Tomato T2 ribonuclease LE is involved in the response to pathogens

T2 ribonucleases (RNases) are RNA-degrading enzymes that function in various cellular processes, mostly via RNA metabolism. T2 RNase-encoding genes have been identified in various organisms, from bacteria to mammals, and are most diverse in plants. The existence of T2 RNase genes in almost every organism suggests an important biological function that has been conserved through evolution. In plants, T2 RNases are suggested to be involved in phosphate scavenging and recycling, and are implicated in defence responses to pathogens. We investigated the function of the tomato T2 RNase LE, known to be induced by phosphate deficiency and wounding. The possible involvement of LE in pathogen responses was examined. Expression analysis showed LE induction during fungal infection and by stimuli known to be associated with pathogen inoculation, including oxalic acid and hydrogen peroxide. Analysis of LE-suppressed transgenic tomato lines revealed higher susceptibility to oxalic acid, a cell death-inducing factor, compared to the wild type. This elevated sensitivity of LE-suppressed lines was evidenced by visual signs of necrosis, and increased ion leakage and reactive oxygen species levels, indicating acceleration of cell death. Challenge of the LE-suppressed lines with the necrotrophic pathogen Botrytis cinerea resulted in accelerated development of disease symptoms compared to the wild type, associated with suppressed expression of pathogenesis-related marker genes. The results suggest a role for plant endogenous T2 RNases in antifungal activity.     

Cross talk between gibberellin and cytokinin: the Arabidopsis GA response inhibitor SPINDLY plays a positive role in cytokinin signaling

SPINDLY (SPY) is a negative regulator of gibberellin (GA) responses; however, spy mutants exhibit various phenotypic alterations not found in GA-treated plants. Assaying for additional roles for SPY revealed that spy mutants are resistant to exogenously applied cytokinin. GA also repressed the effects of cytokinin, suggesting that there is cross talk between the two hormone-response pathways, which may involve SPY function. Two spy alleles showing severe (spy-4) and mild (spy-3) GA-associated phenotypes exhibited similar resistance to cytokinin, suggesting that SPY enhances cytokinin responses and inhibits GA signaling through distinct mechanisms. GA and spy repressed numerous cytokinin responses, from seedling development to senescence, indicating that cross talk occurs early in the cytokinin-signaling pathway. Because GA3 and spy-4 inhibited induction of the cytokinin primary-response gene, type-A Arabidopsis response regulator 5, SPY may interact with and modify elements from the phosphorelay cascade of the cytokinin signal transduction pathway. Cytokinin, on the other hand, had no effect on GA biosynthesis or responses. Our results demonstrate that SPY acts as both a repressor of GA responses and a positive regulator of cytokinin signaling. Hence, SPY may play a central role in the regulation of GA/cytokinin cross talk during plant development.     

NADH Binds and Stabilizes the 26S Proteasomes Independent of ATP

The 26S proteasome is the end point of the ubiquitin- and ATP-dependent degradation pathway. The 26S proteasome complex (26S PC) integrity and function has been shown to be highly dependent on ATP and its homolog nucleotides. We report here that the redox molecule NADH binds the 26S PC and is sufficient in maintaining 26S PC integrity even in the absence of ATP. Five of the 19S proteasome complex subunits contain a putative NADH binding motif (GxGxxG) including the AAA-ATPase subunit, Psmc1 (Rpt2). We demonstrate that recombinant Psmc1 binds NADH via the GxGxxG motif. Introducing the ΔGxGxxG Psmc1 mutant into cells results in reduced NADH-stabilized 26S proteasomes and decreased viability following redox stress induced by the mitochondrial inhibitor rotenone. The newly identified NADH binding of 26S proteasomes advances our understanding of the molecular mechanisms of protein degradation and highlights a new link between protein homeostasis and the cellular metabolic/redox state.     

Adaptive Resistance Mutations at Suprainhibitory Concentrations Independent of SOS Mutagenesis

Emergence of resistant bacteria during antimicrobial treatment is one of the most critical and universal health threats. It is known that several stress-induced mutagenesis and heteroresistance mechanisms can enhance microbial adaptation to antibiotics. Here, we demonstrate that the pathogen Bartonella can undergo stress-induced mutagenesis despite the fact it lacks error-prone polymerases, the rpoS gene and functional UV-induced mutagenesis. We demonstrate that Bartonella acquire de novo single mutations during rifampicin exposure at suprainhibitory concentrations at a much higher rate than expected from spontaneous fluctuations. This is while exhibiting a minimal heteroresistance capacity. The emerged resistant mutants acquired a single rpoB mutation, whereas no other mutations were found in their whole genome. Interestingly, the emergence of resistance in Bartonella occurred only during gradual exposure to the antibiotic, indicating that Bartonella sense and react to the changing environment. Using a mathematical model, we demonstrated that, to reproduce the experimental results, mutation rates should be transiently increased over 1,000-folds, and a larger population size or greater heteroresistance capacity is required. RNA expression analysis suggests that the increased mutation rate is due to downregulation of key DNA repair genes (mutS, mutY, and recA), associated with DNA breaks caused by massive prophage inductions. These results provide new evidence of the hazard of antibiotic overuse in medicine and agriculture.     

What do patients want? Surgical informed-consent and patient-centered care – An augmented model of information disclosure

The ideal moral standard for surgical informed-consent calls upon surgeons to carry out a disclosure dialogue with patients so they have as full as possible an understanding of the procedure before they sign the informed-consent form. This study is the first to empirically explore patient preferences regarding disclosure dialogue. Twelve Israelis who underwent life-saving surgeries participated in a narrative study. Three themes emerged from the analysis: objectification of patients, anxiety provoking processes and information, and lack of information that was essential for patients. Findings contribute to existing debates among surgeons regarding the scope and importance of some disclosure components. Analysis led to our formulation of an augmented subjective model of information disclosure that participants prefer, which extends beyond the immediate present of the surgery to the period after discharge, and until return to routine. Surgeons should be aware of patient preferences in disclosure, and gaps between perceptions of surgeons, and preferences and needs of patients.     

Cytosolic activity of SPINDLY implies the existence of a DELLA-independent gibberellin-response pathway

Specific plant developmental processes are modulated by cross-talk between gibberellin (GA)- and cytokinin-response pathways. Coordination of the two pathways involves the O-linked N -acetylglucosamine transferase SPINDLY (SPY) that suppresses GA signaling and promotes cytokinin responses in Arabidopsis. Although SPY is a nucleocytoplasmic protein, its site of action and targets are unknown. Several studies have suggested that SPY acts in the nucleus, where it modifies nuclear components such as the DELLA proteins to regulate signaling networks. Using chimeric GFP–SPY fused to a nuclear-export signal or to a glucocorticoid receptor, we show that cytosolic SPY promotes cytokinin responses and suppresses GA signaling. In contrast, nuclear-localized GFP–SPY failed to complement the spy mutation. To examine whether modulation of cytokinin activity by GA and spy is mediated by the nuclear DELLA proteins, cytokinin responses were studied in double and quadruple della mutants lacking the activities of REPRESSOR OF GA1-3 (RGA) and GA-INSENSITIVE (GAI) or RGA, GAI, RGA Like1 (RGL1) and RGL2. Unlike spy , the della mutants were cytokinin-sensitive. Moreover, when GA was applied to a cytokinin-treated quadruple della mutant it was able to suppress various cytokinin responses. These results suggest that cytosolic SPY and GA regulate cytokinin responses via a DELLA-independent pathway(s).