Peroxidase Superfamily Research Articles

Heterologous expression and characterization of a dye-decolorizing peroxidase from Ganoderma lucidum, and its application in decolorization and detoxifization of different types of dyes.

Dye-decolorizing peroxidases (DyPs) belong to a novel superfamily of heme peroxidases that can oxidize recalcitrant compounds. In the current study, the GlDyP2 gene from Ganoderma lucidum was heterologously expressed in Escherichia coli, and the enzymatic properties of the recombinant GlDyP2 protein were investigated. The GlDyP2 protein could oxidize not only the typical peroxidase substrate ABTS but also two lignin substrates, namely guaiacol and 2,6-dimethoxy phenol (DMP). For the ABTS substrate, the optimum pH and temperature of GlDyP2 were 4.0 and 35°C, respectively. The pH stability and thermal stability of GlDyP2 were also measured; the results showed that GlDyP2 could function normally in the acidic environment, with a T50 value of 51°C. Moreover, compared to untreated controls, the activity of GlDyP2 was inhibited by 1.60 mM of Mg2+, Ni2+, Mn2+, and ethanol; 0.16 mM of Cu2+, Zn2+, methanol, isopropyl alcohol, and Na2EDTA·2H2O; and 0.016 mM of Fe2+ and SDS. The kinetic constants of recombinant GlDyP2 for oxidizing ABTS, Reactive Blue 19, guaiacol, and DMP were determined; the results showed that the recombination GlDyP2 exhibited the strongest affinity and the most remarkable catalytic efficiency towards guaiacol in the selected substrates. GlDyP2 also exhibited decolorization and detoxification capabilities towards several dyes, including Reactive Blue 19, Reactive Brilliant Blue X-BR, Reactive Black 5, Methyl Orange, Trypan Blue, and Malachite Green. In conclusion, GlDyP2 has good application potential for treating dye wastewater.

Fine-mapping and candidate gene analysis of tuber eye depth in potato

Eye depth is an important agronomic trait affecting tubers' appearance, quality, and processing suitability. Hence, cultivating varieties with uniform shapes and shallow eye depth are important goals for potato breeding. In this study, based on the primary mapping of the tuber eye-depth locus using a small primary-segregating population, a large secondary-segregating population with 2,100 individuals was used to map the eye-depth locus further. A major quantitative trait locus for eye-depth on chromosome 10 was identified (designated qEyd10.1) using BSA-seq and traditional QTL mapping method. The qEyd10.1 could explain 55.0% of the eye depth phenotypic variation and was further narrowed to a 309.10 kb interval using recombinant analysis. To predict candidate genes, tissue sectioning and RNA-seq of the specific tuber tissues were performed. Genes encoding members of the peroxidase superfamily with likely roles in indole acetic acid regulation were considered the most promising candidates. These results will facilitate marker-assisted selection for the shallow-eye trait in potato breeding and provide a solid basis for eye-depth gene cloning and the analysis of tuber eye-depth regulatory mechanisms.

Transcriptional and physio-chemical responses of Tectona grandis L. triggered by teak defoliator

Tectona grandis (L) is a popular timber species cultivated in the Indian sub-continent and other countries because of its economic significance. However, the lack of quality planting material with special reference to defoliator herbivory is a major constraint because it negatively influences the establishment of thriving plantations. To address this, the so-called pest response in teak seedlings was investigated at the biochemical and molecular levels. The stress was levied by allowing defoliator infestation on 1-year-old seedlings for nine days at 3-day intervals, whereas control plants were maintained as uninfected. A considerable difference was observed for the considered traits among the infested plants at different intervals compared with the control. Total soluble protein, soluble sugars, tannin, and lignin accumulation increased with increasing stress intensity. Furthermore, catalase, polyphenol oxidase, and phenol content increased after defoliator feeding, whereas peroxidase activity decreased compared with control plants. In addition, six herbivory stress-responsive genes revealed the molecular response of teak. Among them, four genes viz. zinc finger protein (ZFP2), ethylene-responsive factor (ERF-1), catalase-1 (Cat1) and peroxidase super family protein (Psfp) were highly expressed in the infected plants as compared with the control plants. In contrast, the MYB-DNA binding domain gene was downregulated in infected plants compared to uninfected plants. Cytochrome P450 was up-regulated in T1 and T2 plants, while it was down-regulated in T3. Genes were highly expressed as the stress frequency increased. This study offers an improved understanding of biochemical and transcriptional defense responses against the teak defoliator, which could further support teak improvement for quality planting material production.

