Pathogen-Induced Defense Signaling and Signal Crosstalk in Arabidopsis

Erwinia carotovora subsp. carotovora is a bacterial phytopathogen that causes soft rot in various agronomically important crop plants. A genetically specified resistance to E. carotovora has not been defined, and plant resistance to this pathogen is established through nonspecific activation of basal defense responses. This, together with the broad host range, makes this pathogen a good model for studying the activation of plant defenses. Production and secretion of plant cell wall-degrading enzymes (PCWDE) are central to the virulence of E. carotovora. It also possesses the type III secretion system (TTSS) utilized by many Gram-negative bacteria to secrete virulence- promoting effector proteins to plant cells. This study elucidated the role of E. carotovora HrpN (HrpNEcc), an effector protein secreted through TTSS, and the contribution of this protein in the virulence of E. carotovora. Treatment of plants with HrpNEcc was demonstrated to induce a hypersensitive response (HR) as well as resistance to E. carotovora. Resistance induced by HrpNEcc required both salicylic acid (SA)- and jasmonate/ethylene (JA/ET)-dependent defense signaling in Arabidopsis. Simultaneous treatment of Arabidopsis with HrpNEcc and PCWDE polygalacturonase PehA elicited accelerated and enhanced induction of defense genes but also increased production of superoxide and lesion formation. This demonstrates mutual amplification of defense signaling by these two virulence factors of E. carotovora. Identification of genes that are rapidly induced in response to a pathogen can provide novel information about the early events occurring in the plant defense response. CHLOROPHYLLASE 1 (AtCLH1) and EARLY RESPONSIVE TO DEHYDRATION 15 (ERD15) are both rapidly triggered by E. carotovora in Arabidopsis. Characterization of AtCLH1 encoding chlorophyll-degrading enzyme chlorophyllase indicated that it might have a role in chlorophyll degradation during plant tissue damage. Silencing of this gene resulted in increased accumulation of reactive oxygen species (ROS) in response to pathogen infection in a light-dependent manner. This led to enhanced SA-dependent defenses and resistance to E. carotovora. Moreover, crosstalk between different defense signaling pathways was observed; JA-dependent defenses and resistance to fungal pathogen Alternaria brassicicola were impaired, indicating antagonism between SA- and JA-dependent signaling. Characterization of ERD15 suggested that it is a novel, negative regulator of abscisic acid (ABA) signaling in Arabidopsis. Overexpression of ERD15 resulted in insensitivity to ABA and reduced tolerance of the plants to dehydration stress. However, simultaneously, the resistance of the plants to E. carotovora was enhanced. Silencing of ERD15 improved freezing and drought tolerance of transgenic plants. This, together with the reducing effect of ABA on seed germination, indicated hypersensitivity to this phytohormone. ERD15 was hypothesized to act as a capacitor that controls the appropriate activation of ABA responses in Arabidopsis.

Promiscuous restriction is a cellular defense strategy that confers fitness advantage to bacteria

Most bacterial genomes harbor restriction-modification systems, encoding a REase and its cognate MTase. On attack by a foreign DNA, the REase recognizes it as nonself and subjects it to restriction. Should REases be highly specific for targeting the invading foreign DNA? It is often considered to be the case. However, when bacteria harboring a promiscuous or high-fidelity variant of the REase were challenged with bacteriophages, fitness was maximal under conditions of catalytic promiscuity. We also delineate possible mechanisms by which the REase recognizes the chromosome as self at the noncanonical sites, thereby preventing lethal dsDNA breaks. This study provides a fundamental understanding of how bacteria exploit an existing defense system to gain fitness advantage during a host-parasite coevolutionary ``arms race.''

Discovery of genes involved in defense/immunity functions in a haemocytes cDNA library from Fenneropenaeus chinensis by ESTs annotation

