Functional modulation of an aquaporin to intensify photosynthesis and abrogate bacterial virulence in rice.

Plant aquaporins are a recently noted biological resource with a great potential to improve crop growth and defense traits. Here, we report the functional modulation of the rice (Oryza sativa) aquaporin OsPIP1;3 to enhance rice photosynthesis and grain production and to control bacterial blight and leaf streak, the most devastating worldwide bacterial diseases in the crop. We characterize OsPIP1;3 as a physiologically relevant CO2 -transporting facilitator, which supports 30% of rice photosynthesis on average. This role is nullified by interaction of OsPIP1;3 with the bacterial protein Hpa1, an essential component of the Type III translocon that supports translocation of the bacterial Type III effectors PthXo1 and TALi into rice cells to induce leaf blight and streak, respectively. Hpa1 binding shifts OsPIP1;3 from CO2 transport to effector translocation, aggravates bacterial virulence, and blocks rice photosynthesis. On the contrary, the external application of isolated Hpa1 to rice plants effectively prevents OsPIP1;3 from interaction with Hpa1 secreted by the bacteria that are infecting the plants. Blockage of the OsPIP1;3-Hpa1 interaction reverts OsPIP1;3 from effector translocation to CO2 transport, abrogates bacterial virulence, and meanwhile induces defense responses in rice. These beneficial effects can combine to enhance photosynthesis by 29-30%, reduce bacterial disease by 58-75%, and increase grain yield by 11-34% in different rice varieties investigated in small-scale field trials conducted during the past years. Our results suggest that crop productivity and immunity can be coordinated by modulating the physiological and pathological functions of a single aquaporin to break the growth-defense tradeoff barrier.

Fusarium graminearum GGA protein is critical for fungal development, virulence and ascospore discharge through its involvement in vesicular trafficking.

Vesicular trafficking is a conserved material transport process in eukaryotic cells. The GGA family proteins are clathrin adaptors that are involved in eukaryotic vesicle transport, but their functions in phytopathogenic filamentous fungi remain unexplored. Here, we examined the only GGA family protein in Fusarium graminearum, FgGga1, which localizes to both the late Golgi and endosomes. In the absence of FgGga1, the fungal mutant exhibited defects in vegetative growth, DON biosynthesis, ascospore discharge and virulence. Fluorescence microscopy analysis revealed that FgGga1 is associated with trans-Golgi network (TGN)-to-plasma membrane, endosome-to-TGN and endosome-to-vacuole transport. Mutational analysis on the five domains of FgGga1 showed that the VHS domain was required for endosome-to-TGN transport while the GAT167-248 and the hinge domains were required for both endosome-to-TGN and endosome-to-vacuole transport. Importantly, the deletion of the FgGga1 domains that are required in vesicular trafficking also inhibited vegetative growth and virulence of F. graminearum. In addition, FgGga1 interacted with the ascospore discharge regulator Ca2+ ATPase FgNeo1, whose transport to the vacuole is dependent on FgGga1-mediated endosome-to-vacuole transport. Our results suggest that FgGga1 is required for fungal development and virulence via FgGga1-mediated vesicular trafficking, and FgGga1-mediated endosome-to-vacuole transport facilitates ascospore discharge in F. graminearum.

Aphid salivary protein Mp1 facilitates infestation by binding phloem protein 2-A1 in Arabidopsis.

We have previously demonstrated that Arabidopsis (Arabidopsis thaliana) phloem protein PP2-A1 is an integral component of resistance to the green peach aphid (Myzus persicae). Here, we report that M. persicae overcomes the resistance of PP2-A1 by using the salivary protein Mp1 as an energetic effector and an interactor of AtPP2-A1. Using the RNA interference technique, we demonstrated that Mp1 plays an essential role in the phloem-feeding activity of M. persicae. When the Mp1 gene was silenced, aphids incurred serious impairments not only in phloem-feeding activity, but also in survival and fertility. In essence, phloem-feeding activity was attributed to the molecular interaction between Mp1 and AtPP2-A1. The Mp1 and AtPP2-A1 interactions were localized to plant cell membranes by co-immunoprecipitation and bimolecular fluorescence complementation experiments. Furthermore, the interaction was found to be required for aphid feeding on Arabidopsis phloem. Overall, our results suggest that Mp1 is an important effector of M. persicae and interacts with AtPP2-A1 to facilitate infestation in the plant tissue by this insect.

