scholarly journals The Golgin Protein RUD3 Regulates Fusarium graminearum Growth and Virulence

2021 ◽  
Vol 87 (6) ◽  
Author(s):  
Chenyu Wang ◽  
Yao Wang ◽  
Liyuan Zhang ◽  
Ziyi Yin ◽  
Yuancun Liang ◽  
...  
Keyword(s):  
Fusarium Graminearum ◽  
Vegetative Growth ◽  
Mammalian Cells ◽  
Cellular Localization ◽  
Pathogenic Fungus ◽  
Coiled Coil ◽  
Effective Control ◽  
Content Type ◽  
Ascospore Discharge ◽  
And Function

ABSTRACT 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. graminearum. IMPORTANCE Fusarium 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.

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

2020 ◽  
Vol 86 (11) ◽  
Author(s):  
Xuefa Chong ◽  
Chenyu Wang ◽  
Yao Wang ◽  
Yixiao Wang ◽  
Liyuan Zhang ◽  
...  
Keyword(s):  
Fusarium Graminearum ◽  
Vegetative Growth ◽  
Animal Feed ◽  
Pathogenic Fungus ◽  
Critical Role ◽  
Effective Control ◽  
Economic Losses ◽  
Head Blight ◽  
Fungal Development ◽  
Content Type

ABSTRACT 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. graminearum. IMPORTANCE Fusarium 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.

scholarly journals Roles of the novel coiled-coil protein Rng10 in septum formation during fission yeast cytokinesis

2016 ◽  
Vol 27 (16) ◽  
pp. 2528-2541 ◽  
Author(s):  
Yajun Liu ◽  
I-Ju Lee ◽  
Mingzhai Sun ◽  
Casey A. Lower ◽  
Kurt W. Runge ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Rho Gtpases ◽  
Cellular Localization ◽  
Affinity Purification ◽  
Coiled Coil ◽  
The Novel ◽  
Septum Formation ◽  
Division Site ◽  
And Function

Rho GAPs are important regulators of Rho GTPases, which are involved in various steps of cytokinesis and other processes. However, regulation of Rho-GAP cellular localization and function is not fully understood. Here we report the characterization of a novel coiled-coil protein Rng10 and its relationship with the Rho-GAP Rga7 in fission yeast. Both rng10Δ and rga7Δ result in defective septum and cell lysis during cytokinesis. Rng10 and Rga7 colocalize on the plasma membrane at the cell tips during interphase and at the division site during cell division. Rng10 physically interacts with Rga7 in affinity purification and coimmunoprecipitation. Of interest, Rga7 localization is nearly abolished without Rng10. Moreover, Rng10 and Rga7 work together to regulate the accumulation and dynamics of glucan synthases for successful septum formation in cytokinesis. Our results show that cellular localization and function of the Rho-GAP Rga7 are regulated by a novel protein, Rng10, during cytokinesis in fission yeast.

scholarly journals Transcription Factor RFX1 Is Crucial for Maintenance of Genome Integrity in Fusarium graminearum

2014 ◽  
Vol 13 (3) ◽  
pp. 427-436 ◽  
Author(s):  
Kyunghun Min ◽  
Hokyoung Son ◽  
Jae Yun Lim ◽  
Gyung Ja Choi ◽  
Jin-Cheol Kim ◽  
...  
Keyword(s):  
Dna Damage ◽  
Transcription Factor ◽  
Fusarium Graminearum ◽  
Fluorescent Protein ◽  
Pathogenic Fungus ◽  
Genome Integrity ◽  
Deletion Mutants ◽  
Factor X ◽  
Content Type ◽  
Septum Formation

ABSTRACT The survival of cellular organisms depends on the faithful replication and transmission of DNA. Regulatory factor X (RFX) transcription factors are well conserved in animals and fungi, but their functions are diverse, ranging from the DNA damage response to ciliary gene regulation. We investigated the role of the sole RFX transcription factor, RFX1, in the plant-pathogenic fungus Fusarium graminearum . Deletion of rfx1 resulted in multiple defects in hyphal growth, conidiation, virulence, and sexual development. Deletion mutants of rfx1 were more sensitive to various types of DNA damage than the wild-type strain. Septum formation was inhibited and micronuclei were produced in the rfx1 deletion mutants. The results of the neutral comet assay demonstrated that disruption of rfx1 function caused spontaneous DNA double-strand breaks (DSBs). The transcript levels of genes involved in DNA DSB repair were upregulated in the rfx1 deletion mutants. DNA DSBs produced micronuclei and delayed septum formation in F. graminearum . Green fluorescent protein (GFP)-tagged RFX1 localized in nuclei and exhibited high expression levels in growing hyphae and conidiophores, where nuclear division was actively occurring. RNA-sequencing-based transcriptomic analysis revealed that RFX1 suppressed the expression of many genes, including those required for the repair of DNA damage. Taken together, these findings indicate that the transcriptional repressor rfx1 performs crucial roles during normal cell growth by maintaining genome integrity.

