Feathers are arranged in a precise pattern in avian skin. They first arise during development in a row along the dorsal midline, with rows of new feather buds added sequentially in a spreading wave. ...We show that the patterning of feathers relies on coupled fibroblast growth factor (FGF) and bone morphogenetic protein (BMP) signalling together with mesenchymal cell movement, acting in a coordinated reaction-diffusion-taxis system. This periodic patterning system is partly mechanochemical, with mechanical-chemical integration occurring through a positive feedback loop centred on FGF20, which induces cell aggregation, mechanically compressing the epidermis to rapidly intensify FGF20 expression. The travelling wave of feather formation is imposed by expanding expression of Ectodysplasin A (EDA), which initiates the expression of FGF20. The EDA wave spreads across a mesenchymal cell density gradient, triggering pattern formation by lowering the threshold of mesenchymal cells required to begin to form a feather bud. These waves, and the precise arrangement of feather primordia, are lost in the flightless emu and ostrich, though via different developmental routes. The ostrich retains the tract arrangement characteristic of birds in general but lays down feather primordia without a wave, akin to the process of hair follicle formation in mammalian embryos. The embryonic emu skin lacks sufficient cells to enact feather formation, causing failure of tract formation, and instead the entire skin gains feather primordia through a later process. This work shows that a reaction-diffusion-taxis system, integrated with mechanical processes, generates the feather array. In flighted birds, the key role of the EDA/Ectodysplasin A receptor (EDAR) pathway in vertebrate skin patterning has been recast to activate this process in a quasi-1-dimensional manner, imposing highly ordered pattern formation.
Cells are exposed to thousands of DNA damage events on a daily basis. This damage must be repaired to preserve genetic information and prevent development of disease. The most deleterious damage is a ...double-strand break (DSB), which is detected and repaired by mechanisms known as non-homologous end-joining (NHEJ) and homologous recombination (HR), which are components of the DNA damage response system. NHEJ is an error-prone first line of defense, whereas HR invokes error-free repair and is the focus of this review. The functions of the protein components of HR-driven DNA repair are regulated by the coordinated action of post-translational modifications including lysine acetylation, phosphorylation, ubiquitination, and SUMOylation. The latter two mechanisms are fundamental for recognition of DSBs and reorganizing chromatin to facilitate repair. We focus on the structures and molecular mechanisms for the protein components underlying synthesis, recognition, and cleavage of K63-linked ubiquitin chains, which are abundant at damage sites and obligatory for DSB repair. The forward flux of the K63-linked ubiquitination cascade is driven by the combined activity of E1 enzyme, the heterodimeric E2 Mms2-Ubc13, and its cognate E3 ligases RNF8 and RNF168, which is balanced through the binding and cleavage of chains by the deubiquitinase BRCC36, and the proteasome, and through the binding of chains by recognition modules on repair proteins such as RAP80. We highlight a number of aspects regarding our current understanding for the role of kinetics and dynamics in determining the function of the enzymes and chain recognition modules that drive K63 ubiquitination.
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•NHEJ or HR repairs DNA DSBs.•HR repair is error-free and regulated by ubiquitination and SUMOylation.•K63 ubiquitin chains are abundant at repair sites and obligatory for repair.•The flux of the chain-building cascade reshapes the chromatin landscape and facilitates repair.•The role of kinetics and dynamics in driving the flux of the chain-building machinery is emerging.
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a key cellular enzyme, with major roles in both glycolysis, and ‘moonlighting’ activities in the nucleus (uracil DNA glycosylase activity, nuclear ...protein nitrosylation), as a regulator of mRNA stability, a transferrin receptor, and as an antimicrobial agent. These activities are dependent, at least in part, on the integrity of an active site Cys residue, and a second neighboring Cys. These residues are differentially sensitive to oxidation, and determine both its catalytic activity and the redox signaling capacity of the protein. Such Cys modification is critical to cellular adaptation to oxidative environments by re-routing metabolic pathways to favor NADPH generation and antioxidant defenses. Despite the susceptibility of GAPDH to oxidation, it remains a puzzle as to how this enzyme acts as a redox signaling hub for oxidants such as hydrogen peroxide (H2O2) in the presence of high concentrations of specialized high-efficiency peroxide-removing enzymes. One possibility is that crowded environments, such as the cell cytosol, alter the oxidation pathways of GAPDH. In this study, we investigated the role of crowding (induced by dextran) on H2O2- and SIN-1-induced GAPDH oxidation, with data for crowded and dilute conditions compared. LC-MS/MS data revealed a lower extent of modification of the catalytic Cys under crowded conditions (i.e. less monomer units modified), but enhanced formation of the sulfonic acid resulting from hyper-oxidation. This effect was not observed with SIN-1. These data indicate that molecular crowding can modulate the oxidation pathways of GAPDH and its extent of oxidation and inactivation.
