Identification of Differential Drought Response Mechanisms in Medicago sativa subsp. sativa and falcata through Comparative Assessments at the Physiological, Biochemical, and Transcriptional Levels

Alfalfa (Medicago sativa L.) is an extensively grown perennial forage legume, and although it is relatively drought tolerant, it consumes high amounts of water and depends upon irrigation in many regions. Given the progressive decline in water available for irrigation, as well as an escalation in climate change-related droughts, there is a critical need to develop alfalfa cultivars with improved drought resilience. M. sativa subsp. falcata is a close relative of the predominantly cultivated M. sativa subsp. sativa, and certain accessions have been demonstrated to exhibit superior performance under drought. As such, we endeavoured to carry out comparative physiological, biochemical, and transcriptomic evaluations of an as of yet unstudied drought-tolerant M. sativa subsp. falcata accession (PI 641381) and a relatively drought-susceptible M. sativa subsp. sativa cultivar (Beaver) to increase our understanding of the molecular mechanisms behind the enhanced ability of falcata to withstand water deficiency. Our findings indicate that unlike the small number of falcata genotypes assessed previously, falcata PI 641381 may exploit smaller, thicker leaves, as well as an increase in the baseline transcriptional levels of genes encoding particular transcription factors, protective proteins, and enzymes involved in the biosynthesis of stress-related compounds. These findings imply that different falcata accessions/genotypes may employ distinct drought response mechanisms, and the study provides a suite of candidate genes to facilitate the breeding of alfalfa with enhanced drought resilience in the future.

Silver Nanostructures: Limited Sensitivity of Detection, Toxicity and Anti-Inflammation Effects

Nanosilver with sizes 1–100 nm at least in one dimension is widely used due to physicochemical, anti-inflammatory, anti-angiogenesis, antiplatelet, antifungal, anticancer, antibacterial, and antiviral properties. Three modes of the nanosilver action were suggested: “Trojan horse”, inductive, and quantum mechanical. The Ag+ cations have an affinity to thiol, amino, phosphate, and carboxyl groups. Multiple mechanisms of action towards proteins, DNA, and membranes reduce a risk of pathogen resistance but inevitably cause toxicity for cells and organisms. Silver nanoparticles (AgNP) are known to generate two reactive oxygen species (ROS)-superoxide (•O2−) and hydroxyl (•OH) radicals, which inhibit the cellular antioxidant enzymes (superoxide dismutase, catalase, and glutathione peroxidase) and cause mechanical damage of membranes. Ag+ release and replacement by electrolyte ions with potential formation of insoluble AgCl result in NP instability and interactions of heavy metals with nucleic acids and proteins. Protein shells protect AgNP core from oxidation, dissolution, and aggregation, and provide specific interactions with ligands. These nanoconjugates can be used for immunoassays and diagnostics, but the sensitivity is limited at 10 pg and specificity is restricted by binding with protective proteins (immunoglobulins, fibrinogen, albumin, and others). Thus, broad implementation of Ag nanostructures revealed limitations such as instability; binding with major blood proteins; damage of proteins, nucleic acids, and membranes; and immunosuppression of the majority of cytokines.

Immunoreactive Protein Repertoires of Ehrlichia chaffeensis and E. canis Reveal the Dominance of Hypothetical Proteins and Conformation-dependent Antibody Epitopes

The immunomes of Ehrlichia chaffeensis ( E. ch. ) and E. canis ( E. ca. ) have recently be revised to include immunodominant hypothetical proteins with conformational antibody epitopes. In this study, we examined 216 E. ch. and 190 E. ca. highly antigenic proteins according to ANTIGENpro and also performed a genome-wide hypothetical protein analysis ( E. ch. n=104; E. ca. n=124) for immunoreactivity. Using cell-free protein expression and immunoanalysis, 118 E. ch. and 39 E. ca . proteins reacted with sera from naturally E. ch. -infected patients or E. ca. -infected dogs. Moreover, 22 E. ch. and 18 E. ca. proteins consistently and strongly reacted with a panel of patient or canine sera. A subset of E. ch. (n=18) and E. ca. (n=9) proteins were identified as immunodominant. Consistent with our previous study, most proteins were classified as hypothetical and the antibody epitopes exhibited complete or partial conformation-dependence. The majority (28/40; 70%) of E. ch. and E. ca. proteins contained transmembrane domains and 19 (48%) were predicted to be secreted effectors. The antigenic repertoires of E. ch. and E. ca. were mostly diverse and suggest that the immunomes of these closely related ehrlichiae are dominated by species-specific conformational antibody epitopes. This study reveals a significant group of previously undefined E. ch. and E. ca. antigens and reaffirms the importance of conformation-dependent epitopes as targets of anti- Ehrlichia immune responses. These findings substantially expand our understanding of host- Ehrlichia immune responses, advance efforts to define the molecular features of protective proteins and improve prospects for effective vaccines for the ehrlichioses.

