Search

The diversity of secondary compounds in the plant kingdom is huge. About 200,000 compounds are known, which are grouped into amines, non-protein amino acids, peptides, alkaloids, glucosinolates, cyanogenic glucosides, organic acids, terpenoids, quinones, polyacetylenes, and phenolics. The group of phenolic compounds consists of polyphenols, oligophenols and monophenols or simple phenolic compounds such as benzoic and cinnamic acids and their hydroxylated derivatives. Among the thousands of compounds present in ecological interactions, simple phenolic acids are the most abundant in soils, and many are described as allelochemicals. Given the physiological and biochemical importance of these compounds, we review their biosynthesis and metabolic actions in plants.

Pyruvate orthophosphate dikinase (PPDK) catalyzes the conversion of pyruvate to phosphoenolpyruvate, and its activity is essential to maintain photosynthetic CO2 assimilation in C4 plants. Thus, it has been described as a promising target for the development of selective herbicides for C4 weeds. We evaluated whether bromoacetic acid, a known PPDK inhibitor of Clostridium symbiosum, can also inhibit the PPDK of a C4 plant, maize, in in vitro experiments. We also evaluated the effects of foliar application of 0.5–10 mM bromoacetic acid on the leaf pyruvate content and gas exchange of maize plants. Bromoacetic acid reduced maize PPDK activity in vitro in a dose-dependent manner; the degree of inhibition ranged from 13.6 to 73.6% with 0.5–10 mM bromoacetic acid. The foliar application of bromoacetic acid increased the leaf pyruvate content, suggesting that this compound may also inhibit PPDK activity in vivo. As a consequence, impairments to the gas exchange parameters of maize plants were noted, with a marked reduction in the photosynthetic rate and an increase in the intercellular CO2 concentration. Despite the abrupt decrease in stomatal conductance and transpiration, the plants showed large dehydration and reduction of leaf fresh weight after 48 h of application to bromoacetic acid. Our data allow us to affirm that bromoacetic acid has a potent herbicidal effect on a C4 plant, maize; the most likely mechanism of action is the inhibition of PPDK.

Made available in DSpace on 2021-06-25T11:56:42Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-05-30 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Foreign Affairs, Trade, and Development Canada (DFATD) grant Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) CIHR ATPases belonging to the AAA+ superfamily are associated with diverse cellular activities and are mainly characterized by a nucleotide-binding domain (NBD) containing the Walker A and Walker B motifs. AAA+ proteins have a range of functions, from DNA replication to protein degradation. Rvbs, also known as RUVBLs, are AAA+ ATPases with one NBD domain and were described from human to yeast as participants of the R2TP (Rvb1-Rvb2-Tah1-Pih1) complex. Although essential for the assembly of multiprotein complexes-containing DNA and RNA, the protozoa Rvb orthologs are less studied. For the first time, this work describes the Rvbs from Leishmania major, one of the causative agents of Tegumentar leishmaniasis in human. Recombinant LmRUVBL1 and LmRUVBL2 his-tagged proteins were successfully purified and investigated using biophysical tools. LmRUVBL1 was able to form a well-folded elongated hexamer in solution, while LmRUVBL2 formed a large aggregate. However, the co-expression of LmRUVBL1 and LmRUVBL2 assembled the proteins into an elongated heterodimer in solution. Thermo-stability and fluorescence experiments indicated that the LmRUVBL1/2 heterodimer had ATPase activity in vitro. This is an interesting result because hexameric LmRUVBL1 alone had low ATPase activity. Additionally, using independent SL-RNAseq libraries, it was possible to show that both proteins are expressed in all L. major life stages. Specific antibodies obtained against LmRUVBLs identified the proteins in promastigotes and metacyclics cell extracts. Together, the results here presented are the first step towards the characterization of Leishmania Rvbs, and may contribute to the development of possible strategies to intervene against leishmaniasis, a neglected tropical disease of great medical importance. Univ Campinas UNICAMP, Inst Chem, BR-13083970 Campinas, SP, Brazil Sao Paulo State Univ, Biosci Inst, Dept Chem & Biol Sci, BR-18618689 Botucatu, SP, Brazil Univ Toronto, Dept Biochem, Toronto, ON M5G 1M1, Canada Univ Toronto, Dept Chem, Toronto, ON M5S 3H6, Canada Sao Paulo State Univ, Biosci Inst, Dept Chem & Biol Sci, BR-18618689 Botucatu, SP, Brazil FAPESP: 2012/50161-8 FAPESP: 2017/261315 FAPESP: 2013/10939-2 FAPESP: 2015/13521-4 FAPESP: 2014/25967-4 FAPESP: 2019/11496-3 CAPES: 99999.004913/2015-09 CIHR: PJT-173491

