A DUF3494 ice-binding protein with a root cap domain in a streptophyte glacier ice alga.

Ice-binding proteins (IBPs) of the DUF3494 type have been found in many ice-associated unicellular photoautotrophs, including chlorophytes, haptophytes, diatoms and a cyanobacterium. Unrelated IBPs have been found in many land plants (streptophytes). Here we looked for IBPs in two streptophyte algae that grow only on glaciers, a group in which IBPs have not previously been examined. The two species, Ancylonema nordenskioeldii and Ancylonema. alaskanum, belong to the class Zygnematophyceae, whose members are the closest relatives to all land plants. We found that one of them, A. nordenskioeldii, expresses a DUF3494-type IBP that is similar to those of their chlorophyte ancestors and that has not previously been found in any streptophytes. The protein is unusual in having what appears to be a perfect array of TXT motifs that have been implicated in water or ice binding. The IBP strongly binds to ice and almost certainly has a role in mitigating the daily freeze-thaw cycles that the alga is exposed to during late summer. No IBP was found in the second species, A. alaskanum, which may rely more on glycerol production for its freeze-thaw tolerance. The IBP is also unusual in having a 280-residue domain with a ß sandwich structure (which we designate as the DPH domain) that is characteristic of root cap proteins of land plants, and that may have a role in forming IBP oligomers. We also examined existing transcriptome data obtained from land plants to better understand the tissue and temperature dependence of expression of this domain.

Correction: Concentrations and properties of ice nucleating substances in exudates from Antarctic sea-ice diatoms.

Correction for 'Concentrations and properties of ice nucleating substances in exudates from Antarctic sea-ice diatoms' by Yu Xi et al., Environ. Sci.: Processes Impacts, 2021, 23, 323-334, DOI: 10.1039/D0EM00398K.

Changes in Australian food anaphylaxis admission rates following introduction of updated allergy prevention guidelines.

BACKGROUND: Food anaphylaxis admission rates have increased steadily in recent decades. Global food allergy prevention guidelines recommending early introduction of allergenic foods were introduced in 2015-2016. Australian guidelines to not delay the introduction of allergenic foods were introduced in 2007-2008. OBJECTIVE: Our aim was to examine whether introduction of Australian guidelines (2007-2008) and global allergy prevention guidelines (2015-2016) were associated with reductions in food anaphylaxis admission rates. METHODS: We compared food anaphylaxis admission rates across 3 periods: 1998-1999 to 2006-2007, 2007-2008 to 2014-2015, and 2015-2016 to 2018-2019. RESULTS: Annual food anaphylaxis admission rates increased 9-fold between 1998-1999 and 2018-2019, from 2.0 per 105 population to 18.2 per 105 population; the highest absolute rates were in those younger than 1 year. When year-on-year rates of change were examined across the 3 time periods, the annual rate of increase slowed after 2007-2008 in those aged 1 to 4 years (17.6%, 6.2%, and 3.9% per year, respectively) and those aged 5 to 9 years (22%, 13.9%, and -2.4%, respectively), and after 2015-2016, in those aged 10 to 14 years (17.5%, 18.0%, and 10.8%, respectively). By contrast, the year-on-year rate of increase accelerated in those younger than 1 year (5.2%, 8.0%, and 18.0%, respectively) and in all age groups older than 15 years. CONCLUSIONS: Although food anaphylaxis continues to increase overall, there is preliminary evidence indicating a slowing in the year-on-year rate of increase among those aged 1 to 4, 5 to 9, and 10 to 14 years, coinciding with introduction of updated infant feeding and allergy prevention guidelines in 2007-2008 and 2015-2016. Changes to the guidelines may have contributed to an attenuated rate of increase in food anaphylaxis in these age groups, as well as to increased rates in those younger than 1 year.

Investigating the Growth of Algae Under Low Atmospheric Pressures for Potential Food and Oxygen Production on Mars.