FgPsd2, a phosphatidylserine decarboxylase of Fusarium graminearum, regulates development and virulence.

Phosphatidylserine decarboxylases (Psds) are enzymes regulating phosphatidylethanolamine biosynthesis in prokaryotes and eukaryotes, and have the central role in lipid metabolism. To date, the functions of Psds in plant pathogenic fungi are not fully understood. In this study, we have characterized two yeast Psd orthologues: FgPsd1 and FgPsd2, in Fusarium graminearum. Our results indicate that FgPsd1 and FgPsd2 are localized in mitochondria and Golgi, respectively. In addition, we have determined that FgPsd1 is a lethal gene and deletion of FgPsd2 resulted in a significant reduction of mycelial growth and conidiation. Futhermore, the FgPsd2 deletion mutant (ΔFgPsd2) is defective in ascospore production and virulence in wheat. Our study has also found that the ΔFgPsd2 mutant is more sensitive to osmotic and oxygen stresses. Moreover, deletion of FgPsd2 reduced the formation of lipid droplets and aggravated autophagy in F. graminearum. In summary, our findings indicate that FgPsd2 is important for mycelial growth, sexual and asexual reproduction, virulence, lipid droplet formation and autophagy in F. graminearum.

The Golgin Protein RUD3 Regulates Fusarium graminearum Growth and Virulence.

Golgins are coiled-coil proteins that play prominent roles in maintaining the structure and function of the Golgi complex. However, the role of golgin proteins in phytopathogenic fungi remains poorly understood. In this study, we functionally characterized the Fusarium graminearum golgin protein RUD3, a homolog of ScRUD3/GMAP-210 in Saccharomyces cerevisiae and mammalian cells. Cellular localization observation revealed that RUD3 is located in the cis-Golgi. Deletion of RUD3 caused defects in vegetative growth, ascospore discharge, deoxynivalenol (DON) production, and virulence. Moreover, the Δrud3 mutant showed reduced expression of tri genes and impairment of the formation of toxisomes, both of which play essential roles in DON biosynthesis. We further used green fluorescent protein (GFP)-tagged SNARE protein SEC22 (SEC22-GFP) as a tool to study the transport between the endoplasmic reticulum (ER) and Golgi and observed that SEC22-GFP was retained in the cis-Golgi in the Δrud3 mutant. RUD3 contains the coiled coil (CC), GRAB-associated 2 (GA2), GRIP-related Arf binding (GRAB), and GRAB-associated 1 (GA1) domains, which except for GA1, are indispensable for normal localization and function of RUD3, whereas only CC is essential for normal RUD3-RUD3 interaction. Together, these results demonstrate how the golgin protein RUD3 mediates retrograde trafficking in the ER-to-Golgi pathway and is necessary for growth, ascospore discharge, DON biosynthesis, and pathogenicity in F. graminearumIMPORTANCEFusarium head blight (FHB) caused by the fungal pathogen Fusarium graminearum is an economically important disease of wheat and other small grain cereal crops worldwide, and limited effective control strategies are available. A better understanding of the regulation mechanisms of F. graminearum development, deoxynivalenol (DON) biosynthesis, and pathogenicity is therefore important for the development of effective control management of this disease. Golgins are attached via their extreme carboxy terminus to the Golgi membrane and are involved in vesicle trafficking and organelle maintenance in eukaryotic cells. In this study, we systematically characterized a highly conserved Golgin protein, RUD3, and found that it is required for vegetative growth, ascospore discharge, DON production, and pathogenicity in F. graminearum Our findings provide a comprehensive characterization of the golgin family protein RUD3 in plant-pathogenic fungus, which could help to identify a new potential target for effective control of this devastating disease.

The Aquaporin TaPIP2;10 Confers Resistance to Two Fungal Diseases in Wheat.