scholarly journals A specific basal body linker protein provides the connection function for basal body inheritance in trypanosomes

2021 ◽  
Vol 118 (8) ◽  
pp. e2014040118
Author(s):  
De-Hua Lai ◽  
Flavia Moreira-Leite ◽  
Zhi-Shen Xu ◽  
Jiong Yang ◽  
Keith Gull
Keyword(s):  
Basal Body ◽  
Mammalian Cells ◽  
De Novo ◽  
Coiled Coil ◽  
Centriole Duplication ◽  
Cytoskeletal Component ◽  
Dividing Cells ◽  
Orthogonal Orientation ◽  
And Function ◽  
Additional Role

Centrioles and basal bodies (CBBs) are found in physically linked pairs, and in mammalian cells intercentriole connections (G1–G2 tether and S–M linker) regulate centriole duplication and function. In trypanosomes BBs are not associated with the spindle and function in flagellum/cilia nucleation with an additional role in mitochondrial genome (kinetoplast DNA [kDNA]) segregation. Here, we describe BBLP, a BB/pro-BB (pBB) linker protein in Trypanosoma brucei predicted to be a large coiled-coil protein conserved in the kinetoplastida. Colocalization with the centriole marker SAS6 showed that BBLP localizes between the BB/pBB pair, throughout the cell cycle, with a stronger signal in the old flagellum BB/pBB pair. Importantly, RNA interference (RNAi) depletion of BBLP leads to a conspicuous splitting of the BB/pBB pair associated only with the new flagellum. BBLP RNAi is lethal in the bloodstream form of the parasite and perturbs mitochondrial kDNA inheritance. Immunogold labeling confirmed that BBLP is localized to a cytoskeletal component of the BB/pBB linker, and tagged protein induction showed that BBLP is incorporated de novo in both new and old flagella BB pairs of dividing cells. We show that the two aspects of CBB disengagement—loss of orthogonal orientation and ability to separate and move apart—are consistent but separable events in evolutionarily diverse cells and we provide a unifying model explaining centriole/BB linkage differences between such cells.

scholarly journals CmAim24 Is Essential for Mitochondrial Morphology, Conidiogenesis, and Mycoparasitism in Coniothyrium minitans

2019 ◽  
Vol 86 (5) ◽  
Author(s):  
Xiaoxiang Yang ◽  
Huizhang Zhao ◽  
Chenwei Luo ◽  
Lei Du ◽  
Jiasen Cheng ◽  
...  
Keyword(s):  
Cellular Localization ◽  
Sequence Similarity ◽  
Critical Role ◽  
Gene Replacement ◽  
Phytopathogenic Fungus ◽  
Mitochondrial Morphology ◽  
Coniothyrium Minitans ◽  
High Sequence Similarity ◽  
Content Type ◽  
And Function

ABSTRACT Coniothyrium minitans is an important mycoparasite of the notorious phytopathogenic fungus Sclerotinia sclerotiorum. The mycoparasitism system of C. minitans-S. sclerotiorum is unique and important in probing fungi and fungal interactions. Here, we report a conidiation-deficient mutant, ZS-1TN1961, which was screened from a transfer DNA (T-DNA) insertional library of C. minitans. A single-copy gene, encoding a protein with high sequence similarity to Aim24 (altered inheritance of mitochondria protein 24) in Saccharomyces cerevisiae, was disrupted by T-DNA insertion in this mutant. Gene replacement and complementation experiments confirmed that mutants lacking CmAim24 exhibited significantly reduced conidial production and germination as well as reduced sclerotial mycoparasitic ability. Furthermore, cellular localization assays showed that CmAim24 localized to mitochondria, and abnormal mitochondria were observed in the ΔCmAim24 mutant. The ΔCmAim24 mutant exhibited significant accumulation of reactive oxygen species (ROS) and a reduced ATP content in mycelia. In summary, our results suggest that CmAim24 plays a key role in mitochondrial architecture and function, conidiogenesis, and mycoparasitism in C. minitans. IMPORTANCE Aim24 proteins are involved in mitochondrial biogenesis and accumulate between the two membranes of a mitochondrion. Their function in prokaryotes and filamentous fungi is as yet unknown. In the present study, we characterized an Aim24 protein, CmAim24, in the mycoparasite Coniothyrium minitans and proved its critical role in mitochondrial morphology and function, conidiogenesis, conidial germination, and mycoparasitism to S. sclerotiorum.