With long-term missions to Mars and beyond that would not allow resupply, a self-sustaining Bioregenerative Life Support System (BLSS) is essential. Algae are promising candidates for BLSS due to their completely edible biomass, fast growth rates and ease of handling. Extremophilic algae such as snow algae and halophilic algae may also be especially suited for a BLSS because of their ability to grow under extreme conditions. However, as indicated from over 50 prior space studies examining algal growth, little is known about the growth of algae at close to Mars-relevant pressures. Here, we explored the potential for five algae species to produce oxygen and food under low-pressure conditions relevant to Mars. These included Chloromonas brevispina, Kremastochrysopsis austriaca, Dunaliella salina, Chlorella vulgaris, and Spirulina plantensis. The cultures were grown in duplicate in a low-pressure growth chamber at 670 ± 20 mbar, 330 ± 20 mbar, 160 ± 20 mbar, and 80 ± 2.5 mbar pressures under continuous light exposure (62-70 µmol m-2 s-1). The atmosphere was evacuated and purged with CO2 after sampling each week. Growth experiments showed that D. salina, C. brevispina, and C. vulgaris were the best candidates to be used for BLSS at low pressure. The highest carrying capacities for each species under low pressure conditions were achieved by D. salina at 160 mbar (30.0 ± 4.6 × 105 cells/ml), followed by C. brevispina at 330 mbar (19.8 ± 0.9 × 105 cells/ml) and C. vulgaris at 160 mbar (13.0 ± 1.5 × 105 cells/ml). C. brevispina, D. salina, and C. vulgaris all also displayed substantial growth at the lowest tested pressure of 80 mbar reaching concentrations of 43.4 ± 2.5 × 104, 15.8 ± 1.3 × 104, and 57.1 ± 4.5 × 104 cells per ml, respectively. These results indicate that these species are promising candidates for the development of a Mars-based BLSS using low pressure (∼200-300 mbar) greenhouses and inflatable structures that have already been conceptualized and designed.

Concentrations and properties of ice nucleating substances in exudates from Antarctic sea-ice diatoms.

The ocean contains ice nucleating substances (INSs), some of which can be emitted to the atmosphere where they can influence the formation and properties of clouds. A possible source of INSs in the ocean is exudates from sea-ice diatoms. Here we examine the concentrations and properties of INSs in supernatant samples from dense sea-ice diatom communities collected from Ross Sea and McMurdo Sound in the Antarctic. The median freezing temperatures of the samples ranged from approximately -17 to -22 °C. Based on our results and a comparison with results reported in the literature, the ice nucleating ability of exudates from sea-ice diatoms is likely not drastically different from the ice nucleating ability of exudates from temperate diatoms. The number of INSs per mass of DOC for the supernatant samples were lower than those reported previously for the sea surface microlayer and bulk sea water collected in the Arctic and Atlantic. The INSs in the supernatant sample collected from Ross Sea were not sensitive to temperatures up to 100 °C, were larger than 300 kDa, and were different from ice shaping and recrystallization inhibiting molecules present in the same sample. Possible candidates for these INSs include polysaccharide containing nanogels. The INSs in the supernatant sample collected from McMurdo Sound were sensitive to temperatures of 80 and 100 °C and were larger than 1000 kDa. Possible candidates for these INSs include protein containing nanogels.

Case Studies Discussing the Pathology, Immunogenicity, and Proposed Mechanism of Toxicity of an Inhaled Anti-TGFß Humanized Fab Antibody in Non-Human Primates and Mice.

Treatment of nonhuman primates and mice with a humanized antigen-binding fragment (Fab) antibody (UCBFab) inhibiting transforming growth factor ß via daily inhalation for up to 13 weeks resulted in low systemic exposure but high local exposure in the lung. Target engagement was demonstrated by reduced levels of signal transducers, phosphoSMAD and plasminogen activator inhibitor-1 in the bronchoalveolar lavage fluid (BALF). Treatment was associated with a high frequency and titer of antidrug antibodies, indicating high local immunogenicity, and local pathology within the lung and draining lymph nodes. Microscopic changes were characterized by perivascular (PV) and peribronchiolar (PB) mononuclear inflammatory cell (MIC) infiltrates that were principally lymphocytic in nature and mixed inflammatory cell infiltrates and/or inflammation within the alveoli. Immunohistochemical investigation revealed a predominantly CD68-positive macrophage and CD3- and CD8>CD4-positive T-cell response in the alveoli, whereas within the airways, there was a variable mixture of CD3-positive T cells, CD20-positive B cells, and CD68-positive macrophages. Increased cellularity of the draining lymph nodes was also noted, indicating the presence of an immune response to the inhaled test article. Morphologic changes did not progress over time, and all changes partially recovered. Increased leukocytes (principally macrophages) in BALF cytology correlated with the changes seen by histopathology.