Plants employ aquaporins (AQPs) of the plasma membrane intrinsic protein (PIP) family to import environmental substrates, thereby affecting various processes, such as the cellular responses regulated by the signaling molecule hydrogen peroxide (H2O2). Common wheat (Triticum aestivum) contains 24 candidate members of the PIP family, designated as TaPIP1;1 to TaPIP1;12 and TaPIP2;1 to TaPIP2;12. None of these TaPIP candidates have been characterized for substrate selectivity or defense responses in their source plant. Here, we report that T. aestivum AQP TaPIP2;10 facilitates the cellular uptake of H2O2 to confer resistance against powdery mildew and Fusarium head blight, two devastating fungal diseases in wheat throughout the world. In wheat, the apoplastic H2O2 signal is induced by fungal attack, while TaPIP2;10 is stimulated to translocate this H2O2 into the cytoplasm, where it activates defense responses to restrict further attack. TaPIP2;10-mediated transport of H2O2 is essential for pathogen-associated molecular pattern-triggered plant immunity (PTI). Typical PTI responses are induced by the fungal infection and intensified by overexpression of the TaPIP2;10 gene. TaPIP2;10 overexpression causes a 70% enhancement in wheat resistance to powdery mildew and an 86% enhancement in resistance to Fusarium head blight. By reducing the disease severities, TaPIP2;10 overexpression brings about >37% increase in wheat grain yield. These results verify the feasibility of using an immunity-relevant AQP to concomitantly improve crop productivity and immunity.

Mitochondrial FgEch1 is responsible for conidiation and full virulence in Fusarium graminearum.

Enoyl-CoA hydratase (Ech) is an important and well-recognized enzyme that functions in the degradation of fatty acids by ß-oxidation. However, its functions in plant pathogenic fungi are not well known. We characterized an Ech1 orthologue, FgEch1, in Fusarium graminearum. The FgEch1 deletion mutant was defective in the utilization of short-chain fatty acids and conidiation, but not in hyphal growth on glucose-rich media or in perithecium formation. The FgEch1 deletion mutant showed reduced deoxynivalenol (DON) production and virulence in plants. Deletion of FgEch1 also led to increased production of lipid droplets and autophagy. FgEch1, which was localized in the mitochondrion, required the MTS domain for mitochondrial localization and function in F. graminearum. Taken together, these data indicate that mitochondrial FgEch1 is important for conidiation, DON production, and plant infection.

The type II phosphoinositide 4-kinase FgLsb6 is important for the development and virulence of Fusarium graminearum.

Fusarium graminearum is the main pathogenic fungus causing Fusarium head blight (FHB), which is a wheat disease with a worldwide prevalence. In eukaryotes, phosphatidylinositol 4-phosphate (PI4P), which participates in many physiological processes, is located primarily in different organelles, including the trans-Golgi network (TGN), plasma membrane and endosomes. Type II phosphatidylinositol 4-kinases (PI4Ks) are involved in regulating the production of PI4P in yeast, plants and mammalian cells. However, the role of these proteins in phytopathogenic fungi is not well understood. In this study, we characterized the type II PI4K protein FgLsb6 in F. graminearum, a homolog of Lsb6 in Saccharomyces cerevisiae. Unlike Lsb6, FgLsb6 localizes to the vacuoles and endosomes. The ΔFglsb6 mutant displayed defects in vegetative growth, deoxynivalenol (DON) production and pathogenicity. Furthermore, the ΔFglsb6 deletion mutant also exhibited increased resistance to osmotic, oxidative and cell wall stresses. Further analyses of the ΔFglsb6 mutant showed that it was defective in the generation of PI4P on endosomes and endocytosis. Collectively, our data suggest that the decreased vegetative growth and pathogenicity of ΔFglsb6 was due to the conservative roles of FgLsb6 in the generation of PI4P on endosomes and endocytosis.

Endocytic FgEde1 regulates virulence and autophagy in Fusarium graminearum.

