), disrupts brain network connectivity and function. While voltage-gated sodium channels (NaChs) and their anchoring proteins at the nodes of Ranvier (NOR) on axons are key elements of the brain’s network signaling machinery, changes in their integrity have not been studied in context with DAI. Here, we utilized a clinically relevant swine model of concussion that induces evolving axonal pathology, demonstrated by accumulation of amyloid precursor protein (APP) across the white matter. Over a two-week follow-up post-concussion with this model, we found widespread loss of NaCh isoform 1.6 (Nav1.6), progressive increases in NOR length, the appearance of void and heminodes and loss of βIV-spectrin, ankyrin G, and neurofascin 186 or their collective diffusion into the paranode. Notably, these changes were in close proximity, yet distinct from APP-immunoreactive swollen axonal profiles, potentially representing a unique, newfound phenotype of axonal pathology in DAI. Since concussion in humans is non-fatal, the clinical relevance of these findings was determined through examination of post-mortem brain tissue from humans with higher levels of acute traumatic brain injury. Here, a similar loss of Nav1.6 and changes in NOR structures in brain white matter were observed as found in the swine model of concussion. Collectively, this widespread and progressive disruption of NaChs and NOR appears to be a form of sodium channelopathy, which may represent an important substrate underlying brain network dysfunction after concussion.

Giant ankyrin-G (gAnkG) coordinates assembly of axon initial segments (AISs), which are sites of action potential generation located in proximal axons of most vertebrate neurons. Here, we identify a mechanism required for normal neural development in humans that ensures ordered recruitment of gAnkG and β4-spectrin to the AIS. We identified 3 human neurodevelopmental missense mutations located in the neurospecific domain of gAnkG that prevent recruitment of β4-spectrin, resulting in a lower density and more elongated pattern for gAnkG and its partners than in the mature AIS. We found that these mutations inhibit transition of gAnkG from a closed configuration with close apposition of N- and C-terminal domains to an extended state that is required for binding and recruitment of β4-spectrin, and normally occurs early in development of the AIS. We further found that the neurospecific domain is highly phosphorylated in mouse brain, and that phosphorylation at 2 sites (S1982 and S2619) is required for the conformational change and for recruitment of β4-spectrin. Together, these findings resolve a discrete intermediate stage in formation of the AIS that is regulated through phosphorylation of the neurospecific domain of gAnkG.

The stability and shape of the erythrocyte membrane is provided by the ankyrin-1 complex, but how it tethers the spectrin–actin cytoskeleton to the lipid bilayer and the nature of its association with the band 3 anion exchanger and the Rhesus glycoproteins remains unknown. Here we present structures of ankyrin-1 complexes purified from human erythrocytes. We reveal the architecture of a core complex of ankyrin-1, the Rhesus proteins RhAG and RhCE, the band 3 anion exchanger, protein 4.2, glycophorin A and glycophorin B. The distinct T-shaped conformation of membrane-bound ankyrin-1 facilitates recognition of RhCE and, unexpectedly, the water channel aquaporin-1. Together, our results uncover the molecular details of ankyrin-1 association with the erythrocyte membrane, and illustrate the mechanism of ankyrin-mediated membrane protein clustering. Cryo-EM structures of human erythrocyte ankyrin-1 complex offer insights into the architecture of the RBC membrane and show how ankyrins can simultaneously recruit different membrane proteins to enable functional organization of membrane transport processes.

The nodes of Ranvier have clustered Na+ and K+ channels necessary for rapid and efficient axonal action potential conduction. However, detailed mechanisms of channel clustering have only recently been identified: they include two independent axon–glia interactions that converge on distinct axonal cytoskeletons. Here, we discuss how glial cell adhesion molecules and the extracellular matrix molecules that bind them assemble combinations of ankyrins, spectrins and other cytoskeletal scaffolding proteins, which cluster ion channels. We present a detailed molecular model, incorporating these overlapping mechanisms, to explain how the nodes of Ranvier are assembled in both the peripheral and central nervous systems.The formation of the nodes of Ranvier in myelinated axons involves a specific clustering of ion channels. In this Review, Rasband and Peles describe two independent, glia-directed mechanisms that converge on the axonal cytoskeleton to cluster and maintain nodal ion channels.

Actin, spectrin, and associated molecules form a periodic sub-membrane lattice structure in axons. How this membrane skeleton is developed and why it preferentially forms in axons are unknown. Here, we studied the developmental mechanism of this lattice structure. We found that this structure emerged early during axon development and propagated from proximal regions to distal ends of axons. Components of the axon initial segment were recruited to the lattice late during development. Formation of the lattice was regulated by the local concentration of βII spectrin, which is higher in axons than in dendrites. Increasing the dendritic concentration of βII spectrin by overexpression or by knocking out ankyrin B induced the formation of the periodic structure in dendrites, demonstrating that the spectrin concentration is a key determinant in the preferential development of this structure in axons and that ankyrin B is critical for the polarized distribution of βII spectrin in neurites. The brain contains hundred types of neurons, but they are all variations on the same basic structure. Each neuron consists of a cell body that is covered in short protrusions called dendrites and a long thin structure called the axon. The dendrites receive incoming signals from neighboring neurons and they transmit these signals via the cell body to the axon, which in turn relays them to the dendrites of the next neuron (or neurons). Like all cells, neurons maintain their structure with the help of an internal cytoskeleton made up of many different proteins. However, it was discovered recently that axons have an additional lattice-like structure underneath their outer membrane. This structure, which consists of rings of actin filaments separated by molecules of a protein called spectrin, is preferentially formed in axons and is found much less frequently in dendrites. Now Zhong, He et al., who are members of the research group that discovered the axonal skeleton, have used ‘super-resolution imaging’ to figure out how this skeleton forms and why it predominantly forms in axons. In brief, a basic version of the sub-membrane periodic skeleton is laid down early in development, starting next to the cell body before gradually spreading down the axon. The skeleton then continues to mature throughout development with the incorporation of several additional types of proteins. The periodic skeleton only forms in regions which contain enough βII spectrin. Under normal conditions, dendrites contain too little βII spectrin to support the growth of such a periodic skeleton. However, artificially increasing the amount of βII spectrin present by overexpressing the corresponding gene, or by knocking out ankyrin B (a molecule that is important for establishing the preferential distribution of βII spectrin in axons), is sufficient to trigger periodic skeleton formation in dendrites. Given that axons and dendrites have distinct roles in neuronal signaling, this uneven distribution of spectrin is likely to be one way in which these regions maintain the specific structures that support their individual functions.