Brilliant green decolorization by dye decolorizing peroxidase producing bacterium, Achromobacter insolitus isolated from Forest soils of Similipal Biosphere Reserve, Odisha

Dye-decolorizing peroxidases (DyPs) are recently identified superfamily of heme-containing peroxidases which has received much attention for its potential application in dye decolourization. In the current research, six bacterial colonies (SDB1-6) showing lignolytic activity were isolated from forest soils of SBR using a minimal salt medium supplemented with alkali lignin and screened for DyP activity using brilliant green dye supplemented luria bertani agar medium. The highest DyP activity showing bacterium was identified using biochemical and 16SrRNA sequencing and was identified as Achromobacter insolitus. A. insolitus was found to be an efficient strain showing the highest DyP activity of 2.7 U/mL/ml at un-optimized condition. The RSM optimization was conducted to increase the DyP enzyme activity. A. insolitus demonstrated DyP activity of 6.20 U/mL/min with an increase of 2.29 times of enzyme activity in optimal condition of pH 7.0, temperature 25 °C, H2O2 concentration 0.1 mM, and time 24 h. The bacterium was further screened for dye decolorization taking crystal violet, methyl orange, methylene blue, brilliant green and methyl red dyes supplemented in LB medium. The bacterium showed maximum brilliant green degradation ability was analyzed using UV-Vis and FTIR analyses to characterize the degraded product. The highest brilliant green degradation activity of the A. insolitus in the present study could be exploited for bioremediation of brilliant green contaminated industrial effluents in a ecofriendly and cost effective way.

Target enrichment sequencing coupled with GWAS identifies MdPRX10 as a candidate gene in the control of budbreak in apple.

The timing of floral budbreak in apple has a significant effect on fruit production and quality. Budbreak occurs as a result of a complex molecular mechanism that relies on accurate integration of external environmental cues, principally temperature. In the pursuit of understanding this mechanism, especially with respect to aiding adaptation to climate change, a QTL at the top of linkage group (LG) 9 has been identified by many studies on budbreak, but the genes underlying it remain elusive. Here, together with a dessert apple core collection of 239 cultivars, we used a targeted capture sequencing approach to increase SNP resolution in apple orthologues of known or suspected A. thaliana flowering time-related genes, as well as approximately 200 genes within the LG9 QTL interval. This increased the 275 223 SNP Axiom® Apple 480K array dataset by an additional 40 857 markers. Robust GWAS analyses identified MdPRX10, a peroxidase superfamily gene, as a strong candidate that demonstrated a dormancy-related expression pattern and down-regulation in response to chilling. In-silico analyses also predicted the residue change resulting from the SNP allele associated with late budbreak could alter protein conformation and likely function. Late budbreak cultivars homozygous for this SNP allele also showed significantly up-regulated expression of C-REPEAT BINDING FACTOR (CBF) genes, which are involved in cold tolerance and perception, compared to reference cultivars, such as Gala. Taken together, these results indicate a role for MdPRX10 in budbreak, potentially via redox-mediated signaling and CBF gene regulation. Moving forward, this provides a focus for developing our understanding of the effects of temperature on flowering time and how redox processes may influence integration of external cues in dormancy pathways.

Heterologous expression, purification and structural features of native Dictyostelium discoideum dye-decolorizing peroxidase bound to a natively incorporated heme.