Endocytosis plays critical roles in cellular processes, including nutrient uptake and signal transduction. Ede1 is an endocytic scaffolding protein that contributes to endocytic site initiation and maturation in yeast. However, the functions of Ede1 in phytopathogenic fungi are not known. Here, we identified functions of FgEde1 (FGSG_05182) in Fusarium graminearum. Deletion of FgEde1 resulted in defects in hyphal growth, conidiation and ascospore development. The FgEde1 deletion mutant showed reduced deoxynivalenol (DON) production and virulence in wheat. Furthermore, the FgEde1 deletion mutant also exhibited increased resistance to osmotic and oxidative stress as well as cell-wall perturbing agents. Importantly, deletion of FgEde1 increased the severity of autophagy in hyphae. Taken together, these results reveal that FgEde1 is involved in hyphal growth, asexual and sexual reproduction, virulence, stress responses, and autophagy in F. graminearum.

The Dynamin-Like GTPase FgSey1 Plays a Critical Role in Fungal Development and Virulence in Fusarium graminearum.

Fusarium graminearum, the main pathogenic fungus causing Fusarium head blight (FHB), produces deoxynivalenol (DON), a key virulence factor, which is synthesized in the endoplasmic reticulum (ER). Sey1/atlastin, a dynamin-like GTPase protein, is known to be required for homotypic fusion of ER membranes, but the functions of this protein are unknown in pathogenic fungi. Here, we characterized Sey1/atlastin homologue FgSey1 in F. graminearum Like Sey1/atlastin, FgSey1 is located in the ER. The FgSEY1 deletion mutant exhibited significantly reduced vegetative growth, asexual development, DON biosynthesis, and virulence. Moreover, the ΔFgsey1 mutant was impaired in the formation of normal lipid droplets (LDs) and toxisomes, both of which participate in DON biosynthesis. The GTPase, helix bundle (HB), transmembrane segment (TM), and cytosolic tail (CT) domains of FgSey1 are essential for its function, but only the TM domain is responsible for its localization. Furthermore, the mutants FgSey1K63A and FgSey1T87A lacked GTPase activity and failed to rescue the defects of the ΔFgsey1 mutant. Collectively, our data suggest that the dynamin-like GTPase protein FgSey1 affects the generation of LDs and toxisomes and is required for DON biosynthesis and pathogenesis in F. graminearumIMPORTANCEFusarium graminearum is a major plant pathogen that causes Fusarium head blight (FHB) of wheats worldwide. In addition to reducing the plant yield, F. graminearum infection of wheats also results in the production of deoxynivalenol (DON) mycotoxins, which are harmful to humans and animals and therefore cause great economic losses through pollution of food products and animal feed. At present, effective strategies for controlling FHB are not available. Therefore, understanding the regulation mechanisms of fungal development, pathogenesis, and DON biosynthesis is important for the development of effective control strategies of this disease. In this study, we demonstrated that a dynamin-like GTPase protein Sey1/atlastin homologue, FgSey1, is required for vegetative growth, DON production, and pathogenicity in F. graminearum Our results provide novel information on critical roles of FgSey1 in fungal pathogenicity; therefore, FgSey1 could be a potential target for effective control of the disease caused by F. graminearum.

A Rab5 GTPase module is important for autophagosome closure.

In the conserved autophagy pathway, the double-membrane autophagosome (AP) engulfs cellular components to be delivered for degradation in the lysosome. While only sealed AP can productively fuse with the lysosome, the molecular mechanism of AP closure is currently unknown. Rab GTPases, which regulate all intracellular trafficking pathways in eukaryotes, also regulate autophagy. Rabs function in GTPase modules together with their activators and downstream effectors. In yeast, an autophagy-specific Ypt1 GTPase module, together with a set of autophagy-related proteins (Atgs) and a phosphatidylinositol-3-phosphate (PI3P) kinase, regulates AP formation. Fusion of APs and endosomes with the vacuole (the yeast lysosome) requires the Ypt7 GTPase module. We have previously shown that the Rab5-related Vps21, within its endocytic GTPase module, regulates autophagy. However, it was not clear which autophagy step it regulates. Here, we show that this module, which includes the Vps9 activator, the Rab5-related Vps21, the CORVET tethering complex, and the Pep12 SNARE, functions after AP expansion and before AP closure. Whereas APs are not formed in mutant cells depleted for Atgs, sealed APs accumulate in cells depleted for the Ypt7 GTPase module members. Importantly, depletion of individual members of the Vps21 module results in a novel phenotype: accumulation of unsealed APs. In addition, we show that Vps21-regulated AP closure precedes another AP maturation step, the previously reported PI3P phosphatase-dependent Atg dissociation. Our results delineate three successive steps in the autophagy pathway regulated by Rabs, Ypt1, Vps21 and Ypt7, and provide the first insight into the upstream regulation of AP closure.

