Cloning, in silico structural characterization and expression analysis of MfAtr4, an ABC transporter from the banana pathogen Mycosphaerella fijiensis

ABC transporters are membrane proteins that use the energy released from the hydrolysis of ATP to drive the transport of compounds across biological membranes. In some plants, pathogenic fungi ABC transporters play a role as virulence factors by mediating the export of plant defense compounds or fungal virulence factors. Mycosphaerella fijiensis, the causal agent of black Sigatoka disease in banana, is the main constraint for the banana industry worldwide. So far, little is known about molecular mechanism that it uses to infect the host. In this study, degenerated primers designed from fungal ABC transporters known to be involved in virulence were used to isolate homologs from M. fijiensis. Here, we reported the full cloning of MfAtr4 a putative ortholog of MgAtr4, an ABC transporter of the related Mycosphaerella graminicola with a function in virulence. Similarities and differences with its presumed ortholog MgAtr4 are described, and the putative function of MfAtr4 are discussed. Analysis of MfAtr4 gene expression in field banana samples exhibiting visible symptoms of black Sigatoka disease indicated a higher expression of MfAtr4 during the first symptomatic stages in comparison to the late necrotrophic phases, suggesting a role for MfAtr4 in the early stages of pathogenic development of M. fijiensis.Key words: ABC transporters, virulence factors, MgAtr4 ortholog, Mycosphaerella fijiensis, black Sigatoka, Musa sp

Ethnobotany and antimicrobial peptide from plants of Solanaceae family: An update and future prospect

The Solanaceae is an important plant family that has been playing an essential role in traditional medicine and human nutrition. Members of the Solanaceae are rich in bioactive metabolites and have been used by different tribes around the world for ages. Antimicrobial peptides (AMPs) from plants have drawn great interest in recent years and raised new hope for developing new antimicrobial agents for meeting the challenges of antibiotic resistance. This review aims to summarize the reported AMPs from plants of the Solanaceae with possible molecular mechanisms of action as well as to correlate their traditional uses with reported antimicrobial actions of the peptides. A systematic literature study was conducted using different databases until August 2019 based on the inclusion and exclusion criteria. According to literature, a variety of AMPs including defensins, protease inhibitor, lectins, thionin-like peptides, vicilin-like peptides, and snaking were isolated from plants of the Solanaceae and were involved in their defense mechanism. These peptides exhibited significant antibacterial, antifungal and antiviral activity against organisms for both plant and human host. Brugmansia, Capsicum, Datura, Nicotiana, Salpichora, Solanum, Petunia, and Withania are the most commonly studied genera for AMPs. Among these genera, Capsicum and the Solanum ranked top according to the total number of studies (35%–38% studies) for different AMPs. The mechanisms of action of the reported AMPs from Solanaceae was not any new rather similar to other reported AMPs including alteration of membrane potential and permeability, membrane pore formation, and cell aggregation. Whereas, induction of cell membrane permiabilization, inhibition of germination and alteration of hyphal growth were reported as mechanisms of antifungal activity. Plants of the Solanaceae have been used traditionally as antimicrobial, insecticidal, and antiinfectious agents, and as poisons. The reported AMPs from the Solanaceae are the products of chemical shields to protect plants from microorganisms and pests which unfold an obvious link with their traditional medicinal use. In summary, it is evident that AMPs from this family possess considerable antimicrobial activity against a wide range of bacterial and fungal pathogens and can be regarded as a potential source for lead molecules to develop new antimicrobial agents

Aluminum inhibits phosphatidic acid formation by blocking the phospholipase C pathway