Poxviruses are large DNA viruses with an arsenal of immune-modulatory genes, many of which remain uncharacterized. Proteins with ankyrin repeats are distinct features of poxviruses, although the biological functions of ankyrin proteins are not fully understood. Lumpy skin disease virus (LSDV) encodes five proteins with ankyrin repeats. Here, we reveal the role of LSDV012, an ankyrin protein, in conferring resistance to type I interferon (IFN) in cells. Deletion of LSDV012 from LSDV significantly impacted viral replication in the presence of type I IFN, highlighting the importance of LSDV012 in antagonizing type I IFN responses. Further investigation revealed that LSDV012 interacted with interferon-induced proteins with tetratricopeptide repeats (IFITs), particularly IFIT1, altering its subcellular localization, interacting with its C-terminus and inhibiting its RNA-binding ability without inducing its degradation. Phylogenetic analysis demonstrated that LSDV012 orthologs are conserved in capripoxviruses and cervidpoxviruses , and exhibit host species-specific interactions with IFIT1. Notably, LSDV012 was able to rescue the degradation of IFIT1 mediated by VACV C9. These findings provide novel insights into the viral strategies employed by LSDV to subvert host antiviral defenses and underscore the evolutionary adaptations of poxvirus ankyrin proteins in host species-specific immune evasion.

Significance Excitable axonal membrane microdomains are unique features of vertebrate nervous systems that are required for normal neuronal signaling and are involved in human neurological disorders. Ankyrin-G is a critical adaptor protein that acquired a giant exon early in vertebrate evolution, resulting in a new nervous system-specific polypeptide that is a master organizer of axonal excitable membranes. Giant ankyrin-G–deficient mice live to weaning and provide a rationale for survival of humans with severe cognitive dysfunction bearing a truncating mutation in the giant exon. The giant exon of ankyrin-G thus was a transformative innovation in evolution of the vertebrate nervous system that now is a potential target in neurodevelopmental disorders. Axon initial segments (AISs) and nodes of Ranvier are sites of clustering of voltage-gated sodium channels (VGSCs) in nervous systems of jawed vertebrates that facilitate fast long-distance electrical signaling. We demonstrate that proximal axonal polarity as well as assembly of the AIS and normal morphogenesis of nodes of Ranvier all require a heretofore uncharacterized alternatively spliced giant exon of ankyrin-G (AnkG). This exon has sequence similarity to I-connectin/Titin and was acquired after the first round of whole-genome duplication by the ancestral ANK2/ANK3 gene in early vertebrates before development of myelin. The giant exon resulted in a new nervous system-specific 480-kDa polypeptide combining previously known features of ANK repeats and β-spectrin–binding activity with a fibrous domain nearly 150 nm in length. We elucidate previously undescribed functions for giant AnkG, including recruitment of β4 spectrin to the AIS that likely is regulated by phosphorylation, and demonstrate that 480-kDa AnkG is a major component of the AIS membrane “undercoat’ imaged by platinum replica electron microscopy. Surprisingly, giant AnkG-knockout neurons completely lacking known AIS components still retain distal axonal polarity and generate action potentials (APs), although with abnormal frequency. Giant AnkG-deficient mice live to weaning and provide a rationale for survival of humans with severe cognitive dysfunction bearing a truncating mutation in the giant exon. The giant exon of AnkG is required for assembly of the AIS and nodes of Ranvier and was a transformative innovation in evolution of the vertebrate nervous system that now is a potential target in neurodevelopmental disorders.

GABAergic circuits are critical for the synchronization and higher order function of brain networks. Defects in this circuitry are linked to neuropsychiatric diseases, including bipolar disorder, schizophrenia, and autism. Work in cultured neurons has shown that ankyrin-G plays a key role in the regulation of GABAergic synapses on the axon initial segment and somatodendritic domain of pyramidal neurons, where it interacts directly with the GABAA receptor-associated protein (GABARAP) to stabilize cell surface GABAA receptors. Here, we generated a knock-in mouse model expressing a mutation that abolishes the ankyrin-G/GABARAP interaction (Ank3 W1989R) to understand how ankyrin-G and GABARAP regulate GABAergic circuitry in vivo. We found that Ank3 W1989R mice exhibit a striking reduction in forebrain GABAergic synapses resulting in pyramidal cell hyperexcitability and disruptions in network synchronization. In addition, we identified changes in pyramidal cell dendritic spines and axon initial segments consistent with compensation for hyperexcitability. Finally, we identified the ANK3 W1989R variant in a family with bipolar disorder, suggesting a potential role of this variant in disease. Our results highlight the importance of ankyrin-G in regulating forebrain circuitry and provide novel insights into how ANK3 loss-of-function variants may contribute to human disease.