Aquaporin1 regulates development, secondary metabolism and stress responses in Fusarium graminearum.

The Ascomycete fungus Fusarium graminearum, the causal agent of Fusarium head blight of wheat and barley, has become a predominant model organism for the study of fungal phytopathogens. Aquaporins (AQPs) have been implicated in the transport of water, glycerol, and a variety of other small molecules in yeast, plants and animals. However, the role of these proteins in phytopathogenic fungi is not well understood. Here, we identified and attempted to elucidate the function of the five aquaporin genes in F. graminearum. The phylogenetic analysis revealed that FgAQPs are divided into two clades, with FgAQP1 in the first clade. The ∆AQP1 mutant formed whitish colonies with longer aerial hyphae and reduced conidiation and perithecium formation. The ∆AQP1 mutant conidia were morphologically abnormal and appeared to undergo abnormal germination. The ∆AQP1 mutant and the wild type strain were equally pathogenic, while the mutant produced significantly higher quantities of deoxynivalenol (DON). The ∆AQP1 mutant also exhibited increased resistance to osmotic and oxidative stress as well as cell-wall perturbing agents. Using FgAQP1-GFP and DAPI staining, we found that FgAQP1 is localized to the nuclear membrane in conidia. Importantly, deletion of FgAQP1 increased the severity of conidium autophagy. Taken together, these results suggest that FgAQP1 is involved in hyphal development, stress responses, secondary metabolism, and sexual and asexual reproduction in F. graminearum. Unlike the ∆AQP1 mutant, the ∆AQP2, ∆AQP3, ∆AQP4 and ∆AQP5 mutants had no variable phenotypes.

Trs130 participates in autophagy through GTPases Ypt31/32 in Saccharomyces cerevisiae.

Trs130 is a specific component of the transport protein particle II complex, which functions as a guanine nucleotide exchange factor (GEF) for Rab GTPases Ypt31/32. Ypt31/32 is known to be involved in autophagy, although the precise mechanism has not been thoroughly studied. In this study, we investigated the potential involvement of Trs130 in autophagy and found that both the cytoplasm-to-vacuole targeting (Cvt) pathway and starvation-induced autophagy were defective in a trs130ts (trs130 temperature-sensitive) mutant. Mutant cells could not transport Atg8 and Atg9 to the pre-autophagosomal structure/phagophore assembly site (PAS) properly, resulting in multiple Atg8 dots and Atg9 dots dispersed in the cytoplasm. Some dots were trapped in the trans-Golgi. Genetic studies showed that the effect of the Trs130 mutation was downstream of Atg5 and upstream of Atg1, Atg13, Atg9 and Atg14 on the autophagic pathway. Furthermore, overexpression of Ypt31 or Ypt32, but not of Ypt1, rescued autophagy defects in trs130ts and trs65ts (Trs130-HA Trs120-myc trs65Δ) mutants. Our data provide mechanistic insight into how Trs130 participates in autophagy and suggest that vesicular trafficking regulated by GTPases/GEFs is important in the transport of autophagy proteins from the trans-Golgi to the PAS.

Bet3 participates in autophagy through GTPase Ypt1 in Saccharomyces cerevisiae.

