Host Physiology and Pathogenic Variation of Cochliobolus heterostrophus Strains with Mutations in the G Protein Alpha Subunit, CGA1

Conserved eukaryotic signaling proteins participate in development and disease in plant-pathogenic fungi. Strains with mutations in CGA1, a heterotrimeric G protein G alpha subunit gene of the maize pathogen Cochliobolus heterostrophus, are defective in several developmental pathways. Conidia from CGA1 mutants germinate as abnormal, straight-growing germ tubes that form few appressoria, and the mutants are female sterile. Nevertheless, these mutants can cause normal lesions on plants, unlike other filamentous fungal plant pathogens in which functional homologues of CGA1 are required for full virulence. Δcga1 mutants of C. heterostrophus were less infective of several maize varieties under most conditions, but not all, as virulence was nearly normal on detached leaves. This difference could be related to the rapid senescence of detached leaves, since delaying senescence with cytokinin also had differential effects on the virulence of the wild type and the Δcga1 mutant. In particular, detached leaves may provide a more readily available nutrient source than attached leaves. Decreased fitness of Δcga1 as a pathogen may reflect conditions under which full virulence requires signal transduction through CGA1-mediated pathways. The virulence of these signal transduction mutants is thus affected differentially by the physiological state of the host.     

Karyomeres in early cleavage embryos of ophryotrocha labronica lagreca and bacci

Summary Karyomeres or chromosome vesicles occur regularly at all cell divisions in cleavage embryos ofOphryotrocha labronica up to the 16-cell stage. They are formed as separate units, containing one or several nucleolus-like bodies (NLB) as well as intranuclear annulate lamellae (IAL), but coalesce later into a compound nucleus, in connection with copious blebbing and simultaneous appearance of cytoplasmic annulate lamellae (CAL). Labelling of the early embryos with³H-thymidine revealed marked localization of the synthesized DNA to the karyomere envelope region, whereas³H-uridine incorporation, indicating RNA synthesis, was sparse and notably absent in the NLB. On the other hand the latter structure like the envelopes preferentially incorporated³H-myoinositol, and displayed considerable labelling with³H-leucine. The mechanism and general significance of karyomere formation is discussed with particular attention to the NLB and their possible involvement in nuclear membrane formation.This work has been supported by the Swedish Natural Science Research Council and Kungliga Fysiografiska Sällskapet, Lund.The excellent technical assistance of Mrs Annagreta Petersen and Mrs Lena Olsson is gratefully acknowledged.     

Manganese Deficiency Can Replace High Oxygen Levels Needed for Lignin Peroxidase Formation by Phanerochaete chrysosporium

The combined effects of Mn and oxygen on lignin peroxidase (LIP) activity and isozyme composition in Phanerochaete chrysosporium were studied by using shallow stationary cultures grown in the presence of limited or excess N. When no Mn was added, LIP was formed in both N-limited and N-excess cultures exposed to air, but no LIP activity was observed at Mn concentrations greater than 13 mg/liter. In oxygen-flushed, N-excess cultures, LIP was formed at all Mn concentrations, and the peak LIP activity values in the extracellular fluid were nearly identical in the presence of Mn concentrations ranging from 3 to 1,500 mg/liter. When the availability of oxygen to cultures exposed to air was increased by growing the fungus under nonimmersed liquid conditions, higher levels of Mn were needed to suppress LIP formation compared with the levels needed in shallow stationary cultures. The composition of LIP isozymes was affected by the levels of N and Mn. Addition of veratryl alcohol to cultures exposed to air did not eliminate the suppressive effect of Mn on LIP formation. A deficiency of Mn in N-excess cultures resulted in lower biomass and a lower rate of glucose consumption than in the presence of Mn. In addition, almost no activity of the antioxidant enzyme Mn superoxide dismutase was observed in Mn-deficient, N-excess cultures, but the activity of this enzyme increased as the Mn concentration increased from 3 to 13 mg/liter. No Zn/Cu superoxide dismutase activity was observed in N-excess cultures regardless of the Mn concentration.     

