Exclusive rewards in mutualisms: ant proteases and plant protease inhibitors create a lock–key system to protect Acacia food bodies from exploitation

Myrmecophytic Acacia species produce food bodies (FBs) to nourish ants of the Pseudomyrmex ferrugineus group, with which they live in an obligate mutualism. We investigated how the FBs are protected from exploiting nonmutualists. Two‐dimensional gel electrophoresis of the FB proteomes and consecutive protein sequencing indicated the presence of several Kunitz‐type protease inhibitors (PIs). PIs extracted from Acacia FBs were biologically active, as they effectively reduced the trypsin‐like and elastase‐like proteolytic activity in the guts of seed‐feeding beetles (Prostephanus truncatus and Zabrotes subfasciatus), which were used as nonadapted herbivores representing potential exploiters. By contrast, the legitimate mutualistic consumers maintained high proteolytic activity dominated by chymotrypsin 1, which was insensitive to the FB PIs. Larvae of an exploiter ant (Pseudomyrmex gracilis) taken from Acacia hosts exhibited lower overall proteolytic activity than the mutualists. The proteases of this exploiter exhibited mainly elastase‐like and to a lower degree chymotrypsin 1‐like activity. We conclude that the mutualist ants possess specifically those proteases that are least sensitive to the PIs in their specific food source, whereas the congeneric exploiter ant appears partly, but not completely, adapted to consume Acacia FBs. By contrast, any consumption of the FBs by nonadapted exploiters would effectively inhibit their digestive capacities. We suggest that the term ‘exclusive rewards’ can be used to describe situations similar to the one that has evolved in myrmecophytic Acacia species, which reward mutualists with FBs but safeguard the reward from exploitation by generalists by making the FBs difficult for the nonadapted consumer to use.     

Anaerobic Elemental Sulfur Reduction by Fungus Fusarium oxysporum

Reduction of inorganic sulfur compounds by the fungus Fusarium oxysporum was examined. When transferred from a normoxic to an anoxic environment, F. oxysporum reduced elemental sulfur to hydrogen sulfide (H₂S). This reaction accompanied fungal growth and oxidation of the carbon source (ethanol) to acetate. Over 2-fold more of H₂S than of acetate was produced, which is the theoretical correlation for the oxidation of ethanol to acetate. NADH-dependent sulfur reductase (SR) activity was detected in cell-free extracts of the H₂S-producing fungus, and was found to be up-regulated under the anaerobic conditions. On the other hands both O₂ consumption by the cells and cytochrome c oxidase activity by the crude mitochondrial fractions decreased. These results indicate that H₂S production involving SR was due to a novel dissimilation mechanism of F. oxysporum, and that the fungus adapts to anaerobic conditions by replacing the energy-producing mechanism of O₂ respiration with sulfur reduction.     

Oxalic acid-induced resistance to Rhizoctonia solani in rice is associated with induction of phenolics,peroxidase and pathogenesis-related proteins

Abstract

Oxalic acid (1 mM) when applied as a foliar spray to rice plants induced resistance to challenge infection with Rhizoctonia solani, the rice sheath blight pathogen. Maximum reduction in sheath blight incidence was observed when the plants were sprayed with oxalic acid three days before inoculation with the fungus. The biochemical alterations in rice plants treated with oxalic acid was also investigated. When rice plants were treated with oxalic acid, a two-fold increase in phenolic content in leaf sheaths was recorded three days after treatment. Phenylalanine ammonia-lyase and peroxidase activities increased significantly starting from two days after treatment. Peroxidase (PO) isozyme analysis indicated that PO-3 and PO-4 were induced two days after treatment with oxalic acid. Western blot analysis revealed that two chitinases (28 and 35 kDa) and two β-1,3-glucanases (30 and 32 kDa) were strongly induced in rice sheaths four to six days after treatment with oxalic acid. Immunoblot analysis of protein extracts from oxalic acid-treated plants demonstrated the induction of a 23 kDa thaumatin-like protein (TLP) cross-reacting with bean TLP antibody. These results suggest that the enhanced activities of defense enzymes and defense-related compounds in oxalic acid-treated rice plants may contribute to resistance against R. solani.     

