Plant proteases in the control of the hypersensitive response

The hypersensitive response (HR) is a plant defence reaction triggered by activation of immune receptors upon pathogen recognition. It results in rapid cell death at the attempted invasion site, confining the pathogen and sending signals to distal parts of the plant that can in turn activate defences for subsequent attacks. HR cell death is a highly controlled phenomenon, requiring the concerted action of diverse plant proteases and regulatory mechanisms to keep it efficient yet confined. Research in the last decade has significantly contributed to a better understanding of the mechanisms leading to HR, although our knowledge about the pathways that regulate this form of programmed cell death (PCD) still remains incomplete. In this review, we explore current knowledge of plant proteases as HR regulators. Proteases are key regulatory enzymes that not only serve degradative purposes, but also have very important signalling roles. In animals, caspases have been shown to be the major regulators and executioners of PCD. Plants do not have caspases, and instead PCD is carried out by the activities of caspase-like and other protease belonging to different protease classes. We summarise the mechanistic roles of plant proteases whose roles in HR regulation are relatively well understood, which includes members of the cysteine, threonine, and serine protease families

Autophagy as an emerging arena for plant-pathogen interactions

Altres ajuts: CERCA Programme/Generalitat de CatalunyaAutophagy is a highly conserved degradation and recycling process that controls cellular homeostasis, stress adaptation, and programmed cell death in eukaryotes. Emerging evidence indicates that autophagy is a key regulator of plant innate immunity and contributes with both pro-death and pro-survival functions to antimicrobial defences, depending on the pathogenic lifestyle. In turn, several pathogens have co-opted and evolved strategies to manipulate host autophagy pathways to the benefit of infection, while some eukaryotic microbes require their own autophagy machinery for successful pathogenesis. In this review, we present and discuss recent advances that exemplify the important role of pro- and antimicrobial autophagy in plant-pathogen interactions

Dying two deaths - programmed cell death regulation in development and disease

Programmed cell death (PCD) is a fundamental cellular process that has adopted a plethora of vital functions in multicellular organisms. In plants, PCD processes are elicited as an inherent part of regular development in specific cell types or tissues, but can also be triggered by biotic and abiotic stresses. Although over the last years we have seen progress in our understanding of the molecular regulation of different plant PCD processes, it is still unclear whether a common core machinery exists that controls cell death in development and disease. In this review, we discuss recent advances in the field, comparing some aspects of the molecular regulation controlling developmental and pathogen-triggered PCD in plants

Enhancing Localized Pesticide Action through Plant Foliage by Silver-Cellulose Hybrid Patches

Efficacy and efficiency of pesticide application in the field through the foliage still face many challenges. There exists a mismatch between the hydrophobic character of the leaf and the active molecule, low dispersion of the pesticides on the leaves' surface, runoff loss, and rolling down of the active molecules to the field, decreasing their efficacy and increasing their accumulation to the soil. We produced bacterial cellulose-silver nanoparticles (BC-AgNPs) hybrid patches by in situ thermal reduction under microwave irradiation in a scalable manner and obtaining AgNPs strongly anchored to the BC. Those hybrids increase the interaction of the pesticide (AgNPs) with the foliage and avoids runoff loss and rolling down of the nanoparticles. The positive antibacterial and antifungal properties were assessed in vitro against the bacteria Escherichia coli and two agro-economically relevant pathogens: the bacterium Pseudomonas syringae and the fungus Botrytis cinerea. We showed in vivo inhibition of the infection in Nicotiana benthamiana and tomato leaves, as proven by the suppression of the expression of defense molecular markers and reactive oxygen species production. The hydrogel-like character of the bacterial cellulose matrix increases the adherence to the foliage of the patches

Transcriptome responses to Ralstonia solanacearum infection in the roots of the wild potato Solanum commersonii

Background: Solanum commersonii is a wild potato species that exhibits high tolerance to both biotic and abiotic stresses and has been used as a source of genes for introgression into cultivated potato. Among the interesting features of S. commersonii is resistance to the bacterial wilt caused by Ralstonia solanacearum, one of the most devastating bacterial diseases of crops. - Results: In this study, we used deep sequencing of S. commersonii RNA (RNA-seq) to analyze the below-ground plant transcriptional responses to R. solanacearum. While a majority of S. commersonii RNA-seq reads could be aligned to the Solanum tuberosum Group Phureja DM reference genome sequence, we identified 2,978 S. commersonii novel transcripts through assembly of unaligned S. commersonii RNA-seq reads. We also used RNA-seq to study gene expression in pathogen-challenged roots of S. commersonii accessions resistant (F118) and susceptible (F97) to the pathogen. Expression profiles obtained from read mapping to the S. tuberosum reference genome and the S. commersonii novel transcripts revealed a differential response to the pathogen in the two accessions, with 221 (F118) and 644 (F97) differentially expressed genes including S. commersonii novel transcripts in the resistant and susceptible genotypes. Interestingly, 22.6% of the F118 and 12.8% of the F97 differentially expressed genes had been previously identified as responsive to biotic stresses and half of those up-regulated in both accessions had been involved in plant pathogen responses. Finally, we compared two different methods to eliminate ribosomal RNA from the plant RNA samples in order to allow dual mapping of RNAseq reads to the host and pathogen genomes and provide insights on the advantages and limitations of each technique. - Conclusions: Our work catalogues the S. commersonii transcriptome and strengthens the notion that this species encodes specific genes that are differentially expressed to respond to bacterial wilt. In addition, a high proportion of S. commersonii-specific transcripts were altered by R. solanacearum only in F118 accession, while phythormone-related genes were highly induced in F97, suggesting a markedly different response to the pathogen in the two plant accessions studied