Titanium dioxide (TiO2) nanoparticles (NPs) are the most widely released nanomaterials in the environment and are considered an emerging contaminant. Although the phytotoxicity mechanism of TiO2 NPs on plants involves the elevated generation of reactive oxygen species (ROS), it is still not well established in soybean. Herein, we evaluated the effects of 250–1000 mg L−1 TiO2 NPs on seed germination, growth, content of ROS, lipid peroxidation, and activity of antioxidant enzymes in roots of soybean plants. Our data revealed that up to 1000 mg L−1 TiO2 NPs did not affect soybean seed germination. Transmission electron microscopy images and determinations of zeta potential and hydrodynamic diameter of a suspension of TiO2 NPs demonstrated that they form aggregates, favoring their adsorption to the root surface with consequent physical damage. The main deleterious effects noted on roots were reduced cell viability, reduced root hair number, striated aspect of root apexes, and reduced fresh and dry weights of roots. In disagreement with other studies, plant exposure to TiO2 NPs reduced the level of total ROS and lipid peroxidation, probably due to increased superoxide dismutase (SOD) activity. Altogether, our data suggest that the toxicity mechanism of TiO2 NPs on soybean roots involves physical damage resulting from their adsorption to the root surface, but not the generation of ROS.

Aluminum oxide (Al2O3) nanoparticles (NPs) are among the nanoparticles most used industrially, but their impacts on living organisms are widely unknown. We evaluated the effects of 50–1000 mg L−1 Al2O3 NPs on the growth, metabolism of lignin and its monomeric composition in soybean plants. Al2O3 NPs did not affect the length of roots and stems. However, at the microscopic level, Al2O3 NPs altered the root surface inducing the formation of cracks near to root apexes and damage to the root cap. The results suggest that Al2O3 NPs were internalized and accumulated into the cytosol and cell wall of roots, probably interacting with organelles such as mitochondria. At the metabolic level, Al2O3 NPs increased soluble and cell wall-bound peroxidase activities in roots and stems but reduced phenylalanine ammonia-lyase activity in stems. Increased lignin contents were also detected in roots and stems. The Al2O3 NPs increased the p-hydroxyphenyl monomer levels in stems but reduced them in roots. The total phenolic content increased in roots and stems; cell wall-esterified p-coumaric and ferulic acids increased in roots, while the content of p-coumaric acid decreased in stems. In roots, the content of ionic aluminum (Al+3) was extremely low, corresponding to 0.0000252% of the aluminum applied in the nanoparticulate form. This finding suggests that all adverse effects observed were due to the Al2O3 NPs only. Altogether, these findings suggest that the structure and properties of the soybean cell wall were altered by the Al2O3 NPs, probably to reduce its uptake and phytotoxicity.

The Hsp104/ClpB protein subfamily is unique in its ability to reactivate protein aggregates, which are implicated in several human and animal diseases. However, when compared to the unicellular yeast Hsp104 and bacterial ClpB disaggregases, the multicellular plant ortholog (Hsp101) is poorly studied. Here, we present a functional characterization of the Hsp101 (SHsp101) from the economically and agriculturally important sugarcane cultivar. Like Hsp104, SHsp101 has in vitro ATP/ATPγS dependent aggregate reactivation activity, confers induced thermotolerance in yeast and its ATPase activity is sensitive to guanidinium chloride. However, SHsp101 has distinct properties not previously reported for Hsp104, including higher ATPase activity at elevated temperatures and ATP independent intrinsic chaperone activity. Hence, SHsp101 has overlapping and distinct features from Hsp104.

