Computational and experimental analysis of foxtail millet under salt stress and selenium supplementation.

Salinity stress is a major threat to crop growth and productivity. Millets are stress-tolerant crops that can withstand the environmental constraints. Foxtail millet is widely recognized as a drought and salinity-tolerant crop owing to its efficient ROS scavenging mechanism. Ascorbate peroxidase (APX) is one of the reactive oxygen species (ROS) scavenging enzymes that leads to hydrogen peroxide (H2O2) detoxification and stabilization of the internal biochemical state of the cell under stress. This inherent capacity of the APX enzyme can further be enhanced by the application of an external mitigant. This study focuses on the impact of salt (NaCl) and selenium (Se) application on the APX enzyme activity of foxtail millet using in silico and in-vitro techniques and mRNA expression studies. The NaCl was applied in the concentrations, i.e., 150 mM and 200 mM, while the Se was applied in 1 µM, 5 µM, and 10 µM concentrations. The in silico studies involved three-dimensional structure modeling and molecular docking. The in vitro studies comprised the morphological and biochemical parameters, alongside mRNA expression studies in foxtail millet under NaCl stress and Se applications. The in silico studies revealed that the APX enzyme showed better interaction with Se as compared to NaCl, thus suggesting the enzyme-modulating role of Se. The morphological and biochemical analysis indicated that Se alleviated the NaCl (150 mM and 200 mM) and induced symptoms at 1 µM as compared to 5 and 10 µM by enhancing the morphological parameters, upregulating the gene expression and enzyme activity of APX, and ultimately reducing the H2O2 content significantly. The transcriptomic studies confirmed the upregulation of chloroplastic APX in response to salt stress and selenium supplementation. Hence, it can be concluded that Se as a mitigant at lower concentrations can alleviate NaCl stress in foxtail millet.

Glucose metabolic reprogramming and modulation in glycerol biosynthesis regulates drug resistance in clinical isolates of Candida.

AIMS: The study is aimed at understanding the novel molecular mechanisms governing drug resistance in the opportunistic fungi belonging to the genus Candida. METHODS AND RESULTS: This is a multipronged study wherein different assays like drug susceptibility and whole cell proteome analysis, stress tolerance assay, measurement of total internal glycerol content, western blot analysis, reactive oxygen species (ROS) measurement, glucose uptake, lactate production, ATP generation, and NADPH measurements were made.The study reveals an incidence of different species of Candida in the northern most part of India (Kashmir valley). Resistant isolates, mostly resistant to azoles were reported across all the species. The study revealed a difference in resistance mechanisms between Candida albicans and C. glabrata clinical isolates. Further, such resistance mechanism (in the case of C. albicans) was mostly mediated by Hexokinase 2 (Hxk2) and Glucose-6-phosphate dehydrogenase (G6pd). Increased expression of Hxk2 was associated with increased glucose uptake, more lactate production, and more ATP generation in drug-resistant C. albicans. At the same time, increased G6pd expression was responsible for the increased production of NADPH, which imparts a better ROS scavenging potential. While in C. glabrata the resistance was linked with glycerol metabolism, where the drug-resistant isolate tends to accumulate more glycerol as an osmolyte in response to external stresses. This glycerol accumulation was found to be triggered by the HOG1-MAPK pathway. CONCLUSION: The study concludes that, like various human malignant tumors, there is a strong correlation between drug resistance and aberrant cellular metabolism in the opportunistic fungi belonging to the genus Candida.

Exogenous zinc mitigates salinity stress by stimulating proline metabolism in proso millet (Panicum miliaceum L.).