Aluminum (Al(3+)) has been recognized as a main toxic factor in crop production in acid lands. Phosphatidic acid (PA) is emerging as an important lipid signaling molecule and has been implicated in various stress-signaling pathways in plants. In this paper, we focus on how PA generation is affected by Al(3+) using Coffea arabica suspension cells. We pre-labeled cells with [(32)P]orthophosphate ((32)Pi) and assayed for (32)P-PA formation in response to Al(3+). Treating cells for 15 min with either AlCl(3) or Al(NO(3))(3) inhibited the formation of PA. In order to test how Al(3+) affected PA signaling, we used the peptide mastoparan-7 (mas-7), which is known as a very potent stimulator of PA formation. The Al(3+) inhibited mas-7 induction of PA response, both before and after Al(3+) incubation. The PA involved in signaling is generated by two distinct phospholipid signaling pathways, via phospholipase D (PLD; EC: 3.1.4.4) or via Phospholipase C (PLC; EC: 3.1.4.3), and diacylglycerol kinase (DGK; EC 2.7.1.107). By labeling with (32)Pi for short periods of time, we found that PA formation was inhibited almost 30% when the cells were incubated with AlCl(3) suggesting the involvement of the PLC/DGK pathway. Incubation of cells with PLC inhibitor, U73122, affected PA formation, like AlCl(3) did. PLD in vivo activation by mas-7 was reduced by Al(3+). These results suggest that PA formation was prevented through the inhibition of the PLC activity, and it provides the first evidence for the role of Al toxicity on PA production

Las proteínas DING, una familia con intrigantes funciones celulares

La familia de las proteínas DING recibe este nombre porque en especies filogenéticamente distantes, dichos aminoácidos están altamente conservados en el extremo N-terminal. Sus integrantes tienen un peso molecular ~40 kDa, están relacionadas con el metabolismo del fosfato, son secretadas y en su mayoría poseen actividad enzimática de fosfatasa. Inicialmente se creyó que las proteínas DING eran exclusivas de Pseudomonas sp., pero ahora se sabe que están distribuidas en los diferentes reinos biológicos. El descubrimiento de esta familia se fundamentó en la secuenciación de aminoácidos debido a que, con excepción de Pseudomonas fluorescens, P. aeruginosa y algunos otros procariontes, los genes que las codifican no han sido encontrados en las bases de genes de los eucariontes cuyos genomas han sido ya secuenciados. Las proteínas DING tienen funciones biológicas controversiales y por ello están siendo objeto de intensa investigación. En células animales se les ha asociado con la aparición de enfermedades como el cáncer de mama y la caquexia, pero también con la protección contra la arterioesclerosis y la litiasis. En vegetales, algunas proteínas DING muestran propiedades citotóxicas sobre células tumorales o de inhibición de la replicación del virus VIH-1. La evidencia biológica muestra que el mecanismo de acción de las proteínas DING puede ser variado y el resultado contrastante. Dada la potencial aplicación terapéutica de estas proteínas, en esta revisión se describen los hallazgos que se han realizando en esta familia debido a que previamente a su aplicación es necesario entender los mecanismos que regulan sus funciones. Abstract The DING family of proteins called because in phylogenetically distant species, these amino acids are highly conserved in the N- terminal. The members have a molecular weight of ~40 kDa, are related to phosphate metabolism, are secreted and have mostly phosphatase enzymatic activity. Initially it was believed that DING proteins were unique to Pseudomonas sp., but is now known they are distributed in different biological kingdoms. The discovery of this family was based on the sequencing of amino acids because, with the exception of Pseudomonas fluorescens, P. aeruginosa and some other prokaryotes, the genes that encode them have not been found on the basis of genes of eukaryotes whose genomes have already been sequenced. The DING proteins have controversial biological functions and are therefore the subject of intense research. In animal cells they have been associated with the occurrence of diseases such as breast cancer and cachexia, but also to protection against atherosclerosis and gallstones. In plants, DING proteins exhibit some cytotoxic properties on tumor cells or on inhibiting the replication of HIV-1 virus. Biological evidence shows that the mechanism of action of the DING proteins can be varied and with contrasting results. Given the potential therapeutic application of these proteins, in this review, we described the findings that have been made in this family, since before its exploitation it is necessary to understand the mechanisms that regulate their functions. Keywords: Hypericum perforatum; Pseudomonas sp., DING proteins, anticancer drugs

