Biology subjects

Thieron, H.

Publications and source records attributed to Thieron, H..

4 recordsLinked to original sources

bioRxiv2024

Broad-scale phenotyping in Arabidopsis reveals varied involvement of RNA interference across diverse plant-microbe interactions

RNA interference (RNAi) is a crucial mechanism that can contribute to immunity against infectious microbes through the action of DICER-LIKE (DCL) and ARGONAUTE (AGO) proteins. In the case of the fungal pathogen Botrytis cinerea and the oomycete Hyaloperonospora arabidopsidis, plant DCL and AGO proteins have proven roles as negative regulators of immunity, suggesting functional specialization of these proteins. To address this aspect in a broader taxonomic context, we characterized the colonization pattern of an informative set of DCL and AGO loss-of-function mutants in Arabidopsis thaliana upon infection with a panel of pathogenic microbes with different lifestyles, and a fungal mutualist. Our results revealed that AGO1 and AGO4 function as positive regulators of immunity to a bacterial and a fungal pathogen, respectively. Additionally, AGO2 and AGO10 positively modulated the colonization by a fungal mutualist. Therefore, analysing the role of RNAi across a broader range of plant-microbe interactions has identified previously unknown functions for AGO proteins. For some pathogen interactions, however, all tested mutants exhibited wild type-like infection phenotypes, suggesting that the roles of AGO and DCL proteins in these interactions may be more complex to elucidate.

plant biology↗

bioRxiv2024

Barley powdery mildew invasion coincides with the dynamic accumulation of leaf apoplastic extracellular vesicles that are associated with host stress response proteins

The mutual exchange of extracellular vesicles across kingdom borders is a feature of many plant-microbe interactions. The occurrence and cargos of extracellular vesicles has been studied in several instances, but their dynamics in the course of infection have remained elusive. Here we used two different procedures, differential high-speed centrifugation and polymer-based enrichment, to collect extracellular vesicles from the apoplastic wash fluid of barley (Hordeum vulgare) leaves challenged by its fungal powdery mildew pathogen, Blumeria hordei. Both methods yielded extracellular vesicles of similar quality and morphological characteristics, though the polymer approach was associated with higher reproducibility. We noted that extracellular vesicles derived from the apoplastic wash fluid constitute polydisperse populations that are selectively responsive to leaf infection by B. hordei. Extracellular vesicles of [~]100 nm - 300 nm diameter became progressively more abundant, in particular from 72 hours post inoculation onwards, resulting in a major peak late during fungal infection. Vesicles of [~]300 nm - 500 nm showed similar accumulation dynamics but reached much lower levels, suggesting they might constitute a separate population. Proteome analysis uncovered an enrichment of biotic stress response proteins associated with the extracellular vesicles. The barley t-SNARE protein Ror2, the ortholog of the PEN1 marker protein of extracellular vesicles in Arabidopsis thaliana, accumulates in extracellular vesicles during powdery mildew infection, hence also qualifying as a potential marker protein. Our study serves as a starting point for investigating the role of extracellular vesicles at different stages of plant-microbe interactions.

plant biology↗

bioRxiv2022

Site-specific analysis reveals candidate cross-kingdom small RNAs, tRNA and rRNA fragments, and signs of fungal RNA phasing in the barley-powdery mildew interaction

The establishment of host-microbe interactions requires molecular communication between both partners, which involves the mutual transfer of noncoding small RNAs. Previous evidence suggests that this is also true for the barley powdery mildew disease, which is caused by the fungal pathogen Blumeria hordei. However, previous studies lacked spatial resolution regarding the accumulation of small RNAs upon host infection by B. hordei. Here, we analysed site-specific small RNA repertoires in the context of the barley-B. hordei interaction. To this end, we dissected infected leaves into separate fractions representing different sites that are key to the pathogenic process: epiphytic fungal mycelium, infected plant epidermis, isolated haustoria, a vesicle-enriched fraction from infected epidermis, and extracellular vesicles. Unexpectedly, we discovered enrichment of specific 31- to 33-base long 5-terminal fragments of barley 5.8S ribosomal RNA (rRNA) in extracellular vesicles and infected epidermis, as well as particular B. hordei tRNA fragments in haustoria. We describe canonical small RNAs from both the plant host and the fungal pathogen that may confer cross-kingdom RNA interference activity. Interestingly, we found first evidence of phased small RNAs (phasiRNAs) in B. hordei, a feature usually attributed to plants, which may be associated with the post-transcriptional control of fungal coding genes, pseudogenes, and transposable elements. Our data suggests a key and possibly site-specific role for cross-kingdom RNA interference and noncoding RNA fragments in the host-pathogen communication between B. hordei and its host barley.

plant biology↗

bioRxiv2018

A fungal ribonuclease-like effector protein inhibits plant host ribosomal RNA degradation

The biotrophic fungal pathogen Blumeria graminis causes the powdery mildew disease of cereals and grasses. Proteins with a predicted ribonuclease (RNase)-like fold (termed RALPHs) comprise the largest set of secreted effector candidates within the B. graminis f. sp. hordei genome. Their exceptional abundance suggests they play crucial functions during pathogenesis. We show that transgenic expression of RALPH CSEP0064/BEC1054 increases susceptibility to infection in monocotyledenous and dicotyledonous plants. CSEP0064/BEC1054 interacts in planta with five host proteins: two translation elongation factors (eEF1 and eEF1{gamma}), two pathogenesis-related proteins (PR5 and PR10) and a glutathione-S-transferase. We present the first crystal structure of a RALPH, CSEP0064/BEC1054, demonstrating it has an RNase-like fold. The protein interacts with total RNA and weakly with DNA. Methyl jasmonate levels modulate susceptibility to aniline-induced host RNA fragmentation. In planta expression of CSEP0064/BEC1054 reduces the formation of this RNA fragment. We propose that CSEP0064/BEC1054 is a pseudoenzyme that binds to host ribosomes, thereby inhibiting the action of plant ribosome-inactivating proteins that would otherwise lead to host cell death, an unviable interaction and demise of the fungus.

plant biology↗