Biology subjects

Moffett, P.

Publications and source records attributed to Moffett, P..

11 recordsLinked to original sources

bioRxiv2026

Extracellular water withdrawal drives disease resistance in the phyllosphere

A central question in immunity is how hosts arrest pathogen growth. Diverse plant pathogens create a water-soaked niche in host tissue essential for pathogenesis, yet how water shapes infection outcome is unknown. Using genetics and hyperspectral imaging, we show that extracellular water status is rate-limiting for both compatible and incompatible interactions. We find that the hypersensitive response of effector-triggered immunity (ETI) is, mechanistically, a desiccation event. Water loss imposes osmotic stress that arrests bacterial division while the pathogen remains alive and metabolically active, rather than killing it. Restoring apoplastic water reverses this stasis and licenses growth despite intact immune signaling and cell death. Water status, not immune signaling per se, gates pathogen growth. This reframes ETI as a controlled desiccation mechanism and identifies hydration as a decisive lever on disease outcome.

plant biology↗

bioRxiv2026

Atypical immunity induced by extracellular water in plants

Microbial pathogens require nutrients and water to support their growth and proliferation. Pathogen-mediated resource acquisition is orchestrated by the secretion of virulence factors that have evolved a diversity of forms (from effector proteins to small toxins), but which have converged in function (resource acquisition). A key observable virulence mechanism employed by a large number of microbial pathogens to cause disease is to induce a water-rich niche in the extracellular space (i.e. the apoplast) of their host, known as water-soaked lesions. Given the ubiquity of this virulence mechanism, we asked whether plants respond to pathogen-induced extracellular water to activate immune programs. We find that inducing a water-soaked apoplast induces an atypical transcriptional response, yet results in a functional immune response that restricts pathogen growth as effectively as canonical pattern-triggered immunity. Genetic and functional analyses reveal a role for the immune phytohormone salicylic acid (SA) in mediating apoplastic water-induced immunity (AWII). These results suggest that plants engage in an immune-priming program in response to apoplastic water accumulation, suggesting that the pathogenic niches induced during infection may be perceived as danger signals.

plant biology↗

bioRxiv2026

Botrytis virus X ORF2 suppresses RNA silencing in a trigger-restricted manner and modulates is associated with altered Dicer-like gene expression in Botrytis cinerea

RNA interference (RNAi) is a central antiviral defense mechanism in fungi, yet relatively few mycoviral suppressors of RNA silencing (VSRs) have been functionally characterized, particularly in phytopathogenic hosts. Botrytis virus X (BVX), a positive-sense RNA virus in the family Alphaflexiviridae, infects Botrytis cinerea and encodes five predicted open reading frames (ORFs), most of which have unknown functions. Here, we screened BVX ORFs 2-5 for RNA silencing suppressor activity using complementary GFP-based assays in Nicotiana benthamiana and examined the leading candidate in the fungal host B. cinerea. BVX ORF2 (X2) enhanced GFP transcript and protein accumulation in assays where silencing is triggered by sense RNA but failed to suppress silencing triggered by hairpin-derived siRNAs or miRNA-guided targeting, indicating a trigger-restricted suppressor phenotype. In B. cinerea, transgenic expression of X2 was associated with reduced induction of the RNAi associated genes BcDCL1 and BcDCL2 compared to empty vector controls, with the strongest effect observed on BcDCL1. In a virus-infected fungal background, X2 expression was also associated with increased viral RNA accumulation. Together, these results identify BVX X2 as a BVX-encoded, trigger-restricted suppressor of RNA silencing and link its expression to altered RNAi-related gene induction and increased viral RNA accumulation in B. cinerea.

