Biology subjects

Ryder, L. S.

Publications and source records attributed to Ryder, L. S..

8 recordsLinked to original sources

bioRxiv2026

A Pmk1-Regulated Mst12-Bip1 Network Coordinates Appressorium Function, Effector Deployment, and Invasive Growth by Magnaporthe oryzae

To cause rice blast disease, the filamentous fungus Magnaporthe oryzae develops a specialised infection cell called an appressorium, which generates enormous turgor to breach the rice leaf cuticle. Although key regulators of appressorium development have been identified, it is not known how they drive the extensive transcriptional reprogramming required for infection-related morphogenesis. Here, we show that the Pmk1 MAP kinase orchestrates plant infection by regulating a transcriptional network controlled by the Mst12 and Bip1 transcription factors. Bip1 is regulated both by Pmk1-dependent phosphorylation and at the transcriptional level, while Mst12 binds to a cis-acting element upstream of BIP1 essential for pathogenesis. Bip1 and Mst12 are both necessary for regulating a set of Pmk1-dependent appressorium-specific genes, including transcriptional regulators, cell wall-degrading enzymes, and effector proteins. In addition, Mst12 specifically regulates functions required for appressorium-mediated penetration, while Bip1 controls a distinct sub-set of effector genes deployed during invasion of plant tissue. When considered together, these findings define a Pmk1-dependent transcriptional network required for plant infection by the blast fungus.

molecular biology↗

bioRxiv2026

Computational design of de novo integrated domains enables rational control of pathogen effector recognition in plant NLR immune receptors.

The rapid evolution of plant pathogens poses a persistent threat to global agricultural sustainability, often outpacing the discovery and deployment of natural disease resistance genes. While bioengineering of plant intracellular immune receptors (NLRs) offers a potential solution, developing bespoke immune recognition remains constrained by the laborious characterisation of natural receptors and plant-pathogen interactions. Here, we describe a programmable framework that leverages generative AI protein design tools, RFdiffusion and ProteinMPNN, to design de novo integrated domains (IDs) against diverse pathogen effectors. By integrating these bespoke binders into the modular rice blast Pik-1/Pik-2 NLR receptor chassis, we successfully engineer recognition of a non-cognate virulence factor (effector) from the Panama disease pathogen, Fusarium oxysporum f. sp. cubense Tropical Race 4. Functional assays in Nicotiana benthamiana demonstrate that these de novo domains facilitate specific effector perception and initiate immune signalling, while structural and biophysical analyses confirm that de novo integrated domains maintain high structural fidelity to the initial designs and associate with their targets via the predicted interaction interfaces. Additionally, our findings provide orthogonal evidence for the role of integrated domains in regulation of NLR signalling, demonstrating integration of de novo IDs can either trigger autoactivity or, in some cases, lead to effector-mediated repression of cell death. By decoupling immune perception from natural evolutionary history through deploying AI-designed sensory domains, this work establishes a design-lead framework for generation of programmable plant immune receptors, providing a new avenue for bioengineering crops against emerging pathogens.

plant biology↗

bioRxiv2025

Synchronous spatio-temporal control of autophagy and organelle trafficking is necessary for appressorium-mediated plant infection by Magnaporthe oryzae

The blast fungus Magnaporthe oryzae infects plants using a specialised infection structure called an appressorium that generates physical force to break the rice leaf cuticle. Appressorium development follows a cell cycle-controlled morphogenetic program, requiring autophagy-associated cell death of the fungal spore from which the infection cell develops. How proliferative growth of the fungus is regulated at the same time as programmed cell death, however, is unknown. In this study, we provide evidence that each cell of the conidium undergoes a separate developmental program, which is necessary for plant infection. Using quantitative live-cell imaging, we monitored trafficking of ten organelle types during appressorium morphogenesis in a wild-type M. oryzae strain and isogenic {Delta}atg8 autophagic mutant. High-resolution microscopy using a photoactivatable green fluorescent protein revealed that organelle trafficking occurs from a single conidium cell into the appressorium, while the remaining two cells undergo autophagy. Organelle inheritance operates independently of cell cycle checkpoints but is always associated with spore germination. We furthermore defined the temporal sequence of organelle movement and de novo organelle biogenesis in the incipient appressorium using photoconvertible fluorescent localisation microscopy. Taken together, our study reveals how synchronous spatiotemporal control of autophagy and organelle trafficking is necessary for rice blast infection.

molecular biology↗

bioRxiv2023

The phosphorylation landscape of infection-related development by the rice blast fungus

Many of the worlds most devastating crop diseases are caused by fungal pathogens which elaborate specialized infection structures to invade plant tissue. Here we present a quantitative mass spectrometry-based phosphoproteomic analysis of infection-related development by the rice blast fungus Magnaporthe oryzae, which threatens global food security. We mapped 8,005 phosphosites on 2,062 fungal proteins, revealing major re-wiring of phosphorylation-based signaling cascades during fungal infection. Comparing phosphosite conservation across 41 fungal species reveals phosphorylation signatures specifically associated with biotrophic and hemibiotrophic fungal infection. We then used parallel reaction monitoring to identify phosphoproteins directly regulated by the Pmk1 MAP kinase that controls plant infection by M. oryzae. We define 33 substrates of Pmk1 and show that Pmk1-dependent phosphorylation of a newly identified regulator, Vts1, is required for rice blast disease. Defining the phosphorylation landscape of infection therefore identifies potential therapeutic interventions for control of plant diseases.

plant biology↗

bioRxiv2022

Rgs1 is a regulator of effector gene expression during plant infection by the rice blast fungus Magnaporthe oryzae

