Search Results (7)

In this study, we optimized the initial concentrations of glycerol and (NH₄)₂SO₄ to enhance 1,3-propanediol (1,3-PDO) production by Clostridium beijerinckii strain Br21. A central composite rotational design (CCRD) was employed, varying glycerol concentrations between 158 and 441 mmol L⁻¹, and (NH₄)₂SO₄ concentrations between 4.4 and 25.8 mmol L⁻¹. The CCRD identified optimal conditions at 441.42 mmol L⁻¹ for glycerol and 25.8 mmol L⁻¹ for (NH₄)₂SO₄. The optimized medium resulted in a 112% increase in 1,3-PDO production compared to the original medium. Analysis of NH₄⁺ and SO₄²⁻ ions under optimal conditions revealed a higher consumption of NH₄⁺ than SO₄²⁻. Furthermore, a quantitative gene expression analysis revealed that while the expression of genes responsible for glycerol uptake and ATP sulfurylase remained unchanged, the expression of the dhaM gene, which encodes the oxidative phosphoenolpyruvate:dihydroxyacetone phosphotransferase, increased approximately 6-fold. In the reductive pathway, the expression of the dhaB1 gene, encoding glycerol dehydratase, and the dhaT gene, encoding 1,3-propanediol dehydrogenase, increased 2.5- and 5-fold, respectively. The upregulation of these genes supports the hypothesis that the optimal concentrations of glycerol and (NH₄)₂SO₄ enhance the 1,3-PDO production by C. beijerinckii Br21. Full article

Iron (Fe) deficiency is one of the most common micronutrient deficiencies limiting crop production globally, especially in arid regions due to decreased availability of Fe in alkaline soils. The ATP sulfurylase (ATPS) gene has been reported to participate in regulating various abiotic stresses. Transcriptome data and qRT-PCR analysis revealed that the ATP sulfurylase gene MhATPS1 was notably induced by Fe-deficiency stress. Consequently, MhATPS1 (103410737) was isolated from Malus halliana, and transgenic tobacco and transgenic apple calli were successfully obtained by genetic transformation. Compared with the wild type (WT), transgenic MhATPS1 lines (transgenic tobacco and transgenic apple calli) displayed stronger resistance to Fe-deficiency treatment. To be specific, transgenic plants exhibited better growth, accumulated more Fe²⁺ content, had higher ferric chelate reductase (FCR) activity, and a greater active oxygen scavenging capacity. Furthermore, transgenic MhATPS1 lines up-regulated the expression of Fe uptake genes under Fe-deficit stress. Additionally, MhATPS1 transgenic lines secreted more H⁺ content compared to the WT. In summary, these findings indicate that the MhATPS1 gene may play a positive role in Fe-deficiency stress in both tobacco and apple calli. Full article

Melatonin is a pleiotropic, nontoxic, regulatory biomolecule with various functions in abiotic stress tolerance. It reverses the adverse effect of heat stress on photosynthesis in plants and helps with sulfur (S) assimilation. Our research objective aimed to find the influence of melatonin, along with excess sulfur (2 mM SO₄²⁻), in reversing heat stress’s impacts on the photosynthetic ability of the mustard (Brassica juncea L.) cultivar SS2, a cultivar with low ATP-sulfurylase activity and a low sulfate transport index (STI). Further, we aimed to substantiate that the effect was a result of ethylene modulation. Melatonin in the presence of excess-S (S) increased S-assimilation and the STI by increasing the ATP-sulfurylase (ATP-S) and serine acetyltransferase (SAT) activity of SS2, and it enhanced the content of cysteine (Cys) and methionine (Met). Under heat stress, melatonin increased S-assimilation and diverted Cys towards the synthesis of more reduced glutathione (GSH), utilizing excess-S at the expense of less methionine and ethylene and resulting in plants’ reduced sensitivity to stress ethylene. The treatment with melatonin plus excess-S increased antioxidant enzyme activity, photosynthetic-S use efficiency (p-SUE), Rubisco activity, photosynthesis, and growth under heat stress. Further, plants receiving melatonin and excess-S in the presence of norbornadiene (NBD; an ethylene action inhibitor) under heat stress showed an inhibited STI and lower photosynthesis and growth. This suggested that ethylene was involved in the melatonin-mediated heat stress reversal effects on photosynthesis in plants. The interaction mechanism between melatonin and ethylene is still elusive. This study provides avenues to explore the melatonin–ethylene-S interaction for heat stress tolerance in plants. Full article