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•Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) has an oxidant-sensitive cysteine (Cys) in its active site.•Effects of macromolecular crowding on GAPDH oxidation by H2O2 and SIN-1 was investigated.•Dextran differentially affects H2O2- and SIN-1-mediated GAPDH cross-linking.•Higher yields of active site Cys-derived sulfonic acids were detected with crowding.
Monoclonal antibodies are one of the fastest growing classes of pharmaceutical products, however, their potential is limited by the high cost of development and manufacturing. Here we present a safe ...and cost-effective platform for in vivo expression of therapeutic antibodies using nucleoside-modified mRNA. To demonstrate feasibility and protective efficacy, nucleoside-modified mRNAs encoding the light and heavy chains of the broadly neutralizing anti-HIV-1 antibody VRC01 are generated and encapsulated into lipid nanoparticles. Systemic administration of 1.4 mg kg
of mRNA into mice results in ∼170 μg ml
VRC01 antibody concentrations in the plasma 24 h post injection. Weekly injections of 1 mg kg
of mRNA into immunodeficient mice maintain trough VRC01 levels above 40 μg ml
. Most importantly, the translated antibody from a single injection of VRC01 mRNA protects humanized mice from intravenous HIV-1 challenge, demonstrating that nucleoside-modified mRNA represents a viable delivery platform for passive immunotherapy against HIV-1 with expansion to a variety of diseases.
Summary Background Prognosis for women with abdominal aortic aneurysm might be worse than the prognosis for men. We aimed to systematically quantify the differences in outcomes between men and women ...being assessed for repair of intact abdominal aortic aneurysm using data from study periods after the year 2000. Methods In these systematic reviews and meta-analysis, we identified studies (randomised, cohort, or cross-sectional) by searching MEDLINE, Embase, CENTRAL, and grey literature published between Jan 1, 2005, and Sept 2, 2016, for two systematic reviews and Jan 1, 2009, and Sept 2, 2016, for one systematic review. Studies were included if they were of both men and women, with data presented for each sex separately, with abdominal aortic aneurysms being assessed for aneurysm repair by either endovascular repair (EVAR) or open repair. We conducted three reviews based on whether studies reported the proportion morphologically suitable (within manufacturers' instructions for use) for EVAR (EVAR suitability review), non-intervention rates (non-intervention review), and 30-day mortality (operative mortality review) after intact aneurysm repair. Studies had to include at least 20 women (for the EVAR suitability review), 20 women (for the non-intervention review), and 50 women (for the operative mortality review). Studies were excluded if they were review articles, editorials, letters, or case reports. For the operative review, studies were also excluded if they only provided hazard ratios or only reported in-hospital mortality. We assessed the quality of the studies using the Newcastle–Ottawa scoring system, and contacted authors for the provision of additional data if needed. We combined results across studies by random-effects meta-analysis. This study is registered with PROSPERO, number CRD42016043227. Findings Five studies assessed the morphological eligibility for EVAR (1507 men, 400 women). The overall pooled proportion of women eligible (34%) for EVAR was lower than it was in men (54%; odds ratio OR 0·44, 95% CI 0·32–0·62). Four single-centre studies reported non-intervention rates (1365 men, 247 women). The overall pooled non-intervention rates were higher in women (34%) than men (19%; OR 2·27, 95% CI 1·21–4·23). The review of 30-day mortality included nine studies (52 018 men, 11 076 women). The overall pooled estimate for EVAR was higher in women (2·3%) than in men (1·4%; OR 1·67, 95% CI 1·38–2·04). The overall estimate for open repair also was higher in women (5·4%) than in men (2·8%; OR 1·76, 95% CI 1·35–2·30). Interpretation Compared with men, a smaller proportion of women are eligible for EVAR, a higher proportion of women are not offered intervention, and operative mortality is much higher in women for both EVAR and open repair. The management of abdominal aortic aneurysm in women needs improvement. Funding National Institute for Health Research (UK).
Precise self-renewal of the germ cell lineage is fundamental to fertility and reproductive success. The early precursors for the germ lineage, primordial germ cells (PGCs), survive and proliferate in ...several embryonic locations during their migration to the embryonic gonad. By elucidating the active signaling pathways in migratory PGCs in vivo, we were able to create culture conditions that recapitulate this embryonic germ cell environment. In defined medium conditions without feeder cells, the growth factors FGF2, insulin, and Activin A, signaling through their cognate-signaling pathways, were sufficient for self-renewal of germline-competent PGCs. Forced expression of constitutively active MEK1, AKT, and SMAD3 proteins could replace their respective upstream growth factors. Unexpectedly, we found that BMP4 could replace Activin A in non-clonal growth conditions. These defined medium conditions identify the key molecular pathways required for PGC self-renewal and will facilitate efforts in biobanking of chicken genetic resources and genome editing.