Salinity is one of the most concerning ecological restrictions influencing plant growth, which poses a devastating threat to global agriculture. Surplus quantities of ROS generated under stress conditions have negative effects on plants' growth and survival by damaging cellular components, including nucleic acids, lipids, proteins and carbohydrates. However, low levels of ROS are also necessary because of their role as signalling molecules in various development-related pathways. Plants possess sophisticated antioxidant systems for scavenging as well as regulating ROS levels to protect cells from damage. Proline is one such crucial non-enzymatic osmolyte of antioxidant machinery that functions in the reduction of stress. There has been extensive research on improving the tolerance, effectiveness, and protection of plants against stress, and to date, various substances have been used to mitigate the adverse effects of salt. In the present study Zinc (Zn) was applied to elucidate its effect on proline metabolism and stress-responsive mechanisms in proso millet. The results of our study indicate the negative impact on growth and development with increasing treatments of NaCl. However, the low doses of exogenous Zn proved beneficial in mitigating the effects of NaCl by improving morphological and biochemical features. In salt-treated plants, the low doses of Zn (1 mg/L, 2 mg/L) rescued the negative impact of salt (150mM) as evidenced by increase in shoot length (SL) by 7.26% and 25.5%, root length (RL) by 21.84% and 39.07% and membrane stability index (MSI) by 132.57% and 151.58% respectively.The proline content improved at all concentrations with maximum increase of 66.65% at 2 mg/L Zn. Similarly, the low doses of Zn also rescued the salt induced stress at 200mM NaCl. The enzymes related to proline biosynthesis were also improved at lower doses of Zn. In salt treated plants (150mM), Zn (1 mg/L, 2 mg/L) increased the activity of P5CS by 19.344% and 21%. The P5CR and OAT activities were also improved with maximum increase of 21.66% and 21.84% at 2 mg/L Zn respectively. Similarly, the low doses of Zn also increased the activities of P5CS, P5CR and OAT at 200mM NaCl. Whereas P5CDH enzyme activity showed a decrease of 82.5% at 2mg/L Zn+150mM NaCl and 56.7% at 2mg/L Zn+200 mM NaCl. These results strongly imply the modulatory role of Zn in maintaining of proline pool during NaCl stress.

Selenate and selenite transporters in proso millet: Genome extensive detection and expression studies under salt stress and selenium.

Crops are susceptible to a variety of stresses and amongst them salinity of soil is a global agronomic challenge that has a detrimental influence on crop yields, thus posing a severe danger to our food security. Therefore, it becomes imperative to examine how plants respond to salt stress, develop a tolerance that allows them to live through higher salt concentrations and choose species that can endure salt stress. From the perspective of food, security millets can be substituted to avoid hardships because of their efficiency in dealing with salt stress. Besides, this problem can also be tackled by using beneficial exogenous elements. Selenium (Se) which exists as selenate or selenite is one such cardinal element that has been reported to alleviate salt stress. The present study aimed for identification of selenate and selenite transporters in proso millet (Panicum miliaceum L.), their expression under NaCl (salt stress) and Na2SeO3 (sodium selenite)treatments. This study identified eight transporters (RLM65282.1, RLN42222.1, RLN18407.1, RLM74477.1, RLN41904.1, RLN17428.1, RLN17268.1, RLM65753.1) that have a potential role in Se uptake in proso millet. We analyzed physicochemical properties, conserved structures, sub-cellular locations, chromosome location, molecular phylogenetic analysis, promoter regions prediction, protein-protein interactions, three-dimensional structure modeling and evaluation of these transporters. The analysis revealed the chromosome location and the number of amino acids present in these transporters as RLM65282.1 (16/646); RLN42222.1 (1/543); RLN18407.1 (2/483); RLM74477.1 (15/474); RLN41904.1 (1/521); RLN17428.1 (2/522); RLN17268.1(2/537);RLM65753.1 (16/539). The sub-cellular locations revealed that all the selenite transporters are located in plasma membrane whereas among selenate transporters RLM65282.1 and RLM74477.1 are located in mitochondria and RLN42222.1 and RLN18407.1 in chloroplast. The transcriptomic studies revealed that NaCl stress decreased the expression of both selenate and selenite transporters in proso millet and the applications of exogenous 1µM Se (Na2SeO3) increased the expression of these Se transporter genes. It was also revealed that selenate shows similar behavior as sulfate, while selenite transport resembles phosphate. Thus, it can be concluded that phosphate and sulphate transporters in millets are responsible for Se uptake.