The Dictyostelium discoideum dye-decolorizing peroxidase (DdDyP) is a newly discovered peroxidase, which belongs to a unique class of heme peroxidase family that lacks homology to the known members of plant peroxidase superfamily. DdDyP catalyzes the H2O2-dependent oxidation of a wide-spectrum of substrates ranging from polycyclic dyes to lignin biomass, holding promise for potential industrial and biotechnological applications. To study the molecular mechanism of DdDyP, highly pure and functional protein with a natively incorporated heme is required, however, obtaining a functional DyP-type peroxidase with a natively bound heme is challenging and often requires addition of expensive biosynthesis precursors. Alternatively, a heme in vitro reconstitution approach followed by a chromatographic purification step to remove the excess heme is often used. Here, we show that expressing the DdDyP peroxidase in ×2 YT enriched medium at low temperature (20°C), without adding heme supplement or biosynthetic precursors, allows for a correct native incorporation of heme into the apo-protein, giving rise to a stable protein with a strong Soret peak at 402nm. Further, we crystallized and determined the native structure of DdDyP at a resolution of 1.95Å, which verifies the correct heme binding and its geometry. The structural analysis also reveals a binding of two water molecules at the distal site of heme plane bridging the catalytic residues (Arg239 and Asp149) of the GXXDG motif to the heme-Fe(III) via hydrogen bonds. Our results provide new insights into the geometry of native DdDyP active site and its implication on DyP catalysis.

The Purification of Heme Peroxidases from Escherichia coli Inclusion Bodies: A Success Story Shown by the Example of Horseradish Peroxidase.

In the following chapter a purification process for recombinant Horseradish peroxidase (HRP) produced in Escherichia coliis described. This enzyme is a secretory plant oxidoreductase belonging to the large peroxidase family III within the peroxidase-catalase superfamily of enzymes. It has high biotechnological significance, however, the isolation of the enzyme from its natural source, the horseradish root, has several shortcomings, which makes the development of a recombinant production strategy interesting. The presented protocol covers all process steps from isolation to the final chromatography step; the enzyme is solubilized from insoluble inclusion bodies, refolded and concentrated to yield a high purity enzyme preparation which is comparable to the commercially available plant-derived HRP. Moreover, we believe that this procedure can also be used to process other peroxidases of family II and III of the plant peroxidase superfamily, as they all share the samerelevant features like disulfide bonds and a heme group.

Insights into factors affecting lactoperoxidase conformation stability and enzymatic activity

Lactoperoxidase is a glycoprotein from the superfamily of mammalian haeme-containing peroxidases and has a great potential as a natural biopreservative in food and medicine due to its antimicrobial activity. For lactoperoxidase to be effectively utilised, its structural stability and consequently enzymatic activity must be maintained. Our measurements showed that two-state denaturation curve was observed at pH 3.5 and 4.0 in 100 mm buffers, whereas aggregation occurred at pH 5.0–7.0. Lactoperoxidase conformation stability was also affected by buffer type and buffer molar concentration and in presence of various ions (Ca2+, Mg2+, Na+). Furthermore, when lactoperoxidase was partially depleted of iron and calcium ions (de-LPO), thermal stability was lower in comparison with that of native lactoperoxidase and decreases in enzymatic activity were also observed. The enzymatic activity of lactoperoxidase was affected by the choice of buffer, pH, molar concentration of buffer, and temperature, but not by the presence of different ions.

New Genomic Regions Identified for Resistance to Spot Blotch and Terminal Heat Stress in an Interspecific Population of Triticum aestivum and T. spelta.

Wheat is one of the most widely grown and consumed food crops in the world. Spot blotch and terminal heat stress are the two significant constraints mainly in the Indo-Gangetic plains of South Asia. The study was undertaken using 185 recombinant lines (RILs) derived from the interspecific hybridization of 'Triticum aestivum (HUW234) × T. spelta (H+26)' to reveal genomic regions associated with tolerance to combined stress to spot blotch and terminal heat. Different physiological (NDVI, canopy temperature, leaf chlorophyll) and grain traits (TGW, grain size) were observed under stressed (spot blotch, terminal heat) and non-stressed environments. The mean maturity duration of RILs under combined stress was reduced by 12 days, whereas the normalized difference vegetation index (NDVI) was 46.03%. Similarly, the grain size was depleted under combined stress by 32.23% and thousand kernel weight (TKW) by 27.56% due to spot blotch and terminal heat stress, respectively. The genetic analysis using 6734 SNP markers identified 37 significant loci for the area under the disease progress curve (AUDPC) and NDVI. The genome-wide functional annotation of the SNP markers revealed gene functions such as plant chitinases, NB-ARC and NBS-LRR, and the peroxidase superfamily Cytochrome P450 have a positive role in the resistance through a hypersensitive response. Zinc finger domains, cysteine protease coding gene, F-box protein, ubiquitin, and associated proteins, play a substantial role in the combined stress of spot blotch and terminal heat in bread wheat, according to genomic domains ascribed to them. The study also highlights T. speltoides as a source of resistance to spot blotch and terminal heat tolerance.