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•Avian primordial germ cell self-renewal is dependent on FGF2, insulin, and Activin A molecules•BMP4 can replace Activin A in non-clonal growth conditions•Defined culture medium conditions will facilitate studies of germ cell self-renewal in other vertebrate species
In this article, McGrew and colleagues define the minimal growth factors and signaling pathways needed for the self-renewal of chicken primordial germ cells. They show that self-renewal is dependent on FGF2, insulin, and Activin A growth factors. BMP4 can replace Activin A in non-clonal growth conditions. These defined conditions identify the key molecular pathways required for PGC self-renewal.
Intracellular Tau inclusions are a pathological hallmark of several neurodegenerative diseases, collectively known as the tauopathies. They include Alzheimer disease, tangle-only dementia, Pick ...disease, argyrophilic grain disease, chronic traumatic encephalopathy, progressive supranuclear palsy, and corticobasal degeneration. Tau pathology appears to spread through intercellular propagation, requiring the formation of assembled “prion-like” species. Several cell and animal models have been described that recapitulate aspects of this phenomenon. However, the molecular characteristics of seed-competent Tau remain unclear. Here, we have used a cell model to understand the relationships between Tau structure/phosphorylation and seeding by aggregated Tau species from the brains of mice transgenic for human mutant P301S Tau and full-length aggregated recombinant P301S Tau. Deletion of motifs 275VQIINK280 and 306VQIVYK311 abolished the seeding activity of recombinant full-length Tau, suggesting that its aggregation was necessary for seeding. We describe conformational differences between native and synthetic Tau aggregates that may account for the higher seeding activity of native assembled Tau. When added to aggregated Tau seeds from the brains of mice transgenic for P301S Tau, soluble recombinant Tau aggregated and acquired the molecular properties of aggregated Tau from transgenic mouse brain. We show that seeding is conferred by aggregated Tau that enters cells through macropinocytosis and seeds the assembly of endogenous Tau into filaments.
Characteristics of seed-competent Tau are unknown.
Native Tau aggregates have a higher seeding potency than recombinant Tau aggregates. Recombinant Tau acquires the conformation and potency of native Tau aggregates by seeded assembly.
Conformation determines the seeding potencies of Tau aggregates.
Understanding the properties of seed-competent Tau gives insight into disease mechanisms.
Summary
Structural color is poorly known in plants relative to animals. In fruits, only a handful of cases have been described, including in Viburnum tinus where the blue color results from a ...disordered multilayered reflector made of lipid droplets. Here, we examine the broader evolutionary context of fruit structural color across the genus Viburnum.
We obtained fresh and herbarium fruit material from 30 Viburnum species spanning the phylogeny and used transmission electron microscopy, optical simulations, and ancestral state reconstruction to identify the presence/absence of photonic structures in each species, understand the mechanism producing structural color in newly identified species, relate the development of cell wall structure to reflectance in Viburnum dentatum, and describe the evolution of cell wall architecture across Viburnum.
We identify at least two (possibly three) origins of blue fruit color in Viburnum in species which produce large photonic structures made of lipid droplets embedded in the cell wall and which reflect blue light.
Examining the full spectrum of mechanisms producing color in pl, including structural color as well as pigments, will yield further insights into the diversity, ecology, and evolution of fruit color.
Avian primordial germ cells (PGCs) have significant potential to be used as a cell-based system for the study and preservation of avian germplasm, and the genetic modification of the avian genome. It ...was previously reported that PGCs from chicken embryos can be propagated in culture and contribute to the germ cell lineage of host birds.
We confirm these results by demonstrating that PGCs from a different layer breed of chickens can be propagated for extended periods in vitro. We demonstrate that intracellular signalling through PI3K and MEK is necessary for PGC growth. We carried out an initial characterisation of these cells. We find that cultured PGCs contain large lipid vacuoles, are glycogen rich, and express the stem cell marker, SSEA-1. These cells also express the germ cell-specific proteins CVH and CDH. Unexpectedly, using RT-PCR we show that cultured PGCs express the pluripotency genes c-Myc, cKlf4, cPouV, cSox2, and cNanog. Finally, we demonstrate that the cultured PGCs will migrate to and colonise the forming gonad of host embryos. Male PGCs will colonise the female gonad and enter meiosis, but are lost from the gonad during sexual development. In male hosts, cultured PGCs form functional gametes as demonstrated by the generation of viable offspring.
The establishment of in vitro cultures of germline competent avian PGCs offers a unique system for the study of early germ cell differentiation and also a comparative system for mammalian germ cell development. Primary PGC lines will form the basis of an alternative technique for the preservation of avian germplasm and will be a valuable tool for transgenic technology, with both research and industrial applications.
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