ß-Nitrostyrene derivatives as broad range potential antifungal agents targeting fungal cell wall.

The prevalence of multidrug resistance has been increasingly witnessed during the past few decades. Resistance of human pathogenic fungi against the currently available antifungal agents has increased the frequency of fungal infections and associated mortality rates. The discovery of novel lead antifungal agents is important to challenge multidrug resistance. The present study examined the antifungal potential of chemically synthesized ß-Nitrostyrene derivatives. Among the eight ß-Nitrostyrene derivatives used in this study, SS45, SS46 and SS47 showed strong antifungal potential. The results show that ß-Nitrostyrene derivatives inhibited the growth of different species of human pathogenic Candida, particularly the highly prevalent C. albicans, C. glabrata and the emerging pathogenic C. auris species. Moreover, ß-Nitrostyrene derivatives also show strong antifungal activities against drug-resistant clinical isolates and drug transporter overexpressing fungal species. The drug susceptibility assays revealed that ß-Nitrostyrene derivatives are fungicidal and show the synergy of action when combined with antifungal drugs caspofungin and fluconazole. The transcriptomic study performed on C. albicans in the presence of ß-Nitrostyrene derivatives revealed the differential expression of genes related to cell wall metabolism. Mechanistically, ß-Nitrostyrene derivatives impact cell wall morphology, enhance ROS generation and modulate drug efflux. Collectively this study reveals that ß-Nitrostyrene derivatives have strong antifungal potential with a particular mode of activity similar to known cell wall perturbing antifungal agents and thus can be exploited as promising potential antifungal agents for further studies.

Role of Epigenetics in Modulating Phenotypic Plasticity against Abiotic Stresses in Plants.

Plants being sessile are always exposed to various environmental stresses, and to overcome these stresses, modifications at the epigenetic level can prove vital for their long-term survival. Epigenomics refers to the large-scale study of epigenetic marks on the genome, which include covalent modifications of histone tails (acetylation, methylation, phosphorylation, ubiquitination, and the small RNA machinery). Studies based on epigenetics have evolved over the years especially in understanding the mechanisms at transcriptional and posttranscriptional levels in plants against various environmental stimuli. Epigenomic changes in plants through induced methylation of specific genes that lead to changes in their expression can help to overcome various stress conditions. Recent studies suggested that epigenomics has a significant potential for crop improvement in plants. By the induction and modulation of various cellular processes like DNA methylation, histone modification, and biogenesis of noncoding RNAs, the plant genome can be activated which can help in achieving a quicker response against various plant stresses. Epigenetic modifications in plants allow them to adjust under varied environmental stresses by modulating their phenotypic plasticity and at the same time ensure the quality and yield of crops. The plasticity of the epigenome helps to adapt the plants during pre- and postdevelopmental processes. The variation in DNA methylation in different organisms exhibits variable phenotypic responses. The epigenetic changes also occur sequentially in the genome. Various studies indicated that environmentally stimulated epimutations produce variable responses especially in differentially methylated regions (DMR) that play a major role in the management of stress conditions in plants. Besides, it has been observed that environmental stresses cause specific changes in the epigenome that are closely associated with phenotypic modifications. However, the relationship between epigenetic modifications and phenotypic plasticity is still debatable. In this review, we will be discussing the role of various factors that allow epigenetic changes to modulate phenotypic plasticity against various abiotic stress in plants.

Synergistic effect of plant extract coupled silver nanoparticles in various therapeutic applications- present insights and bottlenecks.