Ice-Binding Proteins Associated with an Antarctic Cyanobacterium, Nostoc sp. HG1.

Ice-binding proteins (IBPs) have been identified in numerous polar algae and bacteria, but so far not in any cyanobacteria, despite the abundance of cyanobacteria in polar regions. We previously reported strong IBP activity associated with an Antarctic Nostoc species. In this study, to identify the proteins responsible, as well as elucidate their origin, we sequenced the DNA of an environmental sample of this species, designated Nostoc sp. HG1, and its bacterial community and attempted to identify IBPs by looking for known IBPs in the metagenome and by looking for novel IBPs by tandem mass spectrometry (MS/MS) proteomics analyses of ice affinity-purified proteins. The metagenome contained over 116 DUF3494-type IBP genes, the most common type of IBP identified so far. One of the IBPs could be confidently assigned to Nostoc, while the others could be attributed to diverse bacteria, which, surprisingly, accounted for the great majority of the metagenome. Recombinant Nostoc IBPs (nIBPs) had strong ice-structuring activities, and their circular dichroism spectra were consistent with the secondary structure of a DUF3494-type IBP. nIBP is unusual in that it is the only IBP identified so far to have a PEP (amino acid motif) C-terminal signal, a signal that has been associated with anchoring to the outer cell membrane. These results suggest that the observed IBP activity of Nostoc sp. HG1 was due to a combination of endogenous and exogenous IBPs. Amino acid and nucleotide sequence analyses of nIBP raise the possibility that it was acquired from a planctomycete.IMPORTANCE The horizontal transfer of genes encoding ice-binding proteins (IBPs), proteins that confer freeze-thaw tolerance, has allowed many microorganisms to expand their ranges into polar regions. One group of microorganisms for which nothing is known about its IBPs is cyanobacteria. In this study, we identified a cyanobacterial IBP and showed that it was likely acquired from another bacterium, probably a planctomycete. We also showed that a consortium of IBP-producing bacteria living with the Nostoc contribute to its IBP activity.

Glycerol Is an Osmoprotectant in Two Antarctic Chlamydomonas Species From an Ice-Covered Saline Lake and Is Synthesized by an Unusual Bidomain Enzyme.

Glycerol, a compatible solute, has previously been found to act as an osmoprotectant in some marine Chlamydomonas species and several species of Dunaliella from hypersaline ponds. Recently, Chlamydomonas reinhardtii and Dunaliella salina were shown to make glycerol with an unusual bidomain enzyme, which appears to be unique to algae, that contains a phosphoserine phosphatase and glycerol-3-phosphate dehydrogenase. Here we report that two psychrophilic species of Chlamydomonas (C. spp. UWO241 and ICE-MDV) from Lake Bonney, Antarctica also produce high levels of glycerol to survive in the lake's saline waters. Glycerol concentration increased linearly with salinity and at 1.3 M NaCl, exceeded 400 mM in C. sp. UWO241, the more salt-tolerant strain. We also show that both species expressed several isoforms of the bidomain enzyme. An analysis of one of the isoforms of C. sp. UWO241 showed that it was strongly upregulated by NaCl and is thus the likely source of glycerol. These results reveal another adaptation of the Lake Bonney Chlamydomonas species that allow them to survive in an extreme polar environment.

An ice-binding protein from an Arctic population of American dunegrass, Leymus mollis.

