A Phylogenomic Perspective on the Radiation of Ray-Finned Fishes Based upon Targeted Sequencing of Ultraconserved Elements (UCEs)

Ray-finned fishes constitute the dominant radiation of vertebrates with over 32,000 species. Although molecular phylogenetics has begun to disentangle major evolutionary relationships within this vast section of the Tree of Life, there is no widely available approach for efficiently collecting phylogenomic data within fishes, leaving much of the enormous potential of massively parallel sequencing technologies for resolving major radiations in ray-finned fishes unrealized. Here, we provide a genomic perspective on longstanding questions regarding the diversification of major groups of ray-finned fishes through targeted enrichment of ultraconserved nuclear DNA elements (UCEs) and their flanking sequence. Our workflow efficiently and economically generates data sets that are orders of magnitude larger than those produced by traditional approaches and is well-suited to working with museum specimens. Analysis of the UCE data set recovers a well-supported phylogeny at both shallow and deep time-scales that supports a monophyletic relationship between Amia and Lepisosteus (Holostei) and reveals elopomorphs and then osteoglossomorphs to be the earliest diverging teleost lineages. Our approach additionally reveals that sequence capture of UCE regions and their flanking sequence offers enormous potential for resolving phylogenetic relationships within ray-finned fishes.     

Generation and preclinical characterization of an antibody specific for SEMA4D

Semaphorin 4D (SEMA4D or CD100) is a member of the semaphorin family of proteins and an important mediator of the movement and differentiation of multiple cell types, including those of the immune, vascular, and nervous systems. Blocking the binding of SEMA4D to its receptors can result in physiologic changes that may have implications in cancer, autoimmune, and neurological disease. To study the effects of blocking SEMA4D, we generated, in SEMA4D-deficient mice, a panel of SEMA4D-specific hybridomas that react with murine, primate, and human SEMA4D. Utilizing the complementarity-determining regions from one of these hybridomas (mAb 67-2), we generated VX15/2503, a humanized IgG4 monoclonal antibody that is currently in clinical development for the potential treatment of various malignancies and neurodegenerative disorders, including multiple sclerosis and Huntington's disease. This work describes the generation and characterization of VX15/2503, including in vitro functional testing, epitope mapping, and an in vivo demonstration of efficacy in an animal model of rheumatoid arthritis.     

Influenza-Specific Antibody-Dependent Phagocytosis

BackgroundImmunity to human influenza A virus (IAV) infection is only partially understood. Broadly non-neutralizing antibodies may assist in reducing disease but have not been well characterized.MethodsWe measured internalization of opsonized, influenza protein-coated fluorescent beads and live IAV into a monocytic cell line to study antibody-dependent phagocytosis (ADP) against multiple influenza hemagglutinin (HA) subtypes. We analyzed influenza HA-specific ADP in healthy human donors, in preparations of intravenous immunoglobulin (IVIG), and following IAV infection of humans and macaques.ResultsWe found that both sera from healthy adults and IVIG preparations had broad ADP to multiple seasonal HA proteins and weak cross-reactive ADP to non-circulating HA proteins. The ADP in experimentally influenza-infected macaque plasma and naturally influenza-infected human sera mediated phagocytosis of both homologous and heterologous IAVs. Further, the IAV phagocytosed in an antibody-mediated manner had reduced infectivity in vitro.ConclusionWe conclude that IAV infections in humans and macaques leads to the development of influenza-specific ADP that can clear IAV infection in vitro. Repeated exposure of humans to multiple IAV infections likely leads to the development of ADP that is cross-reactive to strains not previously encountered. Further analyses of the protective capacity of broadly reactive influenza-specific ADP is warranted.     

Chemically defined media modifications to lower tryptophan oxidation of biopharmaceuticals

Oxidation of biopharmaceuticals is a major product quality issue with potential impacts on activity and immunogenicity. At Eli Lilly and Company, high tryptophan oxidation was observed for two biopharmaceuticals in development produced in Chinese hamster ovary cells. A switch from historical hydrolysate‐containing media to chemically defined media with a reformulated basal powder was thought to be responsible, so mitigation efforts focused on media modification. Shake flask studies identified that increasing tryptophan, copper, and manganese and decreasing cysteine concentrations were individual approaches to lower tryptophan oxidation. When amino acid and metal changes were combined, the modified formulation had a synergistic impact that led to substantially less tryptophan oxidation for both biopharmaceuticals. Similar results were achieved in shake flasks and benchtop bioreactors, demonstrating the potential to implement these modifications at manufacturing scale. The modified formulation did not negatively impact cell growth and viability, product titer, purity, charge variants, or glycan profile. A potential mechanism of action is presented for each amino acid or metal factor based on its role in oxidation chemistry. This work served not only to mitigate the tryptophan oxidation issue in two Lilly biopharmaceuticals in development, but also to increase our knowledge and appreciation for the impact of media components on product quality. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 32:178–188, 2016     