Abundance of RNase4 and RNase5 mRNA and protein in host defence related tissues and secretions in cattle

Members of the RNaseA family are present in various tissues and secretions but their function is not well understood. Some of the RNases are proposed to participate in host defence. RNase4 and RNase5 are present in cows' milk and have antimicrobial activity. However, their presence in many tissues and secretions has not been characterised. We hypothesised that these two RNases are present in a range of tissues and secretions where they could contribute to host defence. We therefore, determined the relative abundance of RNase4 and RNase5 mRNA as well as protein levels in a range of host defence related and other tissues as well as a range of secretions in cattle, using real time PCR and western blotting. The two RNases were found to be expressed in liver, lung, pancreas, mammary gland, placenta, endometrium, small intestine, seminal vesicle, salivary gland, kidney, spleen, lymph node, skin as well as testes. Corresponding proteins were also detected in many of the above tissues, as well as in seminal fluid, mammary secretions and saliva. This study provides evidence for the presence of RNase4 and RNase5 in a range of tissues and secretions, as well as some major organs in cattle. The data are consistent with the idea that these proteins could contribute to host defence in these locations. This work contributes to growing body of data suggesting that these proteins contribute to the physiology of the organism in a more complex way than acting merely as digestive enzymes.     

Toxin depletion has no effect on antipredator responses in common toad (Bufo bufo) tadpoles

Antipredator responses often involve changes in several phenotypic traits and these changes interactively influence fitness. However, gaining insight into how the overall fitness effect of the overall response comes about is notoriously difficult. One promising avenue is to manipulate a single defensive trait and observe how that modifies fitness as well as the expression of other inducible responses. In chemically‐defended animals, toxins are likely to be costly to produce but it is still unknown how their depletion influences other characteristics. In the present study, we artificially depleted bufadienolide toxin stores in common toad (Bufo bufo) tadpoles, and assessed the effect of this with respect to the interaction with predator presence and limited food availability. We found that toxin depletion in tadpoles did not significantly affect any of the measured life‐history traits. Tadpoles in the predator treatment exhibited an elevated development rate, although this was only apparent when food availability was limited. Also, body mass at metamorphosis was lower in tadpoles exposed to chemical cues indicating a predation threat and when food availability was limited. These results provide evidence that, in larval common toads, the expression of inducible defences may incur fitness costs, whereas chemical defences are either expressed constitutively or, if inducible, elevated toxin production has negligible costs.     

SOBIR1 requires the GxxxG dimerization motif in its transmembrane domain to form constitutive complexes with receptor‐like proteins

Receptor‐like proteins (RLPs), forming an important group of transmembrane receptors in plants, play roles in development and immunity. RLPs contain extracellular leucine‐rich repeats (LRRs) and, in contrast with receptor‐like kinases (RLKs), lack a cytoplasmic kinase required for the initiation of downstream signalling. Recent studies have revealed that the RLK SOBIR1/EVR (SUPPRESSOR OF BIR1‐1/EVERSHED) specifically interacts with RLPs. SOBIR1 stabilizes RLPs and is required for their function. However, the mechanism by which SOBIR1 associates with RLPs and regulates RLP function remains unknown. The Cf immune receptors of tomato (Solanum lycopersicum), mediating resistance to the fungus Cladosporium fulvum, are RLPs that also interact with SOBIR1. Here, we show that both the LRR and kinase domain of SOBIR1 are dispensable for association with the RLP Cf‐4, whereas the highly conserved GxxxGxxxG motif present in the transmembrane domain of SOBIR1 is essential for its interaction with Cf‐4 and additional RLPs. Complementation assays in Nicotiana benthamiana, in which endogenous SOBIR1 levels were knocked down by virus‐induced gene silencing, showed that the LRR domain as well as the kinase activity of SOBIR1 are required for the Cf‐4/Avr4‐triggered hypersensitive response (HR). In contrast, the LRRs and kinase activity of SOBIR1 are not required for facilitation of Cf‐4 accumulation. Together, these results suggest that, in addition to being a stabilizing scaffold for RLPs, SOBIR1 is also required for the initiation of downstream signalling through its kinase domain.