Transformation of Reactive Blue 19 by a recombinant peroxidase DyP.

Wastewater containing recalcitrant dyes causes environmental problems. A new superfamily of heme-containing peroxidases, dye-decolorizing peroxidases (DyPs), has been found to decolorize different kinds of dyes, especial anthraquinone dyes efficiently. However, the mechanism of dyes degradation by DyPs has not been fully understood and the toxicity of dye degradation intermediates by DyPs catalysis to microbes is unclear. In this study, a purified recombinant Thermobifida fusca DyP (TfuDyP) in E. coli BL21(DE3) was used to treat Reactive Blue 19 (RB19), an anthraquinone dye. The reaction intermediates analyzed by ultra performance liquid chromatography/mass spectroscopy (UPLC-MS) indicated the initial site of TfuDyP attack on RB19. In addition, it was found that both RB19 and its incomplete degradation products inhibited the growth of Bacillus subtilis. These findings provided a novel understanding of DyPs catalysis to anthraquinone dyes.

Overexpression of MeH1.2 gene inhibited plant growth and increased branch root differentiation in transgenic cassava

AbstractCassava (Manihot esculenta) is an important tropical crop with extraordinary tolerance to drought stress. Under drought stress, the histone H1 (named MeH1.2) protein of cassava leaf is significantly induced. In this study, the MeH1.2 gene was cloned from cassava to investigate the role of MeH1.2 protein in transcriptional regulation during the development of a multicellular organism in vivo. The MeH1.2 protein was localized to the cell nucleus. The expression of MeH1.2 gene was induced by polyethylene glycol, abscisic acid, and drought stress. Transgenic cassava overexpressing MeH1.2 gene (HOE) was generated from wild type cv.60444 (WT) by Agrobacterium‐mediated transformation. The 60‐d‐old HOE seedlings were significantly smaller and weaker than the WT in Murashige and Skoog medium. Moreover, they exhibited yellow leaves and senescence, and the number of branch roots increased significantly. We found a total of 1,670 down‐regulated differentially expressed genes (DEGs) and 891 up‐regulated DEGs in HOE leaves as indicated by RNA‐seq. Many DEGs were associated with metabolism and stress response, such as amino acid metabolic genes, wound‐responsive family genes, and peroxidase superfamily genes. We hypothesized that MeH1.2 overexpression may retard the growth of HOE plants. The increased proline content and root/shoot ratio of HOE seedlings also supported this hypothesis. Our findings will provide an important basis for further research on the function of the H1.2 gene and its role in drought resistance of cassava.

Biallelic loss-of-function variations in PRDX3 cause cerebellar ataxia.

Peroxiredoxin 3 (PRDX3) belongs to a superfamily of peroxidases that function as protective antioxidant enzymes. Among the six isoforms (PRDX1-PRDX6), PRDX3 is the only protein exclusively localized to the mitochondria, which are the main source of reactive oxygen species. Excessive levels of reactive oxygen species are harmful to cells, inducing mitochondrial dysfunction, DNA damage, lipid and protein oxidation and ultimately apoptosis. Neuronal cell damage induced by oxidative stress has been associated with numerous neurodegenerative disorders including Alzheimer's and Parkinson's diseases. Leveraging the large aggregation of genomic ataxia datasets from the PREPARE (Preparing for Therapies in Autosomal Recessive Ataxias) network, we identified recessive mutations in PRDX3 as the genetic cause of cerebellar ataxia in five unrelated families, providing further evidence for oxidative stress in the pathogenesis of neurodegeneration. The clinical presentation of individuals with PRDX3 mutations consists of mild-to-moderate progressive cerebellar ataxia with concomitant hyper- and hypokinetic movement disorders, severe early-onset cerebellar atrophy, and in part olivary and brainstem degeneration. Patient fibroblasts showed a lack of PRDX3 protein, resulting in decreased glutathione peroxidase activity and decreased mitochondrial maximal respiratory capacity. Moreover, PRDX3 knockdown in cerebellar medulloblastoma cells resulted in significantly decreased cell viability, increased H2O2 levels and increased susceptibility to apoptosis triggered by reactive oxygen species. Pan-neuronal and pan-glial in vivo models of Drosophila revealed aberrant locomotor phenotypes and reduced survival times upon exposure to oxidative stress. Our findings reveal a central role for mitochondria and the implication of oxidative stress in PRDX3 disease pathogenesis and cerebellar vulnerability and suggest targets for future therapeutic approaches.

