Biology subjects

Fisher, S. R.

Publications and source records attributed to Fisher, S. R..

5 recordsLinked to original sources

bioRxiv2025

Receptor-substrate competition for the TonB homologue FusB suggests a model for ferredoxin import in Pectobacterium spp.

TonB-dependent uptake systems of Gram-negative bacterial pathogens constitute prominent virulence factors, allowing nutrient acquisition, primarily siderophore-bound iron, to cross the highly impermeable outer membrane (OM). Remarkably, the ferredoxin uptake system (Fus) of Pectobacteriaceae, a group of soft rot-inducing plant pathogens, imports an entire folded host protein into the periplasm and extracts its bound iron for growth. The inner membrane protein FusB, a TonB homologue, plays two roles in facilitating ferredoxin import. First, like other TonBs, it remodels the globular plug domain obstructing the lumen of the OM receptor FusA to allow ferredoxin passage. Unusually for a TonB protein, FusB then interacts directly with the FusA-bound ferredoxin substrate to facilitate its transport into the periplasm. In this work, we describe structures of FusB-ferredoxin and homodimeric FusB complexes and determine the key features of the binding interfaces formed by FusB with FusA and ferredoxin. We postulate that under resting conditions FusB exists a homodimer, stabilised by an intermolecular R241-D322 salt bridge. The homodimer dissociates when the "FusB-box" of FusA outcompetes one protomer, and FusA D53 displaces FusB D322. Upon ferredoxin binding, FusB undergoes a structural rearrangement, expanding its {beta}-sheet from three to four strands. In agreement with the proposed sequence of events, ferredoxin binding displaces the receptor (FusA) from FusB with Arg241 forming an intramolecular salt bridge with Asp322 to stabilise the newly formed {beta}-hairpin of FusB. We propose a mechanistic model for ferredoxin import in which FusB Arg241 acts as a molecular switch, and two distinct regions function as interaction hotspots.

biophysics↗

bioRxiv2025

Barley BODYGUARD controls cuticular specialisations regulated by SHINE transcription factors

The outer epidermis of land plants secretes a cuticular layer, a hydrophobic diffusion barrier which minimises water loss into the atmosphere and protects from pests, ultraviolet light and organ fusion. Cuticles typically comprise a polyester cutin matrix embedded and overlaid with cuticular waxes, but their exact chemical make-up, structure and functions can vary widely depending on the tissue and species. Barley shows two such cuticular specialisations: (1) deposition of a thick {beta}-diketone-rich wax bloom on multiple organs at reproductive stage, common in other Poeceae species and linked to yield; and, (2) secretion of a sticky layer on the grain fruit (caryopsis) pericarp cuticle which adheres to inner floral hulls, leading to barleys distinctive covered grain used in animal feed and malting. Two SHINE/WAX-INDUCER transcription factors in barley, HvWIN1 and NUD, promote the wax bloom and hull to caryopsis adhesion, respectively, yet little is understood about other genes involved. Leveraging near-isogenic lines of wax-deficient mutants, we identify the barley BODYGUARD1 (HvBDG1) gene encoding an /{beta}-hydrolase essential for leaf cuticular integrity and wax bloom deposition. Modelling of functional and defective alleles suggests that HvBDG1 N-terminal region control of protein flexibility is important for HvBDG1 function. In addition to their role in controlling barley epicuticular wax deposition, we show that both HvBDG1 and HvWIN1 are essential for strong hull to caryopsis adhesion. Along with NUD, these gene products differentially contribute to ultrastructural changes on the pericarp associated with a cuticular building programme driven by NUD and HvWIN1 regulation of cuticle metabolism and transport and cell wall-related genes, and correlate with shifts in pericarp surface chemistry. We also show that the previously asserted grain-specific role of NUD should be revised, as our findings reveal that it is essential for maintaining leaf cuticle integrity. Our analyses in barley suggest that NUD and HvWIN1 control cuticular specialisations and cuticle integrity in part via promotion of HvBDG1 expression, while HvWIN1 and NUD likely act independently from each other. Lastly, mining tetraploid wheat mutant populations followed by crossing to combine mutated homoeologues demonstrated that BDG1 and WIN1 orthologues also control wax bloom in wheat. Taken together, our work greatly expands the genetic networks and molecular activities important for cuticle development in cereals and the underlying mechanisms for both shared and species-specific cuticular specialisations.