Protein Profiling of Psittacanthus calyculatus during Mesquite Infection

Psittacanthus calyculatus is a hemiparasite mistletoe that represents an ecological problem due to the impacts caused to various tree species of ecological and commercial interest. Although the life cycle for the Psittacanthus genus is well established in the literature, the development stages and molecular mechanism implicated in P. calyculatus host infection are poorly understood. In this study, we used a manageable infestation of P. laevigata with P. calyculatus to clearly trace the infection, which allowed us to describe five phenological infective stages of mistletoe on host tree branches: mature seed (T1), holdfast formation (T2), haustorium activation (T3), haustorium penetration (T4), and haustorium connection (T5) with the host tree. Proteomic analyses revealed proteins with a different accumulation and cellular processes in infective stages. Activities of the cell wall-degrading enzymes cellulase and β-1,4-glucosidase were primarily active in haustorium development (T3), while xylanase, endo-glucanase, and peptidase were highly active in the haustorium penetration (T4) and xylem connection (T5). Patterns of auxins and cytokinin showed spatial concentrations in infective stages and moreover were involved in haustorium development. These results are the first evidence of proteins, cell wall-degrading enzymes, and phytohormones that are involved in early infection for the Psittacanthus genus, and thus represent a general infection mechanism for other mistletoe species. These results could help to understand the molecular dialogue in the establishment of P. calyculatus parasitism

Auxin-Cytokinin Cross Talk in Somatic Embryogenesis of <i>Coffea canephora</i>

Cytokinins (CK) are plant growth regulators involved in multiple physiological processes in plants. One less studied aspect is CK homeostasis (HM). The primary genes related to HM are involved in biosynthesis (IPT), degradation (CKX), and signaling (ARR). This paper demonstrates the effect of auxin (Aux) and CK and their cross talk in a Coffea canephora embryogenic system. The transcriptome and RT-qPCR suggest that Aux in pre-treatment represses biosynthesis, degradation, and signal CK genes. However, in the induction, there is an increase of genes implicated in the CK perception/signal, indicating perhaps, as in other species, Aux is repressing CK, and CK are inducing per se genes involved in its HM. This is reflected in the endogenous concentration of CK; pharmacology experiments helped study the effect of each plant growth regulator in our SE system. We conclude that the Aux–CK balance is crucial to directing somatic embryogenesis in C. canephora

Metabolic Responses of the Microalga <i>Neochloris oleoabundans</i> to Extracellular Self- and Nonself-DNA

Stressed organisms identify intracellular molecules released from damaged cells due to trauma or pathogen infection as components of the innate immune response. These molecules called DAMPs (Damage-Associated Molecular Patterns) are extracellular ATP, sugars, and extracellular DNA, among others. Animals and plants can recognize their own DNA applied externally (self-exDNA) as a DAMP with a high degree of specificity. However, little is known about the microalgae responses to damage when exposed to DAMPs and specifically to self-exDNAs. Here we compared the response of the oilseed microalgae Neochloris oleoabundans to self-exDNA, with the stress responses elicited by nonself-exDNA, methyl jasmonate (MeJA) and sodium bicarbonate (NaHCO3). We analyzed the peroxidase enzyme activity related to the production of reactive oxygen species (ROS), as well as the production of polyphenols, lipids, triacylglycerols, and phytohormones. After 5 min of addition, self-exDNA induced peroxidase enzyme activity higher than the other elicitors. Polyphenols and lipids were increased by self-exDNA at 48 and 24 h, respectively. Triacylglycerols were increased with all elicitors from addition and up to 48 h, except with nonself-exDNA. Regarding phytohormones, self-exDNA and MeJA increased gibberellic acid, isopentenyladenine, and benzylaminopurine at 24 h. Results show that Neochloris oleoabundans have self-exDNA specific responses