Structure of Yak Lactoperoxidase at 1.55Å Resolution.

Lactoperoxidase (LPO) is a heme containing oxido-reductase enzyme. It is secreted from mammary, salivary, lachrymal and mucosal glands. It catalyses the conversion of thiocyanate into hypothiocyanate and halides into hypohalides. LPO belongs to the superfamily of mammalian heme peroxidases which also includes myeloperoxidase (MPO), eosinophil peroxidase (EPO) and thyroid peroxidase (TPO). The heme prosthetic group is covalently linked in LPO through two ester bonds involving conserved residues Glu258 and Asp108. It was isolated from colostrum of yak (Bos grunniens), purified to homogeneity and crystallized using ammonium iodide as a precipitating agent. The crystals belonged to monoclinic space group P21 with cell dimensions of a = 53.91Å, b = 78.98Å, c = 67.82Å and β = 92.96°. The structure was determined at 1.55Å resolution. This is the first structure of LPO from yak. Also, this is the highest resolution structure of LPO determined so far from any source. The structure determination revealed that three segments (Ser1-Cys15), (Thr117-Asn138) and (Cys167-Leu175) were disordered and formed one surface of LPO structure. In the substrate binding site, the iodide ions were observed in three subsites which are formed by (1) heme moiety and residues, Gln105, Asp108, His109, Phe113, Arg255, Glu258, Phe380 and Phe381, (2) residues, Asn230, Lys232, Pro236, Cys248, Phe254, Phe381 and Pro424 and (3) residues, Ser198, Leu199 and Arg202. The structure determination also revealed that the side chain of Phe254 was disordered. It was observed to adopt two conformations in the structures of LPO.

Comparative Study of Degradation of Reactive Dyes and Decolorization and Detoxification in Aqueous Solution Applying DyP Peroxidases Isolated from Saccharomonospora viridis (SviDyP) and Thermobifida fusca (TfuDyp)

The wastewater that originates from the widespread usage of synthetic dyes in the industry have become a severe environmental problem. Several efforts have been made to develop new types of treatment which are capable of performing the degradation of the dyes from the environment. Within this scope, much attention has been drawn to enzymatic approaches, mainly the ones applying oxidative enzymes, such as peroxidases. A recently discovered superfamily of peroxidases, the so called dye-decolorizing peroxidases (DyPs) is a promising alternative to further improve the efficiency of these processes. In this work, two of these peroxidases (Saccharomonospora viridis (SviDyP) and Thermobifida fusca (TfuDyp)) were tested together with twelve different reactive dyes in order to evaluate the efficiency of degradation and decolorization, leading to good results. When applying the SviDyP enzyme in experiments carried out for 12 h in pH 3, the degradation efficiencies were above 80% for some dyes. The biodegradation efficiency data and cyclic voltammograms were recorded to obtain the redox potential of the chosen dyes and enzymes. In addition, an electrochemical biosensor was used to gauge the genotoxicity of the generated bioproducts. This analysis showed that bioproducts from dye degradation mostly present a lower degree of genotoxicity when compared to the control reactions.

A Stable Ferryl Porphyrin at the Active Site of Y463M BthA.