The second messenger cyclic diguanylate monophosphate (c-di-GMP) is a central regulator of bacterial lifestyle, controlling several behaviors, including the switch between sessile and motile states. The c-di-GMP levels are controlled by the interplay between diguanylate cyclases (DGCs) and phosphodiesterases, which synthesize and hydrolyze this second messenger, respectively. These enzymes often contain additional domains that regulate activity via binding of small molecules, covalent modification, or protein–protein interactions. A major challenge remains to understand how DGC activity is regulated by these additional domains or interaction partners in specific signaling pathways. Here, we identified a pair of co-transcribed genes (xac2382 and xac2383) in the phytopathogenic, Gram-negative bacterium Xanthomonas citri subsp. citri (Xac), whose mutations resulted in opposing motility phenotypes. We show that the periplasmic cache domain of XAC2382, a membrane-associated DGC, interacts with XAC2383, a periplasmic binding protein, and we provide evidence that this interaction regulates XAC2382 DGC activity. Moreover, we solved the crystal structure of XAC2383 with different ligands, indicating a preference for negatively charged phosphate-containing compounds. We propose that XAC2383 acts as a periplasmic sensor that, upon binding its ligand, inhibits the DGC activity of XAC2382. Of note, we also found that this previously uncharacterized signal transduction system is present in several other bacterial phyla, including Gram-positive bacteria. Phylogenetic analysis of homologs of the XAC2382–XAC2383 pair supports several independent origins that created new combinations of XAC2382 homologs with a conserved periplasmic cache domain with different cytoplasmic output module architectures.

Tribolium castaneum (Coleoptera: Tenebrionidae) is a common pest of stored grains and byproducts and is normally infected by Gregarina cuneata (Apicomplexa: Eugregarinidae). The life cycle of this parasite includes the sporozoite, trophozoite, gamont, gametocyte, and oocyst stages, which occur between the epithelium and lumen of the host's midgut. This study aims to describe the morphofunctional alterations in the midgut and determine the esterase patterns in T. castaneum when parasitized by gregarines. To achieve this purpose, midguts of adult insects were isolated, processed, and analysed using light and electron microscopy. We determined total protein content, amylase activity, and the expression and related activities of the esterases by using polyacrylamide gel electrophoresis (PAGE). The midgut of T. castaneum is formed by digestive, regenerative, and endocrine cells. The effects of parasitism on the digestive cells are severe, because the gregarines remain attached to these cells to absorb all the nutrients they need throughout their development. In these cells, the most common alterations observed include expansion and fragmentation of the rough endoplasmic reticulum, development of the smooth endoplasmic reticulum, changes in mitochondrial cristae, cytoplasmic vacuolization, formation of myelin structures, spherites, large intercellular spaces, autophagic vesicles, expansion of the basal labyrinth, and cytoplasmic protrusions. Deposits of glycogen granules were also observed. Amylase activity was reduced in parasitized insects. Regenerative cells were found in disorganized crypts and did not differentiate into new cells, thus, compromising the restoration of the damaged epithelium. Though few morphological alterations were observed in the endocrine cells, results suggest that the synthesis and/or release of hormones might be impaired. Nine esterases (EST-1 to 9) were identified in the midgut of T. castaneum and were expressed in varying levels in response to parasitism. Two additional isoforms of esterases were exclusively identified in the parasitized insects. The results of this study suggest that gregarines alter the morphology and physiology of the midgut. The changes may result in nutritional depletion and the impairment of other physiological processes, such as reproduction and development of the host. Thus, further studies are needed to uncover the possibility of utilizing gregarines as biological controllers of the insect pest population.

ATPases associated with diverse cellular activities (AAA+) form a superfamily of proteins involved in a variety of functions and are characterized by the presence of an ATPase module containing two conserved motifs known as Walker A and Walker B. ClpB and Hsp104, chaperones that have disaggregase activities, are members of a subset of this superfamily, known as the AAA family, and are characterized by the presence of a second highly conserved motif, known as the second region of homology (SRH). Hsp104 and its homolog Hsp78 (78 kDa heat shock protein) are representatives of the Clp family in yeast. The structure and function of Hsp78 is reviewed and the possible existence of other homologs in metazoans is discussed.