Crustacean haemocytes play important roles in the host immune response including recognition, phagocytosis, melanization, cytotoxicity and inter-cellular signal communication. Expressed sequence tags (ESTs) analysis is proved to be an efficient approach not only for gene discovery, but also for gene expression profiles performance. In order to further understand the innate immune system and defense mechanisms of Chinese shrimp at molecular level, complementary DNA library is constructed from the haemocyte tissue of Fenneropenaeus chinensis. A total of 2371 cDNA clones are successfully sequenced and the average sequence length is 460 bp. About 50% are identified as orthologs of known genes from other organisms by BLASTx and BLASTn program. By sequences comparability and analysis, 34 important genes including 177 ESTs are identified that may be involved in defense or immune functions in shrimp based on the known knowledge. These genes are categorized into five categories according to their putative functions in shrimp immune system: 13 genes are different types of antimicrobial peptides (AMP, penaeidin, antilipopolysaccharide factor, etc.), and their proportion is about 3 8%; 11 genes belong to prophenoloxidase system (prophenoloxidase, serine proteinase, serine proteinase inhibitor, etc.), and their proportion is about 32%; five genes have high homology with clotting protein (lectin, transglutaminase, etc), and their proportion is about 15%; three genes may be involved in inter-cell signal communication (peroxinectin, integrin), and their proportion is about 9%; two genes have been identified to be chaperone proteins (Hsc70, thioredoxin peroxidase), and their proportion is about 6%. These EST sequences enrich our understanding of the immune genes of F chinensis and will help farther experimental research into immune factors and improve our knowledge of the immune mechanisms of shrimp. (c) 2007 Elsevier B.V. All rights reserved.

Chapter Three – Host–Parasite Interactions and the Evolution of Immune Defense

Parasites and pathogens are ubiquitous and act as an important selection pressure on animals. Here, drawing primarily on our own research, mostly on insects, we illustrate how host-parasite interactions have played a role in the evolution of a range of phenomena, including animal coloration, social behavior, foraging ecology, sexual selection, and life-history tradeoffs, as well as how variation in host behavior and ecology can drive variation in parasitism risk and host allocation of resources to immunity and other antiparasite defenses. We conclude by identifying key areas for future study.

Cross-resistance following artificial selection for increased defense against parasitoids in Drosophila melanogaster

An increase in resistance to one natural enemy may result in no correlated change, a positive correlated change, or a negative correlated change in the ability of the host or prey to resist other natural enemies. The type of specificity is important in understanding the evolutionary response to natural enemies and was studied here in a Drosaphila-parasitoid system. Drosophila melanogaster lines selected for increased larval resistance to the endoparasitoid wasps Asobara tabida or Leptopilina boulardi were exposed to attack by A. tabida, L. boulardi and Leptopilina heterotama at 15 degrees C, 20 degrees C, and 25 degrees C. In general, encapsulation ability increased with temperature, with the exception of the lines selected against L. boulardi, which showed the opposite trend. Lines selected against L, boulardi showed large increases in resistance against all three parasitoid species, and showed similar levels of defense against A. tabida to the lines selected against that parasitoid. In contrast, lines selected against A. tabida showed a large increase in resistance to A. tabida and generally to L. heterotoma, but displayed only a small change in their ability to survive attack by L. boulardi. Such asymmetries in correlated responses to selection for increased resistance to natural enemies may influence host-parasitoid community structure.

Interactions between the powdery mildew effector BEC1054 and barley proteins identify candidate host targets

There are over 500 candidate secreted effector proteins (CSEPs) or Blumeria effector candidates (BECs) specific to the barley powdery mildew pathogen Blumeria graminis f.sp. hordei. The CSEP/BEC proteins are expressed and predicted to be secreted by biotrophic feeding structures called haustoria. Eight BECs are required for the formation of functional haustoria. These include the RNase-like effector BEC1054 (synonym CSEP0064). In order to identify host proteins targeted by BEC1054, recombinant BEC1054 was expressed in E. coli, solubilized, and used in pull-down assays from barley protein extracts. Many putative interactors were identified by LC-MS/MS after subtraction of unspecific binders in negative controls. Therefore, a directed yeast-2-hybrid assay, developed to measure the effectiveness of the interactions in yeast, was used to validate putative interactors. We conclude that BEC1054 may target several host proteins, including a glutathione-S-transferase, a malate dehydrogenase, and a pathogen-related-5 protein isoform, indicating a possible role for BEC1054 in compromising well-known key players of defense and response to pathogens. In addition, BEC1054 interacts with an elongation factor 1 gamma. This study already suggests that BEC1054 plays a central role in barley powdery mildew virulence by acting at several levels.