The phytocomponent conjugated silver nanoparticles (AgNPs) have been extensively explored for various therapeutic applications such as antimicrobial, antioxidant, anticancer, anti-inflammatory, antidiabetic and anticoagulant effects. The bio-conjugation of Ag-based nanomaterial with plant extracts reduces their toxicity to biological systems and enhances their therapeutic effectiveness. The diversity of phytochemicals or capping agents provided by the plant extracts and the small size and large surface area of AgNPs permits maximum adsorption of these capping agents onto their surfaces that further promote the therapeutic performance of phytoconjugated AgNPs in various biomedical applications. The mechanistic action involved in antimicrobial and anticancer functions of AgNPs is mainly dependent on the induction of reactive oxygen species (ROS) resulting in cellular apoptosis and necrosis. This review summarizes the recent studies of various plant extract assisted synthesis of AgNPs, potential biomedical applications with the possible mechanism of action and major shortcomings affecting their therapeutic efficacy.

Metabolic flexibility and extensive adaptability governing multiple drug resistance and enhanced virulence in Candida albicans.

Commensal fungus-Candida albicans turn pathogenic during the compromised immunity of the host, causing infections ranging from superficial mucosal to dreadful systemic ones. C. albicans has evolved various adaptive measures which collectively contribute towards its enhanced virulence. Among fitness attributes, metabolic flexibility and vigorous stress response are essential for its pathogenicity and virulence. Metabolic flexibility provides a means for nutrient assimilation and growth in diverse host microenvironments and reduces the vulnerability of the pathogen to various antifungals besides evading host immune response(s). Inside the host micro-environments, C. albicans efficiently utilizes the multiple fermentable and non-fermentable carbon sources to sustain and proliferate in glucose deficit conditions. The utilization of alternative carbon sources further highlights the importance of understanding these pathways as the attractive and potential therapeutic target. A thorough understanding of metabolic flexibility and adaptation to environmental stresses is warranted to decipher in-depth insights into virulence and molecular mechanisms of fungal pathogenicity. In this review, we have attempted to provide a detailed and recent understanding of some key aspects of fungal biology. Particular focus will be placed on processes like nutrient assimilation and utilization, metabolic adaptability, virulence factors, and host immune response in C. albicans leading to its enhanced pathogenicity.

Comparative Study on Phytochemical Profile and Antioxidant Activity of an Epiphyte, Viscum album L. (White Berry Mistletoe), Derived from Different Host Trees.

The study aimed at evaluating the antioxidant profile of a medicinal epiphyte Viscum album L. harvested from three tree species, namely, Populus ciliata L, Ulmus villosa L., and Juglans regia L. The crude extracts were obtained with ethanol, methanol, and water and were evaluated for the total phenol content (TPC), total flavonoid content (TFC), and antioxidant activities using total reducing power (TRP), ferric reducing antioxidant power (FRAP), 1, 1-diphenyl 1-2-picryl-hydrazyl (DPPH), superoxide radical scavenging (SOR), and hydroxyl radical scavenging (•OH) assays. Our results showed that crude leaf extracts of plants harvested from the host Juglans regia L. exhibited higher yields of phytochemical constituents and noticeable antioxidative properties. The ethanolic leaf samples reported the highest phenols (13.46 ± 0.87 mg/g), flavonoids (2.38 ± 0.04 mg/g), FRAP (500.63 ± 12.58 µM Fe II/g DW), and DPPH (87.26% ± 0.30 mg/mL). Moreover, the highest values for TRP (4.24 ± 0.26 µg/mL), SOR (89.79% ± 0.73 mg/mL), and OH (67.16% ± 1.15 mg/mL) were obtained from aqueous leaf extracts. Further, Pearson correlation was used for quantifying the relationship between TPC, TFC, and antioxidant (FRAP, DPPH, SOR, OH) activities in Viscum album L. compared to their hosts. It was revealed that the epiphyte showed variation with the type of host plant and extracting solvent.

Exogenous Application of Zinc to Mitigate the Salt Stress in Vigna radiata (L.) Wilczek-Evaluation of Physiological and Biochemical Processes.