Sitophilus zeamais Motschulsky, 1855 (Coleoptera: Curculionidae) is one of the most aggressive pests of stored grains, causing significant decrease in the nutritional quality of the grains and major losses in economic trade. The ability of this pest to damage grains and other products is directly related to the morphology of the alimentary canal. Considering the importance of the foregut in the digestion, this study aimed to describe the morphology of the crop and proventriculus in S. zeamais adults. The tissues were isolated, processed, and analyzed by using light microscopy, scanning electron microscopy, and transmission electron microscopy. The crop functioned as a storage organ, and its cells showed functional characteristics related to protein synthesis. High densities of prokaryotic microorganisms and spicules were found in the lumen. The proventriculus exhibited eight chitinous teeth, which were responsible for grinding and filtering food particles. This organ formed the stomodeal valve, which controlled the unidirectional flow of food through the alimentary canal. The alimentary canal is the primary surface of contact between the external environment and an insect's internal environment, and knowledge of its morphology is required to better understand the physiology of stored-grain insect pests.

In the present work, the gene xynB2, encoding a β-xylosidase II of the Glycoside Hydrolase 39 (GH39) family, of Caulobacter crescentus was cloned and successfully overexpressed in Escherichia coli DH10B. The recombinant protein (CcXynB2) was purified using nickel-Sepharose affinity chromatography, with a recovery yield of 75.5 %. CcXynB2 appeared as a single band of 60 kDa on a sodium dodecyl sulfate polyacrylamide gel and was recognized by a specific polyclonal antiserum. The predicted CcXynB2 protein showed a high homology with GH39 β-xylosidases of the genus Xanthomonas. CcXynB2 exhibited an optimal activity at 55 °C and a pH of 6. CcXynB2 displayed stability at pH values of 4.5-7.5 for 24 h and thermotolerance up to 50 °C. The K (M) and V (Max) values were 9.3 ± 0.45 mM and 402 ± 19 μmol min(-1) for ρ-nitrophenyl-β-D-xylopyranoside, respectively. The purified recombinant enzyme efficiently produced reducing sugars from birchwood xylan and sugarcane bagasse fibers pre-treated with a purified xylanase. As few bacterial GH39 family β-xylosidases have been characterized, this work provides a good contribution to this group of enzymes.

The process of folding is a seminal event in the life of a protein, as it is essential for proper protein function and therefore cell physiology. Inappropriate folding, or misfolding, can not only lead to loss of function, but also to the formation of protein aggregates, an insoluble association of polypeptides that harm cell physiology, either by themselves or in the process of formation. Several biological processes have evolved to prevent and eliminate the existence of non-functional and amyloidogenic aggregates, as they are associated with several human pathologies. Molecular chaperones and heat shock proteins are specialized in controlling the quality of the proteins in the cell, specifically by aiding proper folding, and dissolution and clearance of already formed protein aggregates. The latter is a function of disaggregases, mainly represented by the ClpB/Hsp104 subfamily of molecular chaperones, that are ubiquitous in all organisms but, surprisingly, have no orthologs in the cytosol of metazoan cells. This review aims to describe the characteristics of disaggregases and to discuss the function of yeast Hsp104, a disaggregase that is also involved in prion propagation and inheritance. O processo de enovelamento é um evento importante para o tempo de vida de uma proteína, uma vez que é essencial para a função adequada da proteína e, por conseguinte, para a fisiologia celular. Enovelamento inadequado, ou mau enovelamento, pode não só conduzir à perda de função, mas também para a formação de agregados de proteína, uma associação insolúvel de polipeptídos que prejudicam a fisiologia celular, quer por si próprios ou no processo de formação. Vários processos biológicos têm evoluído para prevenir e eliminar a existência de agregados não funcionais e amiloidogênicos, uma vez que estão associados com várias patologias humanas. Chaperonas moleculares e proteínas de choque térmico são especializadas no controle de qualidade de proteínas na célula, especificamente, auxiliando o enovelamento correto, dissolução e remoção de agregados de proteína já formados. Esta última é uma função das 'disagregases', representadas principalmente pela subfamília ClpB/ Hsp104 de chaperonas moleculares, que são comuns em todos os organismos, mas, surpreendentemente, não possuem ortólogos no citosol de células de metazoários. Esta revisão tem como objetivo descrever as características das disagregases e discutir a função da Hsp104 de levedura, uma disagregase que também está envolvida na propagação e herança de príons.