Several cold-hardy grasses have been shown to have ice-binding proteins (IBPs) that protect against freeze-thaw injury. Here, we looked for IBP activity in an Alaskan coastal grass, Leymus mollis (Pooidae), that had not previously been examined. Rhizome tissue had strong ice-structuring and ice recrystallization inhibiting (IRI) activities, indicating the probable presence of IBPs. The gene sequence of an IBP was obtained. The sequence encoded a 118-amino acid IRI domain composed of eight repeats and that was 80% identical to the IRI domain of the IBP of perennial ryegrass Lolium perenne. The predicted 3D structure of the IRI domain had eight beta-roll coils like those in L. perenne IBP.

Immune complex disease in a chronic monkey study with a humanised, therapeutic antibody against CCL20 is associated with complement-containing drug aggregates.

Despite the potential for the chemokine class as therapeutic targets in immune mediated disease, success has been limited. Many chemokines can bind to multiple receptors and many receptors have multiple ligands, with few exceptions. One of those exceptions is CCL20, which exclusively pairs to CCR6 and is associated with several immunologic conditions, thus providing a promising therapeutic target. Following successful evaluation in a single dose, first time in human clinical study, GSK3050002-a humanized IgG1 monoclonal antibody against human CCL20-was evaluated in a 26-week cynomolgus monkey toxicology study. A high incidence of unexpected vascular and organ inflammation was observed microscopically, leading to the decision to halt clinical development. Here we report a dose-responsive increase in the incidence and severity of inflammation in multiple organs from monkeys receiving 30 and 300 mg/kg/week by either subcutaneous or intravenous injection. Histomorphological changes resembled an immune complex-mediated pathology, which is often due to formation of anti-drug antibodies in monkeys receiving a human protein therapeutic and thus not predictive of clinical outcome. However, the presentation was atypical in that there was a clear dose response with a very high incidence of inflammation with a low incidence of ADA that did not correlate well individually. Additionally, the immunohistologic presentation was atypical in that the severity and distribution of tissue inflammation was greater than the numbers of associated immune complexes (i.e., granular deposits). An extensive ex vivo analysis of large molecular weight protein complexes in monkey serum from this study and in human serum samples demonstrated a time-dependent aggregation of GSK3050002, that was not predicted by in vitro assays. The aggregates also contained complement components. These findings support the hypothesis that immune complexes of drug aggregates, not necessarily including anti-drug antibodies, can fix complement, accumulate over time, and trigger immune complex disease. A situation which may have increased clinical relevance than typical anti-drug antibody-associated immune complex disease in monkeys administered human antibody proteins.

Formation and Glomerular Deposition of Immune Complexes in Mice Administered Human Antibodies: Evaluation of Dose, Frequency, and Biomarkers.

Administration of human protein-based drugs to animals often leads to formation of antidrug antibodies (ADAs) that may form circulating immune complexes (CICs) with the dosed protein. Circulating immune complexes can activate and bind complement (cCICs), and if large amount of CICs or cCICs is formed, the clearance mechanism potentially becomes saturated, which can lead to immune complex (IC) deposition and inflammation. To obtain a better understanding of the underlying factors, including the relationship between different dose regimes on IC formation and deposition and identification of possible biomarkers of IC deposition and IC-related pathological changes in kidneys, BALB/c and C57BL/6J mice were administered with human anti-tumor necrosis factor α (aTNFα, adalimumab) or a humanized anti-TNP (aTNP) antibody for 13 weeks. Particularly, ADA, CIC, cCIC formation, IC deposition, and glomerulonephritis were observed in C57BL/6J administered with aTNFα, whereas the immunologic response was minor in BALB/c mice administered with aTNFα and in BALB/c and C57BL/6J mice administered aTNP. Changing dose levels or increasing dosing frequency of aTNFα on top of an already-established CIC and cCIC response did not lead to substantial changes in CIC, cCIC formation, or IC deposition. Finally, no association between the presence of CICs or cCIC in plasma and glomerular IC deposition and/or glomerulonephritis was observed.

Ice-Binding Proteins in a Chrysophycean Snow Alga: Acquisition of an Essential Gene by Horizontal Gene Transfer.