To cause rice blast disease the filamentous fungus Magnaporthe oryzae secretes a battery of effector proteins into host plant tissue to facilitate infection. Effector-encoding genes are expressed only during plant infection and show very low expression during other developmental stages. How effector gene expression is regulated in such a precise manner during invasive growth by M. oryzae is not known. Here, we report a forward-genetic screen to identify regulators of effector gene expression, based on selection of mutants that show constitutive effector gene expression. Using this simple screen, we identify Rgs1, a regulator of G-protein signalling (RGS) protein that is necessary for appressorium development, as a novel transcriptional regulator of effector gene expression, which acts prior to plant infection. We show that an N-terminal domain of Rgs1, possessing transactivation activity, is required for effector gene regulation and acts in an RGS-independent manner. Rgs1 controls expression of at least 60 temporally co-regulated effector genes, preventing their transcription during the pre-penetration stage of development prior to plant infection. A regulator of appressorium morphogenesis is therefore also required for orchestration of pathogen gene expression required for invasive growth by M. oryzae during plant infection.

molecular biology↗

bioRxiv2022

Direct measurement of appressorium turgor using a molecular mechanosensor in the rice blast fungus Magnaporthe oryzae

Many plant pathogenic fungi forcibly enter their hosts to cause disease. The rice blast fungus Magnaporthe oryzae, for example, infects plants using a specialised infection cell called an appressorium, which generates enormous turgor to drive a rigid penetration peg through the rice leaf cuticle. While these vast internal pressures are a critical weapon in fungal host penetration, they have remained very challenging to probe directly during host invasion, leaving our understanding of these extreme cellular mechanics incomplete. Here, we combine Fluorescence Lifetime Imaging (FLIM) with a membrane-targeting molecular mechanoprobe to quantify changes in membrane tension as a direct proxy for appressorial turgor in M. oryzae. We report that mature melanin-pigmented M. oryzae appressoria display a heterogeneous low fluorescence lifetime and high membrane tension, consistent with enormous turgor. These extreme pressures lead to large-scale spatial heterogeneities in membrane mechanics, much greater than observed in any other cell type previously, highlighting the extreme mechanics of turgor-driven appressorium-mediated plant infection. By contrast, appressoria of non-pathogenic melanin-deficient mutants, alb1 and buf1, or immature non-melanised appressoria, exhibit high fluorescence lifetime, consistent with low membrane tension and turgor, that remain spatially homogeneous. To evaluate the method, we investigated turgor dynamics in a range of mutants impaired in appressorium function. We show that the turgor sensor kinase mutant{Delta} sln1, recently proposed to generate excess appressorium turgor, displayed a significantly higher membrane tension compared to an isogenic wild type M. oryzae strain. This non-invasive, live cell imaging technique allows direct quantification and visualization of the enormous turgor pressures deployed during pathogen infection.

cell biology↗

bioRxiv2018

Magnaporthe oryzae SMO1 encodes a Ras GTPase-activating protein required for spore morphology, appressorium function and rice blast disease

The pathogenic life cycle of the rice blast fungus Magnaporthe oryzae involves a series of morphogenetic changes, essential for its ability to cause disease. The smo mutation was identified more than twenty-five years ago and affects the shape and development of diverse cell types in M. oryzae, including conidia, appressoria and asci. All attempts to clone the SMO1 gene by map-based cloning and/or complementation, have failed over many years. Here, we report the identification of SMO1 by a combination of bulk segregant analysis and comparative genome analysis. SMO1 encodes a GTPase-activating protein (GAP), which regulates Ras signalling during infection-related development. Targeted deletion of SMO1 results in abnormal, non-adherent conidia, impaired in their production of spore tip mucilage. Smo1 mutants also develop smaller appressoria, with a severely reduced capacity to infect rice plants. SMO1 is necessary for organisation of microtubules and for septin-dependent remodelling of the F-actin cytoskeleton at the appressorium pore. Smo1 physically interacts with components of the Ras2 signaling complex, and a range of other signalling and cytoskeletal components, including the four core septins. SMO1 is therefore necessary for regulation of RAS activation required for conidial morphogenesis and septin-mediated plant infection.

microbiology↗

bioRxiv2018

The MoT3 assay does not distinguish between Magnaporthe oryzae wheat and rice blast isolates from Bangladesh

The blast fungus Magnaporthe oryzae is comprised of lineages that exhibit varying degrees of specificity on about 50 grass hosts, including rice, wheat and barley. Reliable diagnostic tools are essential given that the pathogen has a propensity to jump to new hosts and spread to new geographic regions. Of particular concern is wheat blast, which has suddenly appeared in Bangladesh in 2016 before spreading to neighboring India. In these Asian countries, wheat blast strains are now co-occurring with the destructive rice blast pathogen raising the possibility of genetic exchange between these destructive pathogens. We assessed the recently described MoT3 diagnostic assay and found that it did not distinguish between wheat and rice blast isolates from Bangladesh. The assay is based on primers matching the WB12 sequence corresponding to a fragment of the M. oryzae MGG_02337 gene annotated as a short chain dehydrogenase. These primers could not reliably distinguish between wheat and rice blast isolates from Bangladesh based on DNA amplification experiments performed in separate laboratories in Bangladesh and in the UK. In addition, comparative genomics of the WB12 sequence revealed a complex underlying genetic structure with related sequences across M. oryzae strains and in both rice and wheat blast isolates. We, therefore, caution against the indiscriminate use of this assay to identify wheat blast.

plant biology↗