Type III secretion inhibitors for the management of bacterial plant diseases

Altres ajuts: COST Action SUSTAIN (FA1208) from the European Union.The identification of chemical compounds that prevent and combat bacterial diseases is fundamental for crop production. Bacterial virulence inhibitors are a promising alternative to classical control treatments, because they have a low environmental impact and are less likely to generate bacterial resistance. The major virulence determinant of most animal and plant bacterial pathogens is the type III secretion system (T3SS). In this work, we screened nine plant extracts and 12 isolated compounds-including molecules effective against human pathogens-for their capacity to inhibit the T3SS of plant pathogens and for their applicability as virulence inhibitors for crop protection. The screen was performed using a luminescent reporter system developed in the model pathogenic bacterium Ralstonia solanacearum. Five synthetic molecules, one natural product and two plant extracts were found to down-regulate T3SS transcription, most through the inhibition of the regulator hrpB. In addition, for three of the molecules, corresponding to salicylidene acylhydrazide derivatives, the inhibitory effect caused a dramatic decrease in the secretion capacity, which was translated into impaired plant responses. These candidate virulence inhibitors were then tested for their ability to protect plants. We demonstrated that salicylidene acylhydrazides can limit R. solanacearum multiplication in planta and protect tomato plants from bacterial speck caused by Pseudomonas syringae pv. tomato. Our work validates the efficiency of transcription reporters to discover compounds or natural product extracts that can be potentially applied to prevent bacterial plant disease

Protease activities triggered by Ralstonia solanacearum infection in susceptible and tolerant tomato lines

Altres ajuts: CERCA Programme/Generalitat de CatalunyaActivity-based protein profiling (ABPP) is a powerful proteomic technique to display protein activities in a proteome. It is based on the use of small molecular probes that react with the active site of proteins in an activity-dependent manner. We used ABPP to dissect the protein activity changes that occur in the intercellular spaces of tolerant (Hawaii 7996) and susceptible (Marmande) tomato plants in response to R. solanacearum, the causing agent of bacterial wilt, one of the most destructive bacterial diseases in plants. The intercellular space -or apoplast- is the first battlefield where the plant faces R. solanacearum. Here, we explore the possibility that the limited R. solanacearum colonization reported in the apoplast of tolerant tomato is partly determined by its active proteome. Our work reveals specific activation of papain-like cysteine proteases (PLCPs) and serine hydrolases (SHs) in the leaf apoplast of the tolerant tomato Hawaii 7996 on R. solanacearum infection. The P69 family members P69C and P69F, and an unannotated lipase (Solyc02g077110.2.1), were found to be post-translationally activated. In addition, protein network analysis showed that deeper changes in network topology take place in the susceptible tomato variety, suggesting that the tolerant cultivar might be more prepared to face R. solanacearum in its basal state. Altogether this work identifies significant changes in the activity of 4 PLCPs and 27 SHs in the tomato leaf apoplast in response to R. solanacearum, most of which are yet to be characterized. Our findings denote the importance of novel proteomic approaches such as ABPP to provide new insights on old and elusive questions regarding the molecular basis of resistance to R. solanacearum

Dynamic expression of Ralstonia solanacearum virulence factors and metabolism-controlling genes during plant infection

