Attribution of Space-Time Variability in Global-Ocean Dissolved Inorganic Carbon.

The inventory and variability of oceanic dissolved inorganic carbon (DIC) is driven by the interplay of physical, chemical, and biological processes. Quantifying the spatiotemporal variability of these drivers is crucial for a mechanistic understanding of the ocean carbon sink and its future trajectory. Here, we use the Estimating the Circulation and Climate of the Ocean-Darwin ocean biogeochemistry state estimate to generate a global-ocean, data-constrained DIC budget and investigate how spatial and seasonal-to-interannual variability in three-dimensional circulation, air-sea CO2 flux, and biological processes have modulated the ocean sink for 1995-2018. Our results demonstrate substantial compensation between budget terms, resulting in distinct upper-ocean carbon regimes. For example, boundary current regions have strong contributions from vertical diffusion while equatorial regions exhibit compensation between upwelling and biological processes. When integrated across the full ocean depth, the 24-year DIC mass increase of 64 Pg C (2.7 Pg C year-1) primarily tracks the anthropogenic CO2 growth rate, with biological processes providing a small contribution of 2% (1.4 Pg C). In the upper 100 m, which stores roughly 13% (8.1 Pg C) of the global increase, we find that circulation provides the largest DIC gain (6.3 Pg C year-1) and biological processes are the largest loss (8.6 Pg C year-1). Interannual variability is dominated by vertical advection in equatorial regions, with the 1997-1998 El Niño-Southern Oscillation causing the largest year-to-year change in upper-ocean DIC (2.1 Pg C). Our results provide a novel, data-constrained framework for an improved mechanistic understanding of natural and anthropogenic perturbations to the ocean sink.

Global-scale dispersal and connectivity in mangroves.

Dispersal provides a key mechanism for geographical range shifts in response to changing environmental conditions. For mangroves, which are highly susceptible to climate change, the spatial scale of dispersal remains largely unknown. Here we use a high-resolution, eddy- and tide-resolving numerical ocean model to simulate mangrove propagule dispersal across the global ocean and generate connectivity matrices between mangrove habitats using a range of floating periods. We find high rates of along-coast transport and transoceanic dispersal across the Atlantic, Pacific, and Indian Oceans. No connectivity is observed between populations on either side of the American and African continents. Archipelagos, such as the Galapagos and those found in Polynesia, Micronesia, and Melanesia, act as critical stepping-stones for dispersal across the Pacific Ocean. Direct and reciprocal dispersal routes across the Indian Ocean via the South Equatorial Current and seasonally reversing monsoon currents, respectively, allow connectivity between western Indian Ocean and Indo-West Pacific sites. We demonstrate the isolation of the Hawaii Islands and help explain the presence of mangroves on the latitudinal outlier Bermuda. Finally, we find that dispersal distance and connectivity are highly sensitive to the minimum and maximum floating periods. We anticipate that our findings will guide future research agendas to quantify biophysical factors that determine mangrove dispersal and connectivity, including the influence of ocean surface water properties on metabolic processes and buoyancy behavior, which may determine the potential of viably reaching a suitable habitat. Ultimately, this will lead to a better understanding of global mangrove species distributions and their response to changing climate conditions.

The Lyme disease spirochete's BpuR DNA/RNA-binding protein is differentially expressed during the mammal-tick infectious cycle, which affects translation of the SodA superoxide dismutase.

When the Lyme disease spirochete, Borrelia burgdorferi, transfers from a feeding tick into a human or other vertebrate host, the bacterium produces vertebrate-specific proteins and represses factors needed for arthropod colonization. Previous studies determined that the B. burgdorferi BpuR protein binds to its own mRNA and autoregulates its translation, and also serves as co-repressor of erp transcription. Here, we demonstrate that B. burgdorferi controls transcription of bpuR, expressing high levels of bpuR during tick colonization but significantly less during mammalian infection. The master regulator of chromosomal replication, DnaA, was found to bind specifically to a DNA sequence that overlaps the bpuR promoter. Cultured B. burgdorferi that were genetically manipulated to produce elevated levels of BpuR exhibited altered levels of several proteins, although BpuR did not impact mRNA levels. Among these was the SodA superoxide dismutase, which is essential for mammalian infection. BpuR bound to sodA mRNA in live B. burgdorferi, and a specific BpuR-binding site was mapped 5' of the sodA open reading frame. Recognition of posttranscriptional regulation of protein levels by BpuR adds another layer to our understanding of the B. burgdorferi regulome, and provides further evidence that bacterial protein levels do not always correlate directly with mRNA levels.