All ice-associated algae examined so far have genes for ice-binding proteins (IBPs), which suggest that these proteins are essential for survival in icy habitats. The most common type of IBP, type 1 IBPs (also referred to as DUF3494 IBPs), is also found in ice-associated bacteria and fungi. Previous studies have suggested that algal IBP genes were acquired by horizontal transfer from other microorganisms (probably bacteria). However, it remains unclear whether this is also the case for algae distantly related to the ones examined so far and whether microorganisms other than bacteria could be the donors. Furthermore, there is only limited evidence that these proteins are expressed at low temperature. Here, we show that Kremastochrysopsis austriaca (Chrysophyceae), an Austrian snow alga that is not closely related to any of the ice-associated algae examined so far, also produces IBPs, although their activity was weak. Sequencing the algal genome and the transcriptomes of cells grown at 1 and 15°C revealed three isoforms of a type 1 IBP. In agreement with their putative function, the three isoforms were strongly upregulated by one to two orders of magnitude at 1°C compared to 15°C. In a phylogenetic tree, the K. austriaca IBPs were distant from other algal IBPs, with the closest matches being bacterial proteins. These results suggest that the K. austriaca IBPs were derived from a gene that was acquired from a bacterium unrelated to other IBP donor bacteria and confirm by their presence in yet another alga the essential role of algal IBPs.

Formation and glomerular deposition of immune complexes in mice administered bovine serum albumin: Evaluation of dose, frequency, and biomarkers.

In preclinical toxicity studies, species-foreign proteins administered to animals frequently leads to formation of anti-drug antibodies (ADA). Such antibodies may form circulating immune complexes (CIC) with the administered protein. These CIC can activate the classical complement pathway, thereby forming complement-bound CIC (cCIC); if large of amounts of CIC or cCIC is formed, the clearance mechanism may become saturated which potentially leads to vascular immune complex (IC) deposition and inflammation. Limited information is available on the effect of different treatment related procedures as well as biomarkers of IC-related vascular disease. In order to explore the effect of different dose regimens on IC formation and deposition, and identification of possible biomarkers of IC deposition and IC-related pathological changes, C57BL/6J and BALB/c mice were dosed subcutaneously twice weekly with bovine serum albumin (BSA) for 13 weeks without adjuvant. After 6 and 13 weeks, CIC and cCIC were detected in plasma; after 13 weeks, IC deposition was detected in kidney glomeruli. In particular immunohistochemistry double-staining was shown to be useful for detection of IC deposition. Increasing dosing frequency or changing BSA dose level on top of an already established CIC and cCIC response did not cause changes in IC deposition, but CIC and cCIC concentrations tended to decrease with increased dose level, and increased cCIC formation was observed after more frequent dosing. The presence of CIC in plasma was associated with glomerular IC deposits in the dose regimen study; however, the use of CIC or cCIC as potential biomarkers for IC deposition and IC-related pathological changes, needs to be explored further.

Editor's Highlight: Complete Attenuation of Mouse Lung Cell Proliferation and Tumorigenicity in CYP2F2 Knockout and CYP2F1 Humanized Mice Exposed to Inhaled Styrene for up to 2 Years Supports a Lack of Human Relevance.

Styrene is a mouse-specific lung carcinogen, and short-term mode of action studies have demonstrated that cytotoxicity and/or cell proliferation, and genomic changes are dependent on CYP2F2 metabolism. The current study examined histopathology, cell proliferation, and genomic changes in CD-1, C57BL/6 (WT), CYP2F2(-/-) (KO), and CYP2F2(-/-) (CYP2F1, 2B6, 2A13-transgene) (TG; humanized) mice following exposure for up to 104 weeks to 0- or 120-ppm styrene vapor. Five mice per treatment group were sacrificed at 1, 26, 52, and 78 weeks. Additional 50 mice per treatment group were followed until death or 104 weeks of exposure. Cytotoxicity was present in the terminal bronchioles of some CD-1 and WT mice exposed to styrene, but not in KO or TG mice. Hyperplasia in the terminal bronchioles was present in CD-1 and WT mice exposed to styrene, but not in KO or TG mice. Increased cell proliferation, measured by KI-67 staining, occurred in CD-1 and WT mice exposed to styrene for 1 week, but not after 26, 52, or 78 weeks, nor in KO or TG mice. Styrene increased the incidence of bronchioloalveolar adenomas and carcinomas in CD-1 mice. No increase in lung tumors was found in WT despite clear evidence of lung toxicity, or, KO or TG mice. The absence of preneoplastic lesions and tumorigenicity in KO and TG mice indicates that mouse-specific CYP2F2 metabolism is responsible for both the short-term and chronic toxicity and tumorigenicity of styrene, and activation of styrene by CYP2F2 is a rodent MOA that is neither quantitatively or qualitatively relevant to humans.