Resource defense polygyny shifts to female defense polygyny over the course of the reproductive season of a Neotropical harvestman

Although studies classify the polygynous mating system of a given species into female defense polygyny (FDP) or resource defense polygyny (RDP), the boundary between these two categories is often slight. Males of some species may even shift between these two types of polygyny in response to temporal variation in social and environmental conditions. Here, we examine the mating system of the Neotropical harvestman Acutisoma proximum and, in order to assess if mate acquisition in males corresponds to FDP or RDP, we tested four contrasting predictions derived from the mating system theory. At the beginning of the reproductive season, males fight with other males for the possession of territories on the vegetation where females will later oviposit, as expected in RDP. Females present a marked preference for specific host plant species, and males establish their territories in areas where these host plants are specially abundant, which is also expected in RDP. Later in the reproductive season, males reduce their patrolling activity and focus on defending individual females that are ovipositing inside their territories, as what occurs in FDP. This is the first described case of an arachnid that exhibits a shift in mating system over the reproductive season, revealing that we should be cautious when defining the mating system of a species based on few observations concentrated in a brief period.

HOST-MEDIATED INDUCTION OF alpha-AMYLASES BY LARVAE OF THE MEXICAN BEAN WEEVIL Zabrotes subfasciatus (COLEOPTERA: CHRYSOMELIDAE: BRUCHINAE) IS IRREVERSIBLE AND OBSERVED FROM THE INITIATION OF THE FEEDING PERIOD

Larvae of Zabrotes subfasciatus secrete alpha-amylases that are insensitive to the alpha-amylase inhibitor found in seeds of Phaseolus vulgaris. By analyzing amylase activities during larval development on P. vulgaris, we detected activity of the constitutive amylase and the two inducible amylase isoforms at all stages. When larvae were transferred from the non alpha-amylase inhibitor containing seeds of Vigna unguiculata to P. vulgaris, the inducible alpha-amylases were expressed at the same level as in control larvae fed on P. vulgaris. Interestingly, when larvae were transferred from seeds of P. vulgaris to those of V. unguiculata, inducible alpha-amylases continued to be expressed at a level similar to that found in control larvae fed P. vulgaris continuously. When 10-day-old larvae were removed from seeds of V. unguiculata and transferred into capsules containing flour of P. vulgaris cotyledons, and thus maintained until completing 17 days ( age when the larvae stopped feeding), we could detect higher activity of the inducible alpha-amylases. However, when larvae of the same age were transferred from P. vulgaris into capsules containing flour of V. unguiculata, the inducible alpha-amylases remained up-regulated. These results suggest that the larvae of Z. subfasciatus have the ability to induce insensitive amylases early in their development. A short period of feeding on P. vulgaris cotyledon flour was sufficient to irreversibly induce the inducible alpha-amylase isoforms. Incubations of brush border membrane vesicles with the alpha-amylase inhibitor 1 from P. vulgaris suggest that the inhibitor is recognized by putative receptors found in the midgut microvillar membranes. (C) 2010 Wiley Periodicals, Inc.

Immunity of an Alternative Host Can Be Overcome by Higher Densities of Its Parasitoids Palmistichus elaeisis and Trichospilus diatraeae

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Mistletoes Play Different Roles in a Modular Host-Parasite Network

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Interaction of pathogenic fungi with host cells: Molecular and cellular approaches

This review provides an overview of several molecular and cellular approaches that are likely to supply insights into the host-fungus interaction. Fungi present intra- and/or extracellular host-parasite interfaces, the parasitism phenomenon being dependent on complementary surface molecules. The entry of the pathogen into the host cell is initiated by the fungus adhering to the cell surface, which generates an uptake signal that may induce its cytoplasmatic internalization. Furthermore, microbial pathogens use a variety of their surface molecules to bind to host extracellular matrix (ECM) components to establish an effective infection. on the other hand, integrins mediate the tight adhesion of cells to the ECM at sites referred to as focal adhesions and also play a role in cell signaling. The phosphorylation process is an important mechanism of cell signaling and regulation; it has been implicated recently in defense strategies against a variety of pathogens that alter host-signaling pathways in order to facilitate their invasion and survival within host cells. The study of signal transduction pathways in virulent fungi is especially important in view of their putative role in the regulation of pathogenicity. This review discusses fungal adherence, changes in cytoskeletal organization and signal transduction in relation to host-fungus interaction. (c) 2005 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.

Differential gene expression by Paracoccidioides brasiliensis in host interaction conditions: Representational difference analysis identifies candidate genes associated with fungal pathogenesis

Paracoccidioides brasiliensis causes infection by the host inhalation of airborne propagules of the mycelia phase of the fungus. These particles reach the lungs, and disseminate to virtually all organs. Here we describe the identification of differentially expressed genes in studies of host-fungus interaction. We analyzed two cDNA populations of P. brasiliensis, one obtained from infected animals and the other an admixture of fungus and human blood thus mimicking the hematologic events of the fungal dissemination. Our analysis identified transcripts differentially expressed. Genes related to iron acquisition, melanin synthesis and cell defense were specially upregulated in the mouse model of infection. The upregulated transcripts of yeast cells during incubation with human blood were those predominantly related to cell wall remodeling/synthesis. The expression pattern of genes was independently confirmed in host conditions, revealing their potential role in the infection process. This work can facilitate functional studies of novel regulated genes that may be important for the survival and growth strategies of P. brasiliensis in humans. (c) 2006 Elsevier Masson SAS. All rights reserved.