Surface Engineering of 3D Gas Diffusion Electrodes for High‐Performance H2 Production with Nonprecious Metal Catalysts

In this work, a methodology is demonstrated to engineer gas diffusion electrodes for nonprecious metal catalysts. Highly active transition metal phosphides are prepared on carbon‐based gas diffusion electrodes with low catalyst loadings by modifying the O/C ratio at the surface of the electrode. These nonprecious metal catalysts yield extraordinary performance as measured by low overpotentials (51 mV at ?10 mA cm^?2), unprecedented mass activities (>800 A g^?1 at 100 mV overpotential), high turnover frequencies (6.96 H₂ s^?1 at 100 mV overpotential), and high durability for a precious metal‐free catalyst in acidic media. It is found that a high O/C ratio induces a more hydrophilic surface directly impacting the morphology of the CoP catalyst. The improved hydrophilicity, stemming from introduced oxyl groups on the carbon electrode, creates an electrode surface that yields a well‐distributed growth of cobalt electrodeposits and thus a well‐dispersed catalyst layer with high surface area upon phosphidation. This report demonstrates the high‐performance achievable by CoP at low loadings which facilitates further cost reduction, an important part of enabling the large‐scale commercialization of non‐platinum group metal catalysts. The fabrication strategies described herein offer a pathway to lower catalyst loading while achieving high efficiency and promising stability on a 3D electrode.     

Marine biology of the pacific lamprey Entosphenus tridentatus

Reviews in Fish Biology and Fisheries - Pacific lamprey Entosphenus tridentatus has an anadromous life cycle that begins with larvae that filter-feed in freshwater, followed by transformation into...     

OPA1 cleavage depends on decreased mitochondrial ATP level and bivalent metals

OPA1, an intra-mitochondrial dynamin GTPase, is a key actor of outer and inner mitochondrial membrane dynamic. OPA1 amino-terminal cleavage by PARL and m-AAA proteases was recently proposed to participate to the mitochondrial network dynamic in a DeltaPsi(m)-dependent way, and to apoptosis. Here, by an in vitro approach combining the use of purified mitochondrial fractions and mitochondrial targeting drugs, we intended to identify the central stimulus responsible for OPA1 cleavage. We confirm that apoptosis induction and PTPore opening, as well as DeltaPsi(m) dissipation induce OPA1 cleavage. Nevertheless, our experiments evidenced that decreased mitochondrial ATP levels, either generated by apoptosis induction, DeltaPsi(m) dissipation or inhibition of ATP synthase, is the common and crucial stimulus that controls OPA1 processing. In addition, we report that ectopic iron addition activates OPA1 cleavage, whereas zinc inhibits this process. These results suggest that the ATP-dependent OPA1 processing plays a central role in correlating the energetic metabolism to mitochondrial dynamic and might be involved in the pathophysiology of diseases associated to excess of iron or depletion of zinc and ATP.     

Physiological consequences of an altered flow regime on Alabama bass (Micropterus henshalli)

There are numerous studies on the effects of dams on aquatic biota, yet relatively little is known about whether hydropeaking activities cause physiological change in fish. Using Alabama bass (Micropterus henshalli) as a model, we evaluated whether hydropeaking in a regulated river altered glucocorticoid stress responsiveness relative to fish from an unregulated tributary. Blood samples were collected at the time of capture (baseline) and then collected again after a 1‐hr period of confinement (response). Leukocyte profiles (blood smears) were created and plasma was extracted to assess plasma cortisol levels and neutrophils and lymphocyte (N:L) ratios, between sites and times to evaluate differences between sites and the two sampling periods. Baseline cortisol levels were higher in fish collected from the regulated river compared to those from unregulated site, but response levels of cortisol were similar between sites. Baseline and response level N:L ratios did not differ between sites. High baseline levels of cortisol suggested that fish exposed to regulated flows expressed an altered stress response and were likely in an allostatic state, i.e., attempting to acclimate. Further research is needed to understand how altered stress responses due to hydropeaking flows may be affecting fish.