BthA is a diheme enzyme that is a member of the bacterial cytochrome c peroxidase superfamily, capable of generating a highly unusual Fe(IV)Fe(IV)═O oxidation state, known to be responsible for long-range oxidative chemistry in the enzyme MauG. Here, we show that installing a canonical Met ligand in lieu of the Tyr found at the heme of MauG associated with electron transfer, results in a construct that yields an unusually stable Fe(IV)═O porphyrin at the peroxidatic heme. This state is spontaneously formed at ambient conditions using either molecular O2 or H2O2. The resulting data illustrate how a ferryl iron, with unforeseen stability, may be achieved in biology.

Regulatory Phenomena in the Glutathione Peroxidase Superfamily.

Significance: The selenium-containing Glutathione peroxidases (GPxs)1-4 protect against oxidative challenge, inhibit inflammation and oxidant-induced regulated cell death. Recent Advances: GPx1 and GPx4 dampen phosphorylation cascades predominantly via prevention of inactivation of phosphatases by H2O2 or lipid hydroperoxides. GPx2 regulates the balance between regeneration and apoptotic cell shedding in the intestine. It inhibits inflammation-induced carcinogenesis in the gut but promotes growth of established cancers. GPx3 deficiency facilitates platelet aggregation likely via disinhibition of thromboxane biosynthesis. It is also considered a tumor suppressor. GPx4 is expressed in three different forms. The cytosolic form proved to inhibit interleukin-1-driven nuclear factor κB activation and leukotriene biosynthesis. Moreover, it is a key regulator of ferroptosis, because it reduces hydroperoxy groups of complex lipids and silences lipoxygenases. By alternate substrate use, the nuclear form contributes to chromatin compaction. Mitochondrial GPx4 forms the mitochondrial sheath of spermatozoa and, thus, guarantees male fertility. Out of the less characterized GPxs, the cysteine-containing GPx7 and GPx8 are unique in contributing to oxidative protein folding in the endoplasmic reticulum by reacting with protein isomerase as an alternate substrate. A yeast 2-Cysteine glutathione peroxidase equipped with CP and CR was reported to sense H2O2 for inducing an adaptive response. Critical Issues: Most of the findings compiled are derived from tissue culture and/or animal studies only. Their impact on human physiology is sometimes questionable. Future Directions: The expression of individual GPxs and GPx-dependent regulatory phenomena are to be further investigated, in particular in respect to human health.

Integrated physiologic, proteomic, and metabolomic analyses of Malus halliana adaptation to saline\u2013alkali stress

Saline–alkali stress is a severely adverse abiotic stress limiting plant growth. Malus halliana Koehne is an apple rootstock that is tolerant to saline–alkali stress. To understand the molecular mechanisms underlying the tolerance of M. halliana to saline–alkali stress, an integrated metabolomic and proteomic approach was used to analyze the plant pathways involved in the stress response of the plant and its regulatory mechanisms. A total of 179 differentially expressed proteins (DEPs) and 140 differentially expressed metabolites (DEMs) were identified. We found that two metabolite-related enzymes (PPD and PAO) were associated with senescence and involved in porphyrin and chlorophyll metabolism; six photosynthesis proteins (PSAH2, PSAK, PSBO2, PSBP1, and PSBQ2) were significantly upregulated, especially PSBO2, and could act as regulators of photosystem II (PSII) repair. Sucrose, acting as a signaling molecule, directly mediated the accumulation of D-phenylalanine, tryptophan, and alkaloid (vindoline and ecgonine) and the expression of proteins related to aspartate and glutamate (ASP3, ASN1, NIT4, and GLN1−1). These responses play a central role in maintaining osmotic balance and removing reactive oxygen species (ROS). In addition, sucrose signaling induced flavonoid biosynthesis by activating the expression of CYP75B1 to regulate the homeostasis of ROS and promoted auxin signaling by activating the expression of T31B5_170 to enhance the resistance of M. halliana to saline–alkali stress. The decrease in peroxidase superfamily protein (PER) and ALDH2C4 during lignin synthesis further triggered a plant saline–alkali response. Overall, this study provides an important starting point for improving saline–alkali tolerance in M. halliana via genetic engineering.

The Relationship of NADPH Oxidases and Heme Peroxidases: Fallin' in and Out.