Multiple ice-binding proteins of probable prokaryotic origin in an Antarctic lake alga, Chlamydomonas sp. ICE-MDV (Chlorophyceae).

Ice-associated algae produce ice-binding proteins (IBPs) to prevent freezing damage. The IBPs of the three chlorophytes that have been examined so far share little similarity across species, making it likely that they were acquired by horizontal gene transfer (HGT). To clarify the importance and source of IBPs in chlorophytes, we sequenced the IBP genes of another Antarctic chlorophyte, Chlamydomonas sp. ICE-MDV (Chlamy-ICE). Genomic DNA and total RNA were sequenced and screened for known ice-associated genes. Chlamy-ICE has as many as 50 IBP isoforms, indicating that they have an important role in survival. The IBPs are of the DUF3494 type and have similar exon structures. The DUF3494 sequences are much more closely related to prokaryotic sequences than they are to sequences in other chlorophytes, and the chlorophyte IBP and ribosomal 18S phylogenies are dissimilar. The multiple IBP isoforms found in Chlamy-ICE and other algae may allow the algae to adapt to a greater variety of ice conditions than prokaryotes, which typically have a single IBP gene. The predicted structure of the DUF3494 domain has an ice-binding face with an orderly array of hydrophilic side chains. The results indicate that Chlamy-ICE acquired its IBP genes by HGT in a single event. The acquisitions of IBP genes by this and other species of Antarctic algae by HGT appear to be key evolutionary events that allowed algae to extend their ranges into polar environments.

Modeling Trait Anxiety: From Computational Processes to Personality.

Computational methods are increasingly being applied to the study of psychiatric disorders. Often, this involves fitting models to the behavior of individuals with subclinical character traits that are known vulnerability factors for the development of psychiatric conditions. Anxiety disorders can be examined with reference to the behavior of individuals high in "trait" anxiety, which is a known vulnerability factor for the development of anxiety and mood disorders. However, it is not clear how this self-report measure relates to neural and behavioral processes captured by computational models. This paper reviews emerging computational approaches to the study of trait anxiety, specifying how interacting processes susceptible to analysis using computational models could drive a tendency to experience frequent anxious states and promote vulnerability to the development of clinical disorders. Existing computational studies are described in the light of this perspective and appropriate targets for future studies are discussed.

Evolutionary genomics of the cold-adapted diatom Fragilariopsis cylindrus.

The Southern Ocean houses a diverse and productive community of organisms. Unicellular eukaryotic diatoms are the main primary producers in this environment, where photosynthesis is limited by low concentrations of dissolved iron and large seasonal fluctuations in light, temperature and the extent of sea ice. How diatoms have adapted to this extreme environment is largely unknown. Here we present insights into the genome evolution of a cold-adapted diatom from the Southern Ocean, Fragilariopsis cylindrus, based on a comparison with temperate diatoms. We find that approximately 24.7 per cent of the diploid F. cylindrus genome consists of genetic loci with alleles that are highly divergent (15.1 megabases of the total genome size of 61.1 megabases). These divergent alleles were differentially expressed across environmental conditions, including darkness, low iron, freezing, elevated temperature and increased CO2. Alleles with the largest ratio of non-synonymous to synonymous nucleotide substitutions also show the most pronounced condition-dependent expression, suggesting a correlation between diversifying selection and allelic differentiation. Divergent alleles may be involved in adaptation to environmental fluctuations in the Southern Ocean.