Salt stress adversely affects the growth and productivity of crops. However, reports suggest that the application of various micronutrients could help the plant to cope with this stress. Hence, the objective of the study was to examine the effect of exogenous application of Zinc (Zn) on salt tolerance in Vigna radiata (L.) Wilczek (mungbean). Mungbean is considered to be an economically important crop and possess a strategic position in Southeast Asian countries for sustainable crop production. It is rich in quality proteins, minerals and vitamins. Three weeks old grown seedlings were subjected to NaCl (150 mM and 200 mM) alone or with Zn (250 µM). After 21 days of treatment, plants were harvested for investigating morphological, physiological and biochemical changes. We found that the Zn application mitigates the negative effect upon plant growth to a variable extent. This may be attributed to the increased shoot and root length, improved chlorophyll and carotenoid contents, enhanced total soluble sugar (TSS), total soluble protein (TSP) and proline accumulation, decreased H2O2 content and increased enzymatic antioxidant activities. Zn's application improved the performance of the enzymes such as phenylalanine ammonia-lyase (PAL) and tyrosine ammonia-lyase (TAL) of the secondary metabolism, which resulted in the improvement of total phenol and flavonoids. The antioxidant activities such as 1,1diphenyl 2-picryl hydrazine (DPPH) and ferrous reducing antioxidant power assay (FRAP) of the plants also showed improved results in their salt only treatments. Furthermore, hydrogen peroxide (H2O2) and superoxide radical (SOD) scavenging activity were also improved upon the application of 250 µM zinc. Thus, Zn application in low doses offers promising potential for recovering plants suffering from salinity stress. In conclusion, we assume that zinc application improved salt tolerance in mungbean through the improvement of various physiological and photochemical processes which could prove to be useful in nutrient mediated management for crop improvement.

Understanding the Integrated Pathways and Mechanisms of Transporters, Protein Kinases, and Transcription Factors in Plants under Salt Stress.

Abiotic stress is the major threat confronted by modern-day agriculture. Salinity is one of the major abiotic stresses that influence geographical distribution, survival, and productivity of various crops across the globe. Plants perceive salt stress cues and communicate specific signals, which lead to the initiation of defence response against it. Stress signalling involves the transporters, which are critical for water transport and ion homeostasis. Various cytoplasmic components like calcium and kinases are critical for any type of signalling within the cell which elicits molecular responses. Stress signalling instils regulatory proteins and transcription factors (TFs), which induce stress-responsive genes. In this review, we discuss the role of ion transporters, protein kinases, and TFs in plants to overcome the salt stress. Understanding stress responses by components collectively will enhance our ability in understanding the underlying mechanism, which could be utilized for crop improvement strategies for achieving food security.

Aluminium stress modulates the osmolytes and enzyme defense system in Fagopyrum species.

The present investigation describes aluminum-induced changes in the leaves of two buckwheat species using both physiological and biochemical indices. With increasing levels of Al (viz. 100, 200 and 300 µM), the mean length of root, shoot as well as their biomass accumulation decreased linearly with respect to control. Tolerance test of F. kashmirianum revealed that it was more tolerant to Al-stress than F. tataricum as revealed by higher accumulation of Al in its roots without any significant damage. Translocation factor (TF) values of both species were found to be < 1, indicating more Al is restrained in roots. Total chlorophyll showed a non-significant increase in F. tataricum while as decreased in F. kashmirianum at 300 µM concentration besides, the carotenoid content exhibited inclined trend in F. tataricum and showed a concomitant decrease in F. kashmirianum. The anthocyanin level showed a non-significant decline in F. kashmirianum. Exposure to different Al-treatments enhances malondialdehyde (MDA), H2O2 and membrane stability index (MSI) in both species, with increases being greater in F. kashmirianum than F. tataricum as also revealed by DAB-mediated in vivo histo-chemical detection method. The osmolyte level in general were elevated in both buckwheat species however, enhancement was more in F. tataricum than F. kashmirianum. The activities of antioxidant enzymes viz. superoxide dismutase (SOD), ascorbate peroxidase (APX), guaiacol peroxidase (POD), glutathione reductase (GR), glutathione-S-transferase (GST) were positively correlated with Al-treatment except catalase (CAT) which exhibits a reverse outcome in F. kashmirianum. The present investigation could play an essential role to better understand the detoxification mechanisms of Al in plants.