Helicobacter pylori infection: Host immune response, implications on gene expression and microRNAs

Helicobacter pylori (H. pylori) infection is the most common bacterial infection worldwide. Persistent infection of the gastric mucosa leads to inflammatory processes and may remain silent for decades or progress causing more severe diseases, such as gastric adenocarcinoma. The clinical consequences of H. pylori infection are determined by multiple factors, including host genetic predisposition, gene regulation, environmental factors and heterogeneity of H. pylori virulence factors. After decades of studies of this successful relationship between pathogen and human host, various mechanisms have been elucidated. In this review, we have made an introduction on H. pylori infection and its virulence factors, and focused mainly on modulation of host immune response triggered by bacteria, changes in the pattern of gene expression in H. pylori-infected gastric mucosa, with activation of gene transcription involved in defense mechanisms, inflammatory and immunological response, cell proliferation and apoptosis. We also highlighted the role of bacteria eradication on gene expression levels. In addition, we addressed the recent involvement of different microRNAs in precancerous lesions, gastric cancer, and inflammatory processes induced by bacteria. New discoveries in this field may allow a better understanding of the role of major factors involved in the pathogenic mechanisms of H. pylori. (C) 2014 Baishideng Publishing Group Co., Limited. All rights reserved.

The contribution of a murine CNS-TB model for the understanding of the host-pathogen interactions in the formation of granulomas

Central nervous system (CNS) tuberculosis (TB) is the most severe form of TB, characterized morphologically by brain granulomas and tuberculous meningitis (TBM). Experimental strategies for the study of the host-pathogen interaction through the analysis of granulomas and its intrinsic molecular mechanisms could provide new insights into the neuropathology of TB. To verify whether cerebellar mycobacterial infection induces the main features of the disease in human CNS and better understand the physiological mechanisms underlying the disease, we injected bacillus Calmette-Guerin (BCG) into the mouse cerebellum. BCG-induced CNS-TB is characterized by the formation of granulomas and TBM, a build up of bacterial loads in these lesions, and microglial recruitment into the lesion sites. In addition, there is an enhanced expression of signaling molecules such as nuclear factor-kappa B (NF-kappa B) and there is a presence of inducible nitric oxide synthase (iNOS) in the lesions and surrounding areas. This murine model of cerebellar CNS-TB was characterized by cellular and biochemical immune responses typically found in the human disease. This model could expand our knowledge about granulomas in TB infection of the cerebellum, and help characterize the physiological mechanisms involved with the progression of this serious illness that is responsible for killing millions people every year. (C) 2012 Elsevier B.V. All rights reserved.

Mistletoes Play Different Roles in a Modular Host-Parasite Network

Antagonistic interactions between host plants and mistletoes often form complex networks of interacting species. Adequate characterization of network organization requires a combination of qualitative and quantitative data. Therefore, we assessed the distribution of interactions between mistletoes and hosts in the Brazilian Pantanal and characterized the network structure in relation to nestedness and modularity. Interactions were highly asymmetric, with mistletoes presenting low host specificity (i.e., weak dependence) and with hosts being highly susceptible to mistletoe-specific infections. We found a non-nested and modular pattern of interactions, wherein each mistletoe species interacted with a particular set of host species. Psittacanthus spp. infected more species and individuals and also caused a high number of infections per individual, whereas the other mistletoes showed a more specialized pattern of infection. For this reason, Psittacanthus spp. were regarded as module hubs while the other mistletoe species showed a peripheral role. We hypothesize that this pattern is primarily the result of different seed dispersal systems. Although all mistletoe species in our study are bird dispersed, the frugivorous assemblage of Psittacanthus spp. is composed of a larger suite of birds, whereas Phoradendron are mainly dispersed by Euphonia species. The larger assemblage of bird species dispersing Psittacanthus seeds may also increase the number of hosts colonized and, consequently, its dominance in the study area. Nevertheless, other restrictions on the interactions among species, such as the differential capacity of mistletoe infections, defense strategies of hosts and habitat types, can also generate or enhance the observed pattern.