microbiology↗

bioRxiv2025

Exploring the mycovirome: novel and diverse mycoviruses in Botrytis cinerea

Botrytis cinerea is a necrotrophic fungal pathogen that causes significant economic losses to many crops, including vegetables, fruits, and ornamental plants. The management of B. cinerea is difficult due to a rise in fungicide resistance. Harnessing mycoviruses that cause reduced virulence (hypovirulence) in B. cinerea is a promising alternative. Over 100 mycoviruses have been identified in Botrytis spp. to date, including several hypovirulence-inducing mycoviruses. This research aimed to further explore, for the first time in Canada, the mycovirome of B. cinerea and identify potential hypovirulence-inducing mycoviruses. Isolates of B. cinerea were collected from fruits and vegetables in the province of Quebec. Fitness and pathogenicity criteria, including sclerotia production, colony morphotype, and lesion size were evaluated. A double-stranded RNA (dsRNA) extraction protocol tailored to the detection of mycoviruses was used to sequence dsRNA from 45 isolates with low fitness/pathogenicity, and an in-house bioinformatics workflow was used to profile the mycovirome. Mycoviruses were identified in 44/45 isolates. Most of these had positive single-stranded RNA or dsRNA genomes, and a small number had negative single-stranded RNA, single-stranded DNA, or reverse transcriptase RNA genomes. Following deep analysis of RNA-dependent RNA polymerase and replication initiation proteins, a total of 62 unique contigs were identified belonging to new strains of mycovirus species. Furthermore, four putative novel mycovirus species belonging to Endornaviridae, Botybirnaviridae, Peribunyaviridae, and Bunyavirales taxa were identified. Several mycovirus species positively and/or negatively co-occurred with B. cinerea isolates collected from strawberry or raspberry. This study revealed a high degree of diversity in the mycovirome of B. cinerea. Species accumulation curve analysis indicated that, with the number of isolates characterized, we were unable to capture the full extent of expected diversity. Nevertheless, we identified potential hypovirulence-inducing mycoviruses, including Botrytis cinerea mitovirus 1, Botrytis cinerea hypovirus 1, and Botrytis porri botybirnavirus 1. Some of these novel mycoviruses belonged to taxa known to produce viral particles, which can be an interesting feature for their use as biocontrol agents (BCA). ImportanceThis study provides the first comprehensive profiling of mycoviruses infecting Botrytis cinerea in Canada, a significant step in understanding how these viruses can naturally limit crop disease. Due to growing resistance against conventional fungicides, new biological methods to control B. cinerea are crucial. By profiling mycoviruses in fungal samples collected in Quebec, we identified several novel viruses that appear to reduce the pathogenicity of B. cinerea. These viruses, known as hypovirulence-inducing mycoviruses, could be used to develop biocontrol agents (BCA), offering a more sustainable disease management alternative. Notably, we found virus families with extracellular potential, which may enable easier application as BCAs in agriculture. This research not only broadens the understanding of fungal virology but also holds promise for innovative, eco-friendly approaches to managing Botrytis cinerea in Canada.

genomics↗

bioRxiv2024

Chloroplastic ROS bursts initiate salicylic acid biosynthesis in plant immunity

Chloroplasts are essential centers of signal integration and transduction in plants. They are involved in the biosynthesis of primary and specialized metabolites, including salicylic acid (SA), a key defense phytohormone synthesized via the conserved chorismate biosynthetic pathway. However, the identity of the signal(s) that ultimately triggers SA induction in chloroplasts upon perception of a biotic threat has remained elusive. Here, we provide evidence of a functional link between chloroplast-derived reactive oxygen species (cROS) and SA production. We observe that inhibiting ROS bursts generated from photosystem II during plant immune activation completely abrogates the induction of SA synthesis in response to immunity-inducing signals, without affecting SA-independent immune responses. Indeed, time course analyses show that the induction of SA marker genes parallels that of cROS production during an immune response. Consistent with this, preventing cROS induction is sufficient to nullify the immune protection normally conferred by activating immunity prior to an infection. Analyses of transcriptomes and photosynthetic efficiency show that two conserved effectors from the phytopathogen Pseudomonas syringae, HopM1 and AvrE1, redundantly disrupt photosynthesis and cROS bursts. These effects reduce SA accumulation and are mediated via the impact of HopM1 and AvrE1 in inducting host abscisic acid signaling. Our results suggest that a change in chloroplastic redox homeostasis induced by biotic stressors acts as an initiator of plant immunity through the production of SA, and that this response is targeted by conserved pathogen effector proteins.

plant biology↗

bioRxiv2023

Molecular responses of agroinfiltrated Nicotiana benthamiana leaves expressing suppressor of silencing P19 and influenza virus-like particles