Proteomic analysis of necroptotic extracellular vesicles

Necroptosis is a regulated and inflammatory form of cell death. We, and others, have previously reported that necroptotic cells release extracellular vesicles (EVs). We have found that necroptotic EVs are loaded with proteins, including the phosphorylated form of the key necroptosis-executing factor, mixed lineage kinase domain-like kinase (MLKL). However, neither the exact protein composition, nor the impact, of necroptotic EVs have been delineated. To characterize their content, EVs from necroptotic and untreated U937 cells were isolated and analyzed by mass spectrometry-based proteomics. A total of 3337 proteins were identified, sharing a high degree of similarity with exosome proteome databases, and clearly distinguishing necroptotic and control EVs. A total of 352 proteins were significantly upregulated in the necroptotic EVs. Among these were MLKL and caspase-8, as validated by immunoblot. Components of the ESCRTIII machinery and inflammatory signaling were also upregulated in the necroptotic EVs, as well as currently unreported components of vesicle formation and transport, and necroptotic signaling pathways. Moreover, we found that necroptotic EVs can be phagocytosed by macrophages to modulate cytokine and chemokine secretion. Finally, we uncovered that necroptotic EVs contain tumor neoantigens, and are enriched with components of antigen processing and presentation. In summary, our study reveals a new layer of regulation during the early stage of necroptosis, mediated by the secretion of specific EVs that influences the microenvironment and may instigate innate and adaptive immune responses. This study sheds light on new potential players in necroptotic signaling and its related EVs, and uncovers the functional tasks accomplished by the cargo of these necroptotic EVs.Subject terms: Necroptosis, Cell death and immune response     

High‐Temperature Treatment of Li‐Rich Cathode Materials with Ammonia: Improved Capacity and Mean Voltage Stability during Cycling

Li‐rich electrode materials of the family x Li₂MnO₃·(1?x )LiNi_a Co_b Mn_c O₂ (a + b + c = 1) suffer a voltage fade upon cycling that limits their utilization in commercial batteries despite their extremely high discharge capacity, ≈250 mA h g^?1. Li‐rich, 0.35Li₂MnO₃·0.65LiNi_0.35Mn_0.45Co_0.20O₂, is exposed to NH₃ at 400 °C, producing materials with improved characteristics: enhanced electrode capacity and a limited average voltage fade during 100 cycles in half cells versus Li. Three main changes caused by NH₃ treatment are established. First, a general bulk reduction of Co and Mn is observed via X‐ray photoelectron spectroscopy and X‐ray absorption near edge structure. Next, a structural rearrangement lowers the coordination number of Co? O and Mn? O bonds, as well as formation of a surface spinel‐like structure. Additionally, Li⁺ removal from the bulk causes the formation of surface LiOH, Li₂CO₃, and Li₂O. These structural and surface changes can enhance the voltage and capacity stability of the Li‐rich material electrodes after moderate NH₃ treatment times of 1–2 h.     

High Functional Diversity in Mycobacterium tuberculosis Driven by Genetic Drift and Human Demography

Mycobacterium tuberculosis infects one third of the human world population and kills someone every 15 seconds. For more than a century, scientists and clinicians have been distinguishing between the human- and animal-adapted members of the M. tuberculosis complex (MTBC). However, all human-adapted strains of MTBC have traditionally been considered to be essentially identical. We surveyed sequence diversity within a global collection of strains belonging to MTBC using seven megabase pairs of DNA sequence data. We show that the members of MTBC affecting humans are more genetically diverse than generally assumed, and that this diversity can be linked to human demographic and migratory events. We further demonstrate that these organisms are under extremely reduced purifying selection and that, as a result of increased genetic drift, much of this genetic diversity is likely to have functional consequences. Our findings suggest that the current increases in human population, urbanization, and global travel, combined with the population genetic characteristics of M. tuberculosis described here, could contribute to the emergence and spread of drug-resistant tuberculosis.     

Cell-Wall Interactions and the Selective Bacteriostatic Activity of a Miniature Oligo-Acyl-Lysyl

The oligo-acyl-lysyl, C_12(ω7)K-β₁₂, is comprised of only three Lys residues. Despite its small size, it exhibits potent bacteriostatic activity against Gram-positive bacteria, but it is ∼10-fold less potent against Gram-negative bacteria. We followed the interactions of C_12(ω7)K-β₁₂ from its initial contact with the bacterial surface across the cell wall down to the cytoplasmic membrane. Binding to anionic lipids, as well as to negatively charged LPS and LTA, occurs with very high affinity. The C_12(ω7)K-β₁₂ does not cross the outer membrane of Gram-negative bacteria; rather, it achieves its action by depositing on the LPS layer, promoting surface adhesion and blocking passage of solutes. In Gram-positive bacteria, the thick peptidoglycan layer containing LTA allows passage of C_12(ω7)K-β₁₂ and promotes its accumulation in the small periplasm. From that location it is then driven to the membrane by strong electrostatic interactions. Despite its high potency against Gram-positive bacteria, this agent is not capable of efficiently breaking down the permeability barrier of the cytoplasmic membrane or of reaching an intracellular target, as suggested by the fact that it does not interact with DNA.