Characterization of mercury-induced stress biomarkers in Fagopyrum tataricum plants.

The effect of mercury stress on antioxidant enzymes, lipid peroxidation, photosynthetic pigments, hydrogen peroxide content, osmolytes, and growth parameters in Tartary buckwheat were investigated. The effect of Hg-exposure was found to be time (15 and 30 days) and concentration (0, 25, 50, and 75 µM) dependent. Hg was readily absorbed by seedlings with higher content in roots and it resulted in reduction of root and shoot length. The root and shoot Hg uptakes were significantly and directly correlated with each other. However, the fresh mass and biomass increased up to 50 µM Hg-treatment at both time periods. A significant positive correlation was observed between biomass accumulation with relative water content. Hg levels were positively correlated with the production of hydrogen peroxide in leaves as evidenced by 3, 3-diaminobenzidine (DAB)-mediated tissue fingerprinting. The osmolyte levels in general were elevated except for proline and protein which showed a decline at 75 µM Hg-treatment at 30-days. Amongst the photosynthetic pigments, chlorophyll showed a decline while as carotenoid and anthocyanin levels were elevated. The activity of antioxidant enzymes such as ascorbate peroxidase (APX), guaiacol peroxidase (POD), glutathione reductase (GR), Glutathione-s-transferase (GST) and superoxide dismutase (SOD) were positively correlated with Hg-treatment except SOD, which declined at 75 µM Hg-treatment in 30-days old seedlings. Catalase (CAT) activity showed a positive correlation up to 50 µM Hg-treatment but at 75 µM Hg-stress it decreases at both 15 and 30 days.

Tomato Rab11a characterization evidenced a difference between SYP121-dependent and SYP122-dependent exocytosis.

The regulatory functions of Rab proteins in membrane trafficking lie in their ability to perform as molecular switches that oscillate between a GTP- and a GDP-bound conformation. The role of tomato LeRab11a in secretion was analyzed in tobacco protoplasts. Green fluorescent protein (GFP)/red fluorescent protein (RFP)-tagged LeRab11a was localized at the trans-Golgi network (TGN) in vivo. Two serines in the GTP-binding site of the protein were mutagenized, giving rise to the three mutants Rab11S22N, Rab11S27N and Rab11S22/27N. The double mutation reduced secretion of a marker protein, secRGUS (secreted rat beta-glucuronidase), by half, whereas each of the single mutations alone had a much smaller effect, showing that both serines have to be mutated to obtain a dominant negative effect on LeRab11a function. The dominant negative mutant was used to determine whether Rab11 is involved in the pathway(s) regulated by the plasma membrane syntaxins SYP121 and SYP122. Co-expression of either of these GFP-tagged syntaxins with the dominant negative Rab11S22/27N mutant led to the appearance of endosomes, but co-expression of GFP-tagged SYP122 also labeled the endoplasmic reticulum and dotted structures. However, co-expression of Rab11S22/27N with SYP121 dominant negative mutants decreased secretion of secRGUS further compared with the expression of Rab11S22/27N alone, whereas co-expression of Rab11S22/27N with SYP122 had no synergistic effect. With the same essay, the difference between SYP121- and SYP122-dependent secretion was then evidenced. The results suggest that Rab11 regulates anterograde transport from the TGN to the plasma membrane and strongly implicate SYP122, rather than SYP121. The differential effect of LeRab11a supports the possibility that SYP121 and SYP122 drive independent secretory events.