The production of influenza vaccines in plants is achieved through transient Agrobacterium-mediated expression of viral hemagglutinins (HAs). These proteins are produced and matured through the secretory pathway of plant cells, before being trafficked to the plasma membrane where they induce formation of virus-like particles (VLPs). Production of VLPs unavoidably impacts plant cells, as do viral suppressors of RNA silencing (VSRs) that are often co-expressed to increase protein yields. However, little information is available on host molecular responses to these foreign proteins. The present work provides a comprehensive overview of transcriptomic, metabolic, and signaling changes occurring in Nicotiana benthamiana leaf cells transiently expressing the VSR P19, or co-expressing P19 and an influenza HA. Our data identifies generic responses to Agrobacterium-mediated expression of foreign proteins, including shutdown of chloroplast gene expression, activation of oxidative stress responses, and reinforcement of the plant cell wall through lignification. Our results also indicate that P19 expression promotes salicylic acid (SA) signaling, a process apparently antagonized by co-expression of HA. As the latter induces specific signatures, with effects on lipid metabolism, lipid distribution, and oxylipin signaling, dampening of P19 responses suggests crosstalk between SA and oxylipin pathways. Consistent with the upregulation of oxidative stress-related genes and proteins, we finally show that reduction of oxidative stress damage through exogenous application of ascorbic acid improves plant biomass quality during production of VLPs. One-sentence summaryAgrobacterium-mediated expression of influenza virus-like particles induces a unique molecular signature in Nicotiana benthamiana leaf cells.

plant biology↗

bioRxiv2023

From Asymptomatic to Symptomatic: Multiomics profiling of the temporal response of grapevine viral-mixed infection

Mixed viral infections are common in grapevines. However, our understanding of the factors and signaling pathways that influence the expression of viral symptoms in mixed infections is still incomplete. In a previous study, we revealed that the presence of grapevine leafroll-associated virus species in mixed infections was randomly associated with the devel-opment of virus-like symptoms. To understand what drives the timing of these virus-like symptoms in mixed infections, we used dsRNA and total RNA sequencing and metabolomic analysis to profile the viromes, metabolites, and transcripts of grapevine leaves collected at two different times of the year (summer and autumn). We demonstrated that neither viral titre nor virome composition changes were associated with symptom expression in autumn. The total phenolic content and antioxidant capacity increased in most plants except for those with early onset symptoms. According to the results of differential gene expression analysis, cell wall biosynthesis pathways were significantly downregulated in all grapevine plants infected with grapevine leafroll-associated virus 3, grapevine asteroid mosaic-associated virus, and grape-vine Pinot gris virus. In addition, polyketide pathways were significantly upregulated in all cultivars, while flavonoid precursor (e.g. abscisic acid) production was significantly reduced in plants that expressed strong virus-like symptoms. In the Vidal cultivar, an uncharacterized double-stranded RNA-binding protein (DRB) appears to play a critical role in the plants an-tiviral defences, supporting the recent hypothesis that DRBs make an important contribution to dominant antiviral responses in plants. The seasonality of the expression virus-like symptoms appears to be a consequence of the dynamic interactions between antiviral factors and viral counter-defences that occur at different developmental stages of grapevine.

plant biology↗

bioRxiv2023

Nanovirseq: dsRNA sequencing for plant virus and viroid detection by Nanopore sequencing

Worldwide, there is a need for certified clean plant materials to limit viral diseases spread. In order to design a robust and proactive viral-like disease certification, diagnostics, and management program, it is essential to have a fast, inexpensive, and user-friendly tool. The purpose of this study was to determine whether dsRNA-based nanopore sequencing can be a reliable method for the detection of viruses and viroids in grapevines or not. Compared to direct RNA sequencing from rRNA-depleted total RNA (rdTotalRNA), direct-cDNA sequencing from dsRNA (dsRNAcD) yielded more viral reads and detected all grapevine viruses and viroids detected using Illumina MiSeq sequencing (dsRNA-MiSeq). With dsRNAcD sequencing it was possible to detect low abundance viruses (e.g., Grapevine red globe virus) where rdTotalRNA sequencing failed to detect them. Indeed, even after removing rRNA, rdTotalRNA sequencing yielded low viral read numbers. rdTotalRNA sequencing was not sensitive enough to detect all the viruses detected by dsRNA-MiSeq. In addition, there was a false positive identification of a viroid in the rdTotalRNA sequencing that was due to misannotation of a host-driven read. For quick and accurate reads classification, two different taxonomical classification workflows based on protein and nucleotide homology were evaluated in this study, namely DIAMOND&MEGAND (DIA&MEG) and Centrifuge&Recentrifuge (Cent&Rec), respectively. Virome profiles from both workflows were similar except for grapevine endophyte endornavirus (GEEV), which was only detected using DIA&MEG. However, because DIA&MEGs classification is based on protein homology, it cannot detect viroid infection despite giving more robust results. Even though Cent&Recs virus and viroid detection workflow was faster (30 minutes) than DIA&MEGs (two hours), it could not provide the details and information DIA&MEG was able to provide. As demonstrated in our study, nanopore dsRNAcD sequencing and the proposed data analysis workflows are suitable and reliable for viruses and viroids detection, especially in grapevine where viral mixed infection is common.