Altres ajuts: CERCA Programme/Generalitat de Catalunya. P. Sebastià received the support of a fellowship (code is LCF/BQ/IN17/11620004) from la Caixa Foundation (identifier [ID] 100010434)Background: Ralstonia solanacearum is the causal agent of bacterial wilt, a devastating plant disease responsible for serious economic losses especially on potato, tomato, and other solanaceous plant species in temperate countries. In R. solanacearum, gene expression analysis has been key to unravel many virulence determinants as well as their regulatory networks. However, most of these assays have been performed using either bacteria grown in minimal medium or in planta, after symptom onset, which occurs at late stages of colonization. Thus, little is known about the genetic program that coordinates virulence gene expression and metabolic adaptation along the different stages of plant infection by R. solanacearum. Results: We performed an RNA-sequencing analysis of the transcriptome of bacteria recovered from potato apoplast and from the xylem of asymptomatic or wilted potato plants, which correspond to three different conditions (Apoplast, Early and Late xylem). Our results show dynamic expression of metabolism-controlling genes and virulence factors during parasitic growth inside the plant. Flagellar motility genes were especially up-regulated in the apoplast and twitching motility genes showed a more sustained expression in planta regardless of the condition. Xylem-induced genes included virulence genes, such as the type III secretion system (T3SS) and most of its related effectors and nitrogen utilisation genes. The upstream regulators of the T3SS were exclusively up-regulated in the apoplast, preceding the induction of their downstream targets. Finally, a large subset of genes involved in central metabolism was exclusively down-regulated in the xylem at late infection stages. Conclusions: This is the first report describing R. solanacearum dynamic transcriptional changes within the plant during infection. Our data define four main genetic programmes that define gene pathogen physiology during plant colonisation. The described expression of virulence genes, which might reflect bacterial states in different infection stages, provides key information on the R. solanacearum potato infection process

Induced ligno-suberin vascular coating and tyramine-derived hydroxycinnamic acid amides restrict Ralstonia solanacearum colonization in resistant tomato

19 páginas.- 9 figuras.- referenciasTomato varieties resistant to the bacterial wilt pathogen Ralstonia solanacearum have the ability to restrict bacterial movement in the plant. Inducible vascular cell wall reinforcements seem to play a key role in confining R. solanacearum into the xylem vasculature of resistant tomato. However, the type of compounds involved in such vascular physico-chemical barriers remain understudied, while being a key component of resistance. Here we use a combination of histological and live-imaging techniques, together with spectroscopy and gene expression analysis to understand the nature of R. solanacearum-induced formation of vascular coatings in resistant tomato. We describe that resistant tomato specifically responds to infection by assembling a vascular structural barrier formed by a ligno-suberin coating and tyramine-derived hydroxycinnamic acid amides. Further, we show that overexpressing genes of the ligno-suberin pathway in a commercial susceptible variety of tomato restricts R. solanacearum movement inside the plant and slows disease progression, enhancing resistance to the pathogen. We propose that the induced barrier in resistant plants does not only restrict the movement of the pathogen, but may also prevent cell wall degradation by the pathogen and confer anti-microbial properties, effectively contributing to resistance.Research is funded by MCIN/AEI/10.13039/501100011033 (NSC, MV), MCIN/AEI/PID2019-110330GB-C21 (MF, OS), MCIN/AEI/PID2020-118968RBI00 (JR), through the ‘Severo Ochoa Programme for Centres of Excellence in R&D’ (SEV-2015-0533, CEX2019-000917 and CEX2019-000902-S funded by MCIN/AEI/ 10.13039/501100011033), and by the Spanish National Research Council (CISC) pie-201620E081 (JR, AG) and the Generalitat de Catalunya (2017SGR765 grant). AK is the recipient of a Netaji Subhas – Indian Council of Agricultural Research International Fellowship. SS acknowledges financial support from DOC-FAM, European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement no. 754397. This work was also supported by the CERCA Program/Generalitat de Catalunya.Peer reviewe

Adipocyte fatty-acid binding protein is overexpressed in cirrhosis and correlates with clinical outcomes

Fatty-acid-binding proteins (FABPs) are small intracellular proteins that coordinate lipid-mediated processes by targeting metabolic and immune response pathways. The aim of the study was to investigate plasma FABPs levels and their relationship with clinical outcomes in cirrhosis. Plasma levels of L-FABP1(liver and kidney), I-FABP2(intestine), and A-FABP4(adipocyte and macrophages) were measured in 274 patients with decompensated cirrhosis. Hepatic gene expression of FABPs was assessed in liver biopsies from patients with decompensated cirrhosis and in liver cell types from mice with cirrhosis. Immunohistochemistry of A-FABP4 in human liver biopsy was also performed. Plasma levels of FABPs were increased in patients with decompensated cirrhosis compared to those of healthy subjects (L-FABP1: 25 (17-39) vs 10 (9-17) ng/mL p = 0.001, I-FABP2: 1.1 (0.5-2.1) vs 0.6 (0.4-1) ng/ mL p = 0.04 and A-FABP4: 37 (20-68) vs 16 (11-33) ng/mL p = 0.002), respectively. Increased A-FABP4 levels were associated with complications of cirrhosis, acute-on-chronic liver failure and poor survival. Hepatic A-FABP4 gene expression was upregulated in decompensated cirrhosis. Macrophages were the main liver cell that over-expressed A-FABP4 in experimental cirrhosis and increased A-FABP4 was found in macrophages of human biopsies by immunohistochemistry. A-FABP4 levels are increased in decompensated cirrhosis and correlate with poor outcomes. Liver macrophages appear to be the main source of A-FABP4 in decompensated cirrhosis