Bpur, the Lyme disease spirochete's PUR domain protein: identification as a transcriptional modulator and characterization of nucleic acid interactions.

The PUR domain is a nucleic acid-binding motif found in critical regulatory proteins of higher eukaryotes and in certain species of bacteria. During investigations into mechanisms by which the Lyme disease spirochete controls synthesis of its Erp surface proteins, it was discovered that the borrelial PUR domain protein, Bpur, binds with high affinity to double-stranded DNA adjacent to the erp transcriptional promoter. Bpur was found to enhance the effects of the erp repressor protein, BpaB. Bpur also bound single-stranded DNA and RNA, with relative affinities RNA > double-stranded DNA > single-stranded DNA. Rational site-directed mutagenesis of Bpur identified amino acid residues and domains critical for interactions with nucleic acids, and it revealed that the PUR domain has a distinct mechanism of interaction with each type of nucleic acid ligand. These data shed light on both gene regulation in the Lyme spirochete and functional mechanisms of the widely distributed PUR domain.

The use of a prodrug approach to minimize potential CNS exposure of next generation quinoline methanols while maintaining efficacy in in vivo animal models.

The use of mefloquine (MQ) for antimalarial treatment and prophylaxis has diminished largely in response to concerns about its neurologic side effects. An analog campaign designed to maintain the efficacy of MQ while minimizing blood-brain barrier (BBB) penetration has resulted in the synthesis of a prodrug with comparable-to-superior in vivo efficacy versus mefloquine in a P. berghei mouse model while exhibiting a sixfold reduction in CNS drug levels. The prodrug, WR319670, performed poorly compared to MQ in in vitro efficacy assays, but had promising in vitro permeability in an MDCK-MDR1 cell line BBB permeability screen. Its metabolite, WR308245, exhibited high predicted BBB penetration with excellent in vitro efficacy. Both WR319670 and WR308245 cured 5/5 animals in separate in vivo efficacy studies. The in vivo efficacy of WR319670 was thought to be due to the formation of a more active metabolite, specifically WR308245. This was supported by pharmacokinetics studies in non-infected mice, which showed that both IV and oral administration of WR319670 produced essentially identical levels of WR319670 and WR308245 in both plasma and brain samples at all time points. In these studies, the levels of WR308245 in the brain were 1/4 and 1/6 that of MQ in similar IV and oral studies, respectively. These data show that the use of WR319670 as an antimalarial prodrug was able to maintain efficacy in in vivo efficacy screens, while significantly lowering overall penetration of drug and metabolites across the BBB.

Characterization of in vivo metabolites of WR319691, a novel compound with activity against Plasmodium falciparum.

WR319691 has been shown to exhibit reasonable Plasmodium falciparum potency in vitro and exhibits reduced permeability across MDCK cell monolayers, which as part of our screening cascade led to further in vivo analysis. Single-dose pharmacokinetics was evaluated after an IV dose of 5 mg/kg in mice. Maximum bound and unbound brain levels of WR319691 were 97 and 0.05 ng/g versus approximately 1,600 and 3.2 ng/g for mefloquine. The half-life of WR319691 in plasma was approximately 13 h versus 23 h for mefloquine. The pharmacokinetics of several N-dealkylated metabolites was also evaluated. Five of six of these metabolites were detected and maximum total and free brain levels were all lower after an IV dose of 5 mg/kg WR319691 compared to mefloquine at the same dose. These data provide proof of concept that it is feasible to substantially lower the brain levels of a 4-position modified quinoline methanol in vivo without substantially decreasing potency against P. falciparum in vitro.