plant biology↗

bioRxiv2025

Transcriptional response to Phytophthora root rot in raspberry identifies RiABP19, a Germin-like protein (GLP) gene with a putative role in resistance

Most phytophthora root rot (PRR) outbreaks in symptomatic commercially cultivated raspberry varieties are associated with the prevalence of Phytophthora rubi. Reduced availability of chemical actives and the persistent presence of Phytophthora oospores in the soil contribute to its devastating impact on raspberry-growing regions. In this study, we examined the variation in root morphology in two contrasting raspberry cultivars, Latham (PRR resistant) and Glen Moy (PRR susceptible). We performed RNA-sequencing on Latham roots challenged with P. rubi, to study the transcriptomic response and uncover mechanisms underpinning resistance. We established a new raspberry reference transcript dataset that allowed quantification of raspberry root gene expression. Transcripts significantly upregulated in Latham challenged with P. rubi, included many with characterised roles in resistance, such as Pathogenesis-related proteins and a Germin-like protein, designated RiABP19. The homologous Glen Moy RiABP19 gene showed no differential transcriptional response to PRR infection, indicating a resistance cultivar-specific induction signature following PRR challenge. Three-dimensional structural modelling predicts that RiABP19 contains conserved active sites implicated in auxin-binding and superoxide dismutase activity and can form a homo-hexamer like true germins. Co-immunoprecipitation assays confirmed that RiABP19 can form both homo- and heterodimers in planta. Virus-induced gene silencing of RiABP19 orthologs of in the model plant Nicotiana benthamiana strongly impacts immune signalling, enhancing Phytophthora infestans colonization and attenuating resistance and cell death triggered by the tomato Cf4/Avr4 interaction. These findings suggest that RiABP19 functions as a positive regulator of immunity and may represent a target for future crop improvement in raspberries.

plant biology↗

bioRxiv2025

The highly conserved aphid effector pair Mp1-Mp58 associates to form an effector complex that targets host trafficking protein VPS52

Pathogen and pest effectors play a crucial role in manipulating plant biological processes, facilitating infection and infestation. While pathogens and pests secrete repertoires of effectors into their host plants, most effector function studies focus on characterising individual proteins. We previously identified a genetically linked and co-regulated gene pair in the aphid Myzus persicae encoding effectors Mp1 and Mp58. Here, we explored the functional link between these effectors. We used ectopic expression assays in Nicotiana benthamiana followed by co-immunoprecipitation assays and confocal microscopy to explore effector-effector and effector-target interactions and their subcellular localisation. We produced recombinant proteins to validate effector interactions and used computational modelling to predict effector complex 3D structures. We revealed that effectors Mp1 and Mp58 interact in planta and in vitro and likely form an oligomeric complex. Both effectors associate with the host target Vacuolar Protein Sorting associated Protein 52 (VPS52) to form an Mp1-Mp58-VPS52 complex which localises at vesicle-like structures. Our findings point to effector complex formation in plant-insect interactions and highlight a further layer of complexity in the molecular dialogue between plants and insects. Our work also shows the importance of considering the context in which effectors may function within a larger effector repertoire.

plant biology↗

bioRxiv2023

Computational prediction of structure, function and interaction of aphid salivary effector proteins

Similar to plant pathogens, phloem-feeding insects such as aphids deliver effector proteins inside their hosts that act to promote host susceptibility and enable feeding and infestation. Despite exciting progress towards identifying and characterizing effector proteins from these insects, their functions remain largely unknown. The recent ground-breaking development in protein structure prediction algorithms combined with the availability of proteomics and transcriptomic datasets for agriculturally important pests, such as the aphid Myzus persicae (green peach aphid), provides new opportunities to explore the structural and functional diversity of effector repertoires. In this study, we sought to gain insight into the the M. persicae effector repertoire by predicting and analysing the structures of a set of 71 effector candidate proteins. We used two protein structure prediction methods, AlphaFold and OmegaFold, which produced mutually consistent results. We observed a wide continuous spectrum of sizes and structures among the effector candidates, from disordered proteins to globular enzymes. We made use of the structural information and state-of-the-art computational methods to predict M. persicae effector protein properties, including function and interaction with host plant proteins. Overall, our investigation provides novel insights into the structure, function, and interaction prediction of aphid effector repertoires and will guide the necessary experimental characterization to address new hypotheses.

biochemistry↗