pathology↗

bioRxiv2022

Light prevents pathogen-induced aqueous microenvironments via potentiation of salicylic acid signaling

Upon establishment of an infection, many plant pathogens induce an aqueous microenvironment in the extracellular space of their host, resulting in water-soaked lesions. In the case of Pseudomonas syringae (Pst), this is accomplished through the activity of water-soaking effectors that stimulate abscisic acid (ABA) production and signaling, which results in stomatal closure. This reduces transpiration and induces a microenvironment favorable for bacterial growth. Stomata are also highly sensitive to environmental conditions, including light and circadian rhythm. Here, we show that a period of darkness is required for water-soaking, and that a constant light regime abrogates the water-soaking activity of Pst effectors. Additionally, we show that constant light induces resistance against Pst and that this effect requires salicylic acid (SA). An increase in SA production upon infection under constant light did not affect effector-induced ABA signaling, but rather abrogated ABAs ability to induce stomatal closure. Indeed, under normal diurnal light regimes, application of a SA analog is sufficient to prevent the ability of the pathogen to induce stomatal closure and a water-rich niche in the apoplast. Our results provide a novel approach to interfering with a common virulence strategy, as well as providing a physiological mechanism by which SA functions in defense against certain pathogens.

plant biology↗

bioRxiv2019

Natural variation in the Arabidopsis AGO2 gene is associated with susceptibility to potato virus X

RNA silencing functions as an anti-viral defence in plants through the action of DICER-like (DCL) and ARGONAUTE (AGO) proteins. However, there are few known examples of functional variation in RNA silencing components. The AGO2 protein is important for antiviral defense against multiple viruses and has been shown to be a major limiting factor to infection by potato virus X (PVX) of Arabidopsis thaliana but not Nicotiana benthamiana. We show that the AGO2 proteins from these two plants have differential activity against PVX, suggesting that variation in AGO2 is important in plant-virus interactions. Consistent with this, we find that the Arabidopsis thaliana AGO2 gene shows a high incidence of polymorphisms between accessions, with evidence of selective pressure. AGO2 protein variants can be assigned to two groups, in near equal frequency, based on an amino acid change and small deletions in the protein N-terminus. Inoculation of a large number of Arabidopsis accessions shows strong correlation between these alleles and resistance or susceptibility to PVX. These observations were validated using genetic and transgenic complementation analysis, which showed that one type of AGO2 variant is specifically affected in its antiviral activity, without interfering with other AGO2-associated functions such as anti-bacterial resistance or DNA methylation. Our results demonstrate a novel type of genetically-encoded virus resistance and suggest that plant-virus interactions have influenced natural variation in RNA silencing components.

plant biology↗

bioRxiv2019

ARGONAUTE5 Mediates Fine-Tuning of Vegetative-to-Reproductive Phase Transition Through Its Interaction with miR156 in Arabidopsis

Vegetative-to-reproductive phase change is a finely tuned process in plants, largely controlled by the age-regulated microRNA156 (miR156), which functions by suppressing the transcripts of SQUAMOSA-PROMOTER BINDING LIKE (SPL) transcription factors. ARGONAUTE proteins (AGO) are essential effectors of miRNA-mediated gene regulation. However, which AGO(s) mediate(s) the control of flowering time remains unclear. Here, we demonstrate a role for AGO5 in vegetative-to-reproductive phase transition through the modulation of SPL transcription factors. We show that AGO5 interacts physically and functionally with miR156 and that ago5 mutants present an early flowering phenotype in Arabidopsis. Furthermore, in ago5 mutants, the repression of flowering caused by miR156 overexpression is largely reversed, whereas leaf morphology remains unaffected. Our results thus indicate a specific role for AGO5 in mediating miR156 activity in meristematic, but not vegetative, tissue. As such, our data suggest a spatiotemporal regulation of the miR156 aging pathway, mediated through different AGO proteins in different tissues.

plant biology↗