Structure-activity relationships amongst 4-position quinoline methanol antimalarials that inhibit the growth of drug sensitive and resistant strains of Plasmodium falciparum.

Utilizing mefloquine as a scaffold, a next generation quinoline methanol (NGQM) library was constructed to identify early lead compounds that possess biological properties consistent with the target product profile for malaria chemoprophylaxis while reducing permeability across the blood-brain barrier. The library of 200 analogs resulted in compounds that inhibit the growth of drug sensitive and resistant strains of Plasmodium falciparum. Herein we report selected chemotypes and the emerging structure-activity relationship for this library of quinoline methanols.

Highly variable iron content modulates iceberg-ocean fertilisation and potential carbon export.

Marine phytoplankton growth at high latitudes is extensively limited by iron availability. Icebergs are a vector transporting the bioessential micronutrient iron into polar oceans. Therefore, increasing iceberg fluxes due to global warming have the potential to increase marine productivity and carbon export, creating a negative climate feedback. However, the magnitude of the iceberg iron flux, the subsequent fertilization effect and the resultant carbon export have not been quantified. Using a global analysis of iceberg samples, we reveal that iceberg iron concentrations vary over 6 orders of magnitude. Our results demonstrate that, whilst icebergs are the largest source of iron to the polar oceans, the heterogeneous iron distribution within ice moderates iron delivery to offshore waters and likely also affects the subsequent ocean iron enrichment. Future marine productivity may therefore be not only sensitive to increasing total iceberg fluxes, but also to changing iceberg properties, internal sediment distribution and melt dynamics.

Hematopoietic Stem Cells: Normal Versus Malignant.

SIGNIFICANCE: The long-term hematopoietic stem cell (LT-HSC) demonstrates characteristics of self-renewal and the ability to manage expansion of the hematopoietic compartment while maintaining the capacity for differentiation into hematopoietic stem/progenitor cell (HSPC) and terminal subpopulations. Deregulation of the HSPC redox environment results in loss of signaling that normally controls HSPC fate, leading to a loss of HSPC function and exhaustion. The characteristics of HSPC exhaustion via redox stress closely mirror phenotypic traits of hematopoietic malignancies and the leukemic stem cell (LSC). These facets elucidate the HSC/LSC redox environment as a druggable target and a growing area of cancer research. Recent Advances: Although myelosuppression and exhaustion of the hematopoietic niche are detrimental side effects of classical chemotherapies, new agents that modify the HSPC/LSC redox environment have demonstrated the potential for protection of normal HSPC function while inducing cytotoxicity within malignant populations. CRITICAL ISSUES: New therapies must preserve, or only slightly disturb normal HSPC redox balance and function, while simultaneously altering the malignant cellular redox state. The cascade nature of redox damage makes this a critical and delicate line for the development of a redox-based therapeutic index. FUTURE DIRECTIONS: Recent evidence demonstrates the potential for redox-based therapies to impact metabolic and epigenetic factors that could contribute to initial LSC transformation. This is balanced by the development of therapies that protect HSPC function. This pushes toward therapies that may alter the HSC/LSC redox state but lead to initiation cell fate signaling lost in malignant transformation while protecting normal HSPC function. Antioxid. Redox Signal.

Extracellular Vesicles Released by Cardiomyocytes in a Doxorubicin-Induced Cardiac Injury Mouse Model Contain Protein Biomarkers of Early Cardiac Injury.