Divalent Metal Transporter 1 Knock-Down Modulates IL-1β Mediated Pancreatic Beta-Cell Pro-Apoptotic Signaling Pathways through the Autophagic Machinery

Pro-inflammatory cytokines promote cellular iron-import through enhanced divalent metal transporter-1 (DMT1) expression in pancreatic β-cells, consequently cell death. Inhibition of β-cell iron-import by DMT1 silencing protects against apoptosis in animal models of diabetes. However, how alterations of signaling networks contribute to the protective action of DMT1 knock-down is unknown. Here, we performed phosphoproteomics using our sequential enrichment strategy of mRNA, protein, and phosphopeptides, which enabled us to explore the concurrent molecular events in the same set of wildtype and DMT1-silenced β-cells during IL-1β exposure. Our findings reveal new phosphosites in the IL-1β-induced proteins that are clearly reverted by DMT1 silencing towards their steady-state levels. We validated the levels of five novel phosphosites of the potential protective proteins using parallel reaction monitoring. We also confirmed the inactivation of autophagic flux that may be relevant for cell survival induced by DMT1 silencing during IL-1β exposure. Additionally, the potential protective proteins induced by DMT1 silencing were related to insulin secretion that may lead to improving β-cell functions upon exposure to IL-1β. This global profiling has shed light on the signal transduction pathways driving the protection against inflammation-induced cell death in β-cells after DMT1 silencing.

Circulating proteins protect against renal decline and progression to end-stage renal disease in patients with diabetes

Diabetic kidney disease (DKD) and its major clinical manifestation, progressive renal decline that leads to end-stage renal disease (ESRD), are a major health burden for individuals with diabetes. The disease process that underlies progressive renal decline comprises factors that increase risk as well as factors that protect against this outcome. Using untargeted proteomic profiling of circulating proteins from individuals in two independent cohorts with type 1 and type 2 diabetes and varying stages of DKD followed for 7 to 15 years, we identified three elevated plasma proteins—fibroblast growth factor 20 (OR, 0.69; 95% CI, 0.54 to 0.88), angiopoietin-1 (OR, 0.72; 95% CI, 0.57 to 0.91), and tumor necrosis factor ligand superfamily member 12 (OR, 0.75; 95% CI, 0.59 to 0.95)—that were associated with protection against progressive renal decline and progression to ESRD. The combined effect of these three protective proteins was demonstrated by very low cumulative risk of ESRD in those who had baseline concentrations above median for all three proteins, whereas the cumulative risk of ESRD was high in those with concentrations below median for these proteins at the beginning of follow-up. This protective effect was shown to be independent from circulating inflammatory proteins and clinical covariates and was confirmed in a third cohort of diabetic individuals with normal renal function. These three protective proteins may serve as biomarkers to stratify diabetic individuals according to risk of progression to ESRD and might also be investigated as potential therapeutics to delay or prevent the onset of ESRD.

Elovanoids downregulate SARS-CoV-2 cell-entry, canonical mediators and enhance protective signaling in human alveolar cells

The pro-homeostatic lipid mediators elovanoids (ELVs) attenuate cell binding and entrance of the SARS-CoV-2 receptor-binding domain (RBD) as well as of the SARS-CoV-2 virus in human primary alveoli cells in culture. We uncovered that very-long-chain polyunsaturated fatty acid precursors (VLC-PUFA, n-3) activate ELV biosynthesis in lung cells. Both ELVs and their precursors reduce the binding to RBD. ELVs downregulate angiotensin-converting enzyme 2 (ACE2) and enhance the expression of a set of protective proteins hindering cell surface virus binding and upregulating defensive proteins against lung damage. In addition, ELVs and their precursors decreased the signal of spike (S) protein found in SARS-CoV-2 infected cells, suggesting that the lipids curb viral infection. These findings open avenues for potential preventive and disease-modifiable therapeutic approaches for COVID-19.