Three TRAPP (transport protein particle) complexes have been identified in Saccharomyces cerevisiae. GTPases Ypt1 and Ypt31/32 suppress autophagic defects in the mutants of TRAPPIII-specific subunit (Trs85) and TRAPPII-specific subunits (Trs130 and Trs120), respectively. However, the roles of the common TRAPP subunits (which also form the TRAPPI complex) in autophagy and their relationship to Rab GTPases in autophagy remain unclear. As Bet3 (a common TRAPP subunit) cannot be mutated together with either Trs85 or Trs130, we examined starvation-induced autophagy and the cytoplasm-to-vacuole targeting (Cvt) pathway in bet3ts cells. The results demonstrated that GFP-Atg8 was dispersed in the cytoplasm and Ape1 accumulated as a unique dot on the vacuolar membrane in bet3ts cells. Further analysis revealed that Ape1 maturation and GFP-Atg8 processing are defective in these cells. However, prApe1 (precursor form of Ape1) and GFP-Atg8 are protease-accessible in bet3ts cells under starvation, which indicates that Bet3 functions before autophagosome closure. Furthermore, active Ypt1, but not Ypt31, partly rescued the autophagic defects of bet3ts cells. We conclude that Bet3 is involved in autophagy and propose that it participates in autophagy through TRAPP complexes mostly via Ypt1 in yeast.

Ypt1 suppresses defects of vesicle trafficking and autophagy in Ypt6 related mutants.

Ypt/Rab GTPases coordinately regulate vesicle trafficking in yeasts. Previously, Ypt1 was shown to suppress growth defects of Ypt6 and its related mutants (ypt6ts, ric1∆, rgp1∆, ric1∆rgp1∆), but the physiological role of this suppression has not been well studied. We have investigated the effects of Ypt1 on two major trafficking pathways, vesicle trafficking and autophagy, in Ypt6 related mutants. Ypt1 restores Snc1 transport to the plasma membrane via Golgi in the exocytic pathway in Ypt6 related mutants under nutrient rich conditions. Overexpression of Ypt1 suppresses autophagic defects under nutrient starvation conditions with increased GFP-Atg8 sorting to vacuoles and GFP-Atg8 to GFP conversion in Ypt6 related mutants. However, overexpression of Ypt1 does not restore Ypt6 intracellular localisation in rgp1∆ cells. We propose that vesicle trafficking and autophagy are closely connected processes, and Ypt1 and Ypt6 have some similar functions in both cellular processes.

AtPIP1;4 and AtPIP2;4 Cooperatively Mediate H2O2 Transport to Regulate Plant Growth and Disease Resistance.

The rapid production of hydrogen peroxide (H2O2) is a hallmark of plants' successful recognition of pathogen infection and plays a crucial role in innate immune signaling. Aquaporins (AQPs) are membrane channels that facilitate the transport of small molecular compounds across cell membranes. In plants, AQPs from the plasma membrane intrinsic protein (PIP) family are utilized for the transport of H2O2, thereby regulating various biological processes. Plants contain two PIP families, PIP1s and PIP2s. However, the specific functions and relationships between these subfamilies in plant growth and immunity remain largely unknown. In this study, we explore the synergistic role of AtPIP1;4 and AtPIP2;4 in regulating plant growth and disease resistance in Arabidopsis. We found that in plant cells treated with H2O2, AtPIP1;4 and AtPIP2;4 act as facilitators of H2O2 across membranes and the translocation of externally applied H2O2 from the apoplast to the cytoplasm. Moreover, AtPIP1;4 and AtPIP2;4 collaborate to transport bacterial pathogens and flg22-induced apoplastic H2O2 into the cytoplasm, leading to increased callose deposition and enhanced defense gene expression to strengthen immunity. These findings suggest that AtPIP1;4 and AtPIP2;4 cooperatively mediate H2O2 transport to regulate plant growth and immunity.

MYB44 regulates PTI by promoting the expression of EIN2 and MPK3/6 in Arabidopsis.

The plant signaling pathway that regulates pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) involves mitogen-activated protein kinase (MAPK) cascades that comprise sequential activation of several protein kinases and the ensuing phosphorylation of MAPKs, which activate transcription factors (TFs) to promote downstream defense responses. To identify plant TFs that regulate MAPKs, we investigated TF-defective mutants of Arabidopsis thaliana and identified MYB44 as an essential constituent of the PTI pathway. MYB44 confers resistance against the bacterial pathogen Pseudomonas syringae by cooperating with MPK3 and MPK6. Under PAMP treatment, MYB44 binds to the promoters of MPK3 and MPK6 to activate their expression, leading to phosphorylation of MPK3 and MPK6 proteins. In turn, phosphorylated MPK3 and MPK6 phosphorylate MYB44 in a functionally redundant manner, thus enabling MYB44 to activate MPK3 and MPK6 expression and further activate downstream defense responses. Activation of defense responses has also been attributed to activation of EIN2 transcription by MYB44, which has previously been shown to affect PAMP recognition and PTI development. AtMYB44 thus functions as an integral component of the PTI pathway by connecting transcriptional and posttranscriptional regulation of the MPK3/6 cascade.