Purpose: Cardiac injury is a major cause of death in cancer survivors, and biomarkers for it are detectable only after tissue injury has occurred. Extracellular vesicles (EV) remove toxic biomolecules from tissues and can be detected in the blood. Here, we evaluate the potential of using circulating EVs as early diagnostic markers for long-term cardiac injury.Experimental Design: Using a mouse model of doxorubicin (DOX)-induced cardiac injury, we quantified serum EVs, analyzed proteomes, measured oxidized protein levels in serum EVs released after DOX treatment, and investigated the alteration of EV content.Results: Treatment with DOX caused a significant increase in circulating EVs (DOX_EV) compared with saline-treated controls. DOX_EVs exhibited a higher level of 4-hydroxynonenal adducted proteins, a lipid peroxidation product linked to DOX-induced cardiotoxicity. Proteomic profiling of DOX_EVs revealed the distinctive presence of brain/heart, muscle, and liver isoforms of glycogen phosphorylase (GP), and their origins were verified to be heart, skeletal muscle, and liver, respectively. The presence of brain/heart GP (PYGB) in DOX_EVs correlated with a reduction of PYGB in heart, but not brain tissues. Manganese superoxide dismutase (MnSOD) overexpression, as well as pretreatment with cardioprotective agents and MnSOD mimetics, resulted in a reduction of EV-associated PYGB in mice treated with DOX. Kinetic studies indicated that EVs containing PYGB were released prior to the rise of cardiac troponin in the blood after DOX treatment, suggesting that PYGB is an early indicator of cardiac injury.Conclusions: EVs containing PYGB are an early and sensitive biomarker of cardiac injury. Clin Cancer Res; 24(7); 1644-53. ©2017 AACRSee related commentary by Zhu and Gius, p. 1516.

Simultaneous quantitation of oxidized and reduced glutathione via LC-MS/MS: An insight into the redox state of hematopoietic stem cells.

Cellular redox balance plays a significant role in the regulation of hematopoietic stem-progenitor cell (HSC/MPP) self-renewal and differentiation. Unregulated changes in cellular redox homeostasis are associated with the onset of most hematological disorders. However, accurate measurement of the redox state in stem cells is difficult because of the scarcity of HSC/MPPs. Glutathione (GSH) constitutes the most abundant pool of cellular antioxidants. Thus, GSH metabolism may play a critical role in hematological disease onset and progression. A major limitation to studying GSH metabolism in HSC/MPPs has been the inability to measure quantitatively GSH concentrations in small numbers of HSC/MPPs. Current methods used to measure GSH levels not only rely on large numbers of cells, but also rely on the chemical/structural modification or enzymatic recycling of GSH and therefore are likely to measure only total glutathione content accurately. Here, we describe the validation of a sensitive method used for the direct and simultaneous quantitation of both oxidized and reduced GSH via liquid chromatography followed by tandem mass spectrometry (LC-MS/MS) in HSC/MPPs isolated from bone marrow. The lower limit of quantitation (LLOQ) was determined to be 5.0ng/mL for GSH and 1.0ng/mL for GSSG with lower limits of detection at 0.5ng/mL for both glutathione species. Standard addition analysis utilizing mouse bone marrow shows that this method is both sensitive and accurate with reproducible analyte recovery. This method combines a simple extraction with a platform for the high-throughput analysis, allows for efficient determination of GSH/GSSG concentrations within the HSC/MPP populations in mouse, chemotherapeutic treatment conditions within cell culture, and human normal/leukemia patient samples. The data implicate the importance of the modulation of GSH/GSSG redox couple in stem cells related diseases.

Eubacterial SpoVG homologs constitute a new family of site-specific DNA-binding proteins.

A site-specific DNA-binding protein was purified from Borrelia burgdorferi cytoplasmic extracts, and determined to be a member of the highly conserved SpoVG family. This is the first time a function has been attributed to any of these ubiquitous bacterial proteins. Further investigations into SpoVG orthologues indicated that the Staphylococcus aureus protein also binds DNA, but interacts preferentially with a distinct nucleic acid sequence. Site-directed mutagenesis and domain swapping between the S. aureus and B. burgdorferi proteins identified that a 6-residue stretch of the SpoVG α-helix contributes to DNA sequence specificity. Two additional, highly conserved amino acid residues on an adjacent ß-sheet are essential for DNA-binding, apparently by contacts with the DNA phosphate backbone. Results of these studies thus identified a novel family of bacterial DNA-binding proteins, developed a model of SpoVG-DNA interactions, and provide direction for future functional studies on these wide-spread proteins.