Peroxidase enzymes can oxidize a multitude of substrates in diverse biological processes. According to the latest phylogenetic analysis, there are four major heme peroxidase superfamilies. In this review, we focus on certain members of the cyclooxygenase-peroxidase superfamily (also labeled as animal heme peroxidases) and their connection to specific NADPH oxidase enzymes which provide H2O2 for the one- and two-electron oxidation of various peroxidase substrates. The family of NADPH oxidases is a group of enzymes dedicated to the production of superoxide and hydrogen peroxide. There is a handful of known and important physiological functions where one of the seven known human NADPH oxidases plays an essential role. In most of these functions NADPH oxidases provide H2O2 for specific heme peroxidases and the concerted action of the two enzymes is indispensable for the accomplishment of the biological function. We discuss human and other metazoan examples of such cooperation between oxidases and peroxidases and analyze the biological importance of their functional interaction. We also review those oxidases and peroxidases where this kind of partnership has not been identified yet.

Downregulation of peroxiredoxin II suppresses the proliferation and metastasis of gastric cancer cells.

Peroxiredoxin (Prx) II is an imperative member of the superfamily of peroxidases. It serves an essential role in scavenging organic hydroperoxide and H2O2. It is involved in the development of various malignant tumors. In order to investigate the significance of Prx II expressions level in gastric cancer (GC), downregulation of Prx II was performed to investigate its role in the proliferation and migration of gastric adenocarcinoma cells. In GC cells and 45 GC specimens, the mRNA and protein expression levels of Prx II were determined using reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis, respectively. The Prx II expression profile in another 116 GC specimens was also detected with immunohistochemistry (IHC). The changes in the proliferation and migration of MKN45 and MGC-803 cells folllowing transfection with small interfering RNA (siRNA) were detected by cell counting kit (CCK)-8, western blot analysis, and Transwell migration and invasion assays. The results revealed that the expression of Prx II in GC tissues and GC cells were significantly upregulated compared with the normal control. There was a significant association between the expression level of Prx II and various factors, including tumor size, histological differentiation, the depth of invasion, the stage of tumor-node-metastasis (TNM) and lymph node metastasis in GC (P<0.05). Survival in patients with higher Prx II expression was significantly decreased compared with those with lower Prx II expression (P<0.01). Prx II, depth of invasion, lymph node metastasis and distant metastasis were identified as independent prognosis factors of GC (P<0.05). Knockdown of Prx II significantly suppressed the proliferation and the migration of GC cells. These experiments revealed that Prx II promotes the development of GC, affecting the survival of patients with GC.

Alginate-derived oligosaccharides promote water stress tolerance in cucumber (Cucumis sativus L.)

Marine alginate-derived oligosaccharides (ADOs) are prepared from degraded alginate. Our experiments were carried out to determine the mechanism of ADOs to improve resistance to water stress in cucumber (Cucumis sativus L.). We evaluated the effects of ADOs on physiological indices, photosynthesis, reactive oxygen species (ROS) levels, antioxidant enzyme activities, and relative expression levels of drought resistance genes. The growth of drought stressed cucumber decreased markedly. However, treatment with ADOs significantly improved the diameter, fresh weight, photosynthetic rate, transpiration rate, stomatal conductance, maximum quantum yield of photosystem II (Fv/Fm) and chlorophyll degradation; thus, reversing the effects of drought stress. Moreover, the antioxidant levels and ROS scavenging enzyme activities also increased in response to the ADOs. Additionally, the genes involved in abscisic aid (ABA) signaling and the drought stress response, such as superoxide dismutase [Cu-Zn] (CsSOD(Cu-Zn)), the peroxidase superfamily protein (CsPOD3), ABA deficient 2 (CsABA2), responsive to ABA 18 (CsRAB18), abscisic acid insensitive 5 (CsABI5), responsive to dehydration 22 (CsRD22), and responsive to dehydration 29A (CsRD29A) were upregulated by ADOs. The ABA content was also improved by ADOs. Our results suggest that ADOs induced the expression of some antioxidant enzyme synthetic genes involved in the ABA signaling pathway by stimulating ABA synthesis to improve the drought resistance capacity in cucumber.