Sulfur plays a vital role in the primary and secondary metabolism of plants, and carries an important function in a large number of different compounds. Despite this importance, compared to other mineral nutrients, relatively little is known about sulfur sensing and signalling, as well as about the mechanisms controlling sulfur metabolism and homeostasis. Sulfur contents in plants vary largely not only among different species, but also among accessions of the same species. We previously used associative transcriptomics to identify several genes potentially controlling variation in sulfate content in the leaves of Brassica napus, including an OASC gene for mitochondrial O-acetylserine thiollyase (OAS-TL), an enzyme involved in cysteine synthesis. Here, we show that loss of OASC in Arabidopsis thaliana lowers not only sulfate, but also glutathione levels in the leaves. The reduced accumulation is caused by lower sulfate uptake and translocation to the shoots; however, the flux through the pathway is not affected. In addition, we identified a single nucleotide polymorphism in the OASC gene among A. thaliana accessions that is linked to variation in sulfate content. Both genetic and transgenic complementation confirmed that the exchange of arginine at position 81 for lysine in numerous accessions resulted in a less active OASC and a lower sulfate content in the leaves. The mitochondrial isoform of OAS-TL is, thus, after the ATPS1 isoform of sulfurylase and the APR2 form of APS reductase 2, the next metabolic enzyme with a role in regulation of sulfate content in Arabidopsis. Full article

Inorganic pyrophosphatase (PPase) is a ubiquitous enzyme that converts pyrophosphate (PP_i) to phosphate and, in this way, controls numerous biosynthetic reactions that produce PP_i as a byproduct. PPase activity is generally assayed by measuring the product of the hydrolysis reaction, phosphate. This reaction is reversible, allowing PP_i synthesis measurements and making PPase an excellent model enzyme for the study of phosphoanhydride bond formation. Here we summarize our long-time experience in measuring PPase activity and overview three types of the assay that are found most useful for (a) low-substrate continuous monitoring of PP_i hydrolysis, (b) continuous and fixed-time measurements of PP_i synthesis, and (c) high-throughput procedure for screening purposes. The assays are based on the color reactions between phosphomolybdic acid and triphenylmethane dyes or use a coupled ATP sulfurylase/luciferase enzyme assay. We also provide procedures to estimate initial velocity from the product formation curve and calculate the assay medium’s composition, whose components are involved in multiple equilibria. Full article

Global warming has two dangerous global consequences for agriculture: drought, due to water scarcity, and salinization, due to the prolonged use of water containing high concentrations of salts. Since the global climate is projected to continue to change over this century and beyond, choosing salt-tolerant plants could represent a potential paramount last resort for exploiting the secondary saline soils. Olive is considered moderately resistant to soil salinity as compared to other fruit trees, and in the present study, we investigated the influence of NaCl solutions (ranging from 0 to 200 mM) in a salt-tolerant (cv Canino) and two of its transgenic lines (Canino AT17-1 and Canino AT17-2), overexpressing tobacco osmotin gene, and in a salt-sensitive (Sirole) olive cultivar. After four weeks, most of the shoots of both Canino and Sirole plants showed stunted growth and ultimate leaf drop by exposure to salt-enriched media, contrary to transgenic lines, that did not show injuries and exhibited a normal growth rate. Malondialdehyde (MDA) content was also measured as an indicator of the lipid peroxidation level. To evaluate the role of the S assimilatory pathway in alleviating the adverse effects of salt stress, thiols levels as well as extractable activities of ATP sulfurylase (ATPS) and O-acetyl serine(thiol)lyase (OASTL), the first and the last enzyme of the S assimilation pathway, respectively, have been estimated. The results have clearly depicted that both transgenic lines overexpressing osmotin gene coped with increasing levels of NaCl by the induction of S metabolism, and particularly increase in OASTL activity closely paralleled changes of NaCl concentration. Linear correlation between salt stress and OASTL activity provides evidence that the S assimilation pathway plays a key role in adaptive response of olive plants under salt stress conditions. Full article

Mitochondrial matrix proteins synthesized in the cytosol often contain amino (N)-terminal targeting sequences (NTSs), or alternately internal targeting sequences (ITSs), which enable them to be properly translocated to the organelle. Such sequences are also required for proteins targeted to mitochondrion-related organelles (MROs) that are present in a few species of anaerobic eukaryotes. Similar to other MROs, the mitosomes of the human intestinal parasite Entamoeba histolytica are highly degenerate, because a majority of the components involved in various processes occurring in the canonical mitochondria are either missing or modified. As of yet, sulfate activation continues to be the only identified role of the relic mitochondria of Entamoeba. Mitosomes influence the parasitic nature of E. histolytica, as the downstream cytosolic products of sulfate activation have been reported to be essential in proliferation and encystation. Here, we investigated the position of the targeting sequence of one of the mitosomal matrix enzymes involved in the sulfate activation pathway, ATP sulfurylase (AS). We confirmed by immunofluorescence assay and subcellular fractionation that hemagluttinin (HA)-tagged EhAS was targeted to mitosomes. However, its ortholog in the δ-proteobacterium Desulfovibrio vulgaris, expressed as DvAS-HA in amoebic trophozoites, indicated cytosolic localization, suggesting a lack of recognizable mitosome targeting sequence in this protein. By expressing chimeric proteins containing swapped sequences between EhAS and DvAS in amoebic cells, we identified the ITSs responsible for mitosome targeting of EhAS. This observation is similar to other parasitic protozoans that harbor MROs, suggesting a convergent feature among various MROs in favoring ITS for the recognition and translocation of targeted proteins. Full article