Expression of protective proteins and morphological changes in the rat brain after prolonged exposure to dust

Introduction. The fundamental mechanisms of the body's pathological reaction to coal and dust exposure are hypoxia, excessive activation of free radical processes, structural and metabolic disorders in various organs. Organ-specific molecular defense mechanisms begin to function in the form of changes in the level of proteins with antihypoxic (HIF-3a), chaperone (HSP72), and antioxidant functions (HOx-1 - heme-oxygenase, Prx-1 - peroxiredoxin) under damaging effects. Its high level contributes to the restoration of cells' functional state or indicates significant damage in tissues. Hypoxia and free radical processes are known to lead to severe brain damage and behavioral disorders. To date, little is known about the expression of protective proteins and morphological changes in the brain under prolonged exposure to coal-rock dust on the body. The study aimed to learn the level of intracellular protective proteins HIF-3a, HSP72, HOx-1, Prx-1, and morphological changes in the brain in the dynamics of long-term dust exposure. Materials and methods. Sixty white male Wistar rats weighing 200-250 g of the same age took part in the experiment. Dust exposure was modeled by way of dynamic inhalation priming of rats with coal-rock dust (coal of a gas-fat brand) in an intermittent mode for 12 weeks. We perform morphological studies of the brain after 1, 3, 6, 9, and 12 weeks of dust exposure. The cytosolic fraction of brain tissue researchers determined the expression level of HIF-3a, HSP72, HOx-1, and Prx-1 by Western blot analysis. We selected the activity of free radical processes in the brain tissue. Results. Long-term exposure to coal-rock dust on the body at the morphological level in the brain revealed changes that indicate the development of hypoxia and activation of free radical processes: microvascular disorders, pericellular edema, severe dystrophic damage to neurons, focal loss of neurons, activation of glial cells. Activation of the protective proteins HIF-3a, HSP72, HOx-1, and Prx-1 in the early stages (1-3 weeks) of coal-dust exposure provided compensation for free radical processes in brain neurons. An increase in the duration of dust exposure of more than six weeks influences a low level of HSP72, but high HIF-3a and Prx-1, indicating an increase in hypoxic and free radical damage brain. Conclusions. The results obtained to expand the understanding of the morphological and molecular mechanisms that occur in the brain tissue during prolonged dust exposure to the body are essential for developing methods for organ-specific pharmacological correction.

EPIGENETIC REGULATION OF ADAPTOGENESIS BY PATHOLOGY AND AGING

В обзоре с точки зрения эпигенетики рассмотрены адаптационные возможности организма при патологии и старении. Апаптация организма к внутренним и внешним факторам осуществляется единой гуморальной защитной системой организма, включающей гипоталамо-гипофизарно-эпифизарную и гипоталамогипофизарно-тимусную оси. Короткие пептиды AEDG, AEDP, EDR, KED, EW, KE являются эпигенетическими регуляторами экспрессии генов и синтеза белков, которые могут быть вовлечены в адаптацию при стрессе и активацию гипоталамо-гипофизарно-эпифизарной и гипоталамогипофизарно-тимусной осей. Указанные короткие пептиды регулируют синтез белков теплового шока, стресспротекторных белков, цитокинов, факторов фибринолиза и гемостаза. Эти пептиды могут участвовать в первичной и отсроченной эпигенетической регуляции адаптивного ответа при стрессе, патологии и старении. Ранняя функциональная диагностика нарушения сопряжения звеньев единой гуморальной защитной системы организма при возраст-ассоциированных заболеваниях позволит выявить недостаточную синхронность эпигенетических механизмов, при которой наступает истощение и снижение резервных возможностей организма. Применение пептидов может нивелировать проявления адаптационного синдрома при стрессе и возрастной патологии. The organism adaptive possibilities by pathology and aging are discussed in account of the epigenetic. The organism adaptation to inner and external factors is carried out by organism unite humoral protective system, inclusive hypothalamus-hypophysis-pineal and hypothalamus-hypophysis-thymus axises. AEDG, AEDP, EDR, KED, EW, KE short peptides are the epigenetic regulators of gene expression and protein synthesis, which can be involve to the adaptation by stress and in the activation of hypothalamus-hypophysispineal and hypothalamus-hypophysis-thymus axises. These short peptides regulate the synthesis of proteins of heat shock, stress-protective proteins, cytocines, fibrinolysis and hemostasis factors and can participate in primary and tardive epigenetic regulation of adaptive response by stress, pathology and aging. The early functional diagnostic of element disturbances of organism unite humoral protective system by age-associative pathology can be usefull for the detection of deficient synchronization of epigenetic mechanisms, by wich the depletion and decrease of organism reserve possibilities occurs. The use of peptide can grade the adaptive syndrome manifestation by the stress and age pathology.

Protective properties of the mucosa of the upper respiratory tract and methods of their activation

The article discusses the issues of the protective mechanisms of the mucous membranes of the upper respiratory tract: mucociliary clearance, production of protective proteins and the mucosal immunity system. The data on the significance and relationship of lymphoid structures associated with mucous membranes of various anatomical localization are presented. The mechanisms of action of topical bacterial lysates are considered, their effectiveness in the treatment and prevention of respiratory diseases and their complications is described.