Mitochondrial Porin Is Involved in Development, Virulence, and Autophagy in Fusarium graminearum.

Mitochondrial porin, the voltage-dependent anion-selective channel (VDAC), is the most abundant protein in the outer membrane, and is critical for the exchange of metabolites and phospholipids in yeast and mammals. However, the functions of porin in phytopathogenic fungi are not known. In this study, we characterized a yeast porin orthologue, Fgporin, in Fusarium graminearum. The deletion of Fgporin resulted in defects in hyphal growth, conidiation, and perithecia development. The Fgporin deletion mutant showed reduced virulence, deoxynivalenol production, and lipid droplet accumulation. In addition, the Fgporin deletion mutant exhibited morphological changes and the dysfunction of mitochondria, and also displayed impaired autophagy in the non-nitrogen medium compared to the wild type. Yeast two-hybrid and bimolecular fluorescence complementation assays indicated that Fgporin interacted with FgUps1/2, but not with FgMdm35. Taken together, these results suggest that Fgporin is involved in hyphal growth, asexual and sexual reproduction, virulence, and autophagy in F. graminearum.

The Endoplasmic Reticulum Cargo Receptor FgErv14 Regulates DON Production, Growth and Virulence in Fusarium graminearum.

Fusarium graminearum is a plant filamentous pathogenic fungi and the predominant causal agent of Fusarium head blight (FHB) in cereals worldwide. The regulators of the secretory pathway contribute significantly to fungal mycotoxin synthesis, development, and virulence. However, their roles in these processes in F. graminearum remain poorly understood. Here, we identified and functionally characterized the endoplasmic reticulum (ER) cargo receptor FgErv14 in F. graminearum. Firstly, it was observed that FgErv14 is mainly localized in the ER. Then, we constructed the FgErv14 deletion mutant (ΔFgerv14) and found that the absence of the FgErv14 caused a serious reduction in vegetative growth, significant defects in asexual and sexual reproduction, and severely impaired virulence. Furthermore, we found that the ΔFgerv14 mutant exhibited a reduced expression of TRI genes and defective toxisome generation, both of which are critical for deoxynivalenol (DON) biosynthesis. Importantly, we found the green fluorescent protein (GFP)-tagged FgRud3 was dispersed in the cytoplasm, whereas GFP-FgSnc1-PEM was partially trapped in the late Golgi in ΔFgerv14 mutant. These results demonstrate that FgErv14 mediates anterograde ER-to-Golgi transport as well as late secretory Golgi-to-Plasma membrane transport and is necessary for DON biosynthesis, asexual and sexual reproduction, vegetative growth, and pathogenicity in F. graminearum.

Phosphorylation of a wheat aquaporin at two sites enhances both plant growth and defense.

Eukaryotic aquaporins share the characteristic of functional multiplicity in transporting distinct substrates and regulating various processes, but the underlying molecular basis for this is largely unknown. Here, we report that the wheat (Triticum aestivum) aquaporin TaPIP2;10 undergoes phosphorylation to promote photosynthesis and productivity and to confer innate immunity against pathogens and a generalist aphid pest. In response to elevated atmospheric CO2 concentrations, TaPIP2;10 is phosphorylated at the serine residue S280 and thereafter transports CO2 into wheat cells, resulting in enhanced photosynthesis and increased grain yield. In response to apoplastic H2O2 induced by pathogen or insect attacks, TaPIP2;10 is phosphorylated at S121 and this phosphorylated form transports H2O2 into the cytoplasm, where H2O2 intensifies host defenses, restricting further attacks. Wheat resistance and grain yield could be simultaneously increased by TaPIP2;10 overexpression or by expressing a TaPIP2;10 phosphomimic with aspartic acid substitutions at S121 and S280, thereby improving both crop productivity and immunity.