Analysis of Barley Leaf Epidermis and Extrahaustorial Proteomes During Powdery Mildew Infection Reveals That the PR5 Thaumatin-Like Protein TLP5 Is Required for Susceptibility Towards Blumeria graminis f. sp. hordei

Abstract

Powdery mildews are biotrophic pathogens causing fungal diseases in many economically important crops, including cereals, which are affected by Blumeria graminis. Powdery mildews only invade the epidermal cell layer of leaf tissues, in which they form haustorial structures. Haustoria are at the center of the biotrophic interaction by taking up nutrients from the host and by delivering effectors in the invaded cells to jeopardize plant immunity. Haustoria are composed of a fungal core delimited by a haustorial plasma membrane and cell wall. Surrounding these is the extrahaustorial complex, of which the extrahaustorial membrane is of plant origin. Although haustoria transcriptomes and proteomes have been investigated for Blumeria, the proteomes of barley epidermis upon infection and the barley components of the extrahaustorial complex remains unexplored. When comparing proteomes of infected and non-infected epidermis, several classical pathogenesis-related (PR) proteins were more abundant in infected epidermis. These included peroxidases, chitinases, cysteine-rich venom secreted proteins/PR1 and two thaumatin-like PR5 protein isoforms, of which TLP5 was previously shown to interact with the Blumeria effector BEC1054 (CSEP0064). Against expectations, transient TLP5 gene silencing suggested that TLP5 does not contribute to resistance but modulates susceptibility towards B. graminis. In a second proteomics comparison, haustorial structures were enriched from infected epidermal strips to identify plant proteins closely associated with the extrahaustorial complex. In these haustoria-enriched samples, relative abundances were higher for several V-type ATP synthase/ATPase subunits, suggesting the generation of proton gradients in the extrahaustorial space. Other haustoria-associated proteins included secreted or membrane proteins such as a PIP2 aquaporin, an early nodulin-like protein 9, an aspartate protease and other proteases, a lipase, and a lipid transfer protein, all of which are potential modulators of immunity, or the targets of pathogen effectors. Moreover, the ER BIP-like HSP70, may link ER stress responses and the idea of ER-like properties previously attributed to the extrahaustorial membrane. This initial investigation exploring the barley proteomes of Blumeria-infected tissues and haustoria, associated with a transient gene silencing approach, is invaluable to gain first insight of key players of resistance and susceptibility.

Physiological and Proteomic Changes in the Apoplast Accompany Leaf Senescence in Arabidopsis

“…Leaf senescence-related changes in the secretome, along with variations in the AF volume and pH resemble pathogen attack responses, that in turn involve the induction of many Senescence Associated Genes, SAGs genes (Little et al, 2007;Schippers et al, 2015). PR5 markedly accumulates in the apoplast in response to a variety of biotic and abiotic stresses (Piofczyk et al, 2015;Lambertucci et al, 2019), suggesting a role in a possible universal stress response, or in a crosstalk point where different stress responses converge. It would be plausible that stress-related and senescence programs trigger common mechanisms under conditions requiring recycling.…”

Identification of virulence associated milRNAs and their bidirectional targets in Rhizoctonia solani and maize during infection

“…Notably, GRMZM2G154449 encoding a Thaumatin-like protein (TLP), was predicted to be targeted by Rhi-milR-4104-5p. Previous studies have revealed that plant TLP is classified into the PR protein family 5 (PR5) and exhibits an antifungal property [46]. These results indicated that Rhi-milR-4104-5p may negatively regulate resistance of maize by targeting TLP genes.…”

An interolog-based barley interactome as an integration framework for immune signaling

“…Chitinases, which we also found overexpressed at this timepoint, are secreted by the host cell to degrade fungal walls and elicit PAMP-triggered immunity (PTI) through chitin recognition (Pusztahelyi, 2018). Accumulation of chitinases has also been reported in the extra-haustorial complex during Bgh infection (Lambertucci et al, 2019). Similarly, other protein families that were also reported in the extrahaustorial complex were also found as DM DE at 32 HAI including genes associated with the TCA cycle and pyruvate metabolism, germin-like protein, and cysteine synthase.…”

“…Similarly, other protein families that were also reported in the extrahaustorial complex were also found as DM DE at 32 HAI including genes associated with the TCA cycle and pyruvate metabolism, germin-like protein, and cysteine synthase. Genes involved in pyruvate metabolism are upregulated during haustorial development to contribute to energy production in defense and provide with precursors of other signaling pathways (Qiu et al, 2020;Lambertucci et al, 2019). Remarkably, among this group there is also a malate dehydrogenase (HORVU.MOREX.r2.1HG0067490) that was characterized as target of the Bgh effector BEC1054/CSEP0064 (Pennington et al, 2016).…”

“…This article is a US Government work. It is not subject to copyright under 17 USC The copyright holder for this preprint this version posted November 4, 2021. ; https://doi.org/10.1101/2021.11.02.466982 doi: bioRxiv preprint involved in pyruvate metabolism are upregulated during haustorial development to contribute to energy production in defense and provide with precursors of other signaling pathways (Qiu et al, 2020;Lambertucci et al, 2019). Remarkably, among this group there is also a malate dehydrogenase (HORVU.MOREX.r2.1HG0067490) that was characterized as target of the Bgh effector BEC1054/CSEP0064 (Pennington et al, 2016).…”