scholarly journals Mid1, a Mechanosensitive Calcium Ion Channel, Affects Growth, Development, and Ascospore Discharge in the Filamentous Fungus Gibberella zeae

2011 ◽  
Vol 10 (6) ◽  
pp. 832-841 ◽  
Author(s):  
Brad Cavinder ◽  
Ahmed Hamam ◽  
Roger R. Lew ◽  
Frances Trail
Keyword(s):  
Ion Channel ◽  
Calcium Ion ◽  
Vegetative Growth ◽  
Gibberella Zeae ◽  
Calcium Flux ◽  
Wild Type ◽  
Content Type ◽  
Ascospore Discharge ◽  
Ascospore Development

ABSTRACT The role of Mid1, a stretch-activated ion channel capable of being permeated by calcium, in ascospore development and forcible discharge from asci was examined in the pathogenic fungus Gibberella zeae (anamorph Fusarium graminearum ). The Δ mid1 mutants exhibited a >12-fold reduction in ascospore discharge activity and produced predominately abnormal two-celled ascospores with constricted and fragile septae. The vegetative growth rate of the mutants was ∼50% of the wild-type rate, and production of macroconidia was >10-fold lower than in the wild type. To better understand the role of calcium flux, Δ mid1 Δ cch1 double mutants were also examined, as Cch1, an L-type calcium ion channel, is associated with Mid1 in Saccharomyces cerevisiae . The phenotype of the Δ mid1 Δ cch1 double mutants was similar to but more severe than the phenotype of the Δ mid1 mutants for all categories. Potential and current-voltage measurements were taken in the vegetative hyphae of the Δ mid1 and Δ cch1 mutants and the wild type, and the measurements for all three strains were remarkably similar, indicating that neither protein contributes significantly to the overall electrical properties of the plasma membrane. Pathogenicity of the Δ mid1 and Δ mid1 Δ cch1 mutants on the host (wheat) was not affected by the mutations. Exogenous calcium supplementation partially restored the ascospore discharge and vegetative growth defects for all mutants, but abnormal ascospores were still produced. These results extend the known roles of Mid1 to ascospore development and forcible discharge. However, Neurospora crassa Δ mid1 mutants were also examined and did not exhibit defects in ascospore development or in ascospore discharge. In comparison to ion channels in other ascomycetes, Mid1 shows remarkable adaptability of roles, particularly with regard to niche-specific adaptation.

scholarly journals O-Polysaccharide Glycosylation Is Required for Stability and Function of the Collagen Adhesin EmaA of Aggregatibacter actinomycetemcomitans

2012 ◽  
Vol 80 (8) ◽  
pp. 2868-2877 ◽  
Author(s):  
Gaoyan Tang ◽  
Teresa Ruiz ◽  
Keith P. Mintz
Keyword(s):  
Cellular Localization ◽  
Proteolytic Enzymes ◽  
Aggregatibacter Actinomycetemcomitans ◽  
Binding Activity ◽  
Mrna Levels ◽  
Collagen Binding ◽  
Content Type ◽  
Protein Levels ◽  
Mutant Strains ◽  
And Function

ABSTRACTAggregatibacter actinomycetemcomitansis hypothesized to colonize through the interaction with collagen and establish a reservoir for further dissemination. The trimeric adhesin EmaA ofA. actinomycetemcomitansbinds to collagen and is modified with sugars mediated by an O-antigen polysaccharide ligase (WaaL) that is associated with lipopolysaccharide (LPS) biosynthesis (G. Tang and K. Mintz, J. Bacteriol.192:1395–1404, 2010). This investigation characterized the function and cellular localization of EmaA glycosylation. The interruption of LPS biogenesis by using genetic and pharmacological methods changed the amount and biophysical properties of EmaA molecules in the outer membrane. InrmlCandwaaLmutant strains, the membrane-associated EmaA was reduced by 50% compared with the wild-type strain, without changes in mRNA levels. The membrane-associated EmaA protein levels were recovered by complementation with the corresponding O-polysaccharide (O-PS) biosynthetic genes. In contrast, another trimeric autotransporter, epithelial adhesin ApiA, was not affected in the same mutant background. The inhibition of undecaprenyl pyrophosphate recycling by bacitracin resulted in a similar decrease in the membrane-associated EmaA protein. This effect was reversed by removal of the compound. A significant decrease in collagen binding activity was observed in strains expressing the nonglycosylated form of EmaA. Furthermore, the electrophoretic mobility shifts of the EmaA monomers found in the O-PS mutant strains were associated only with the membrane-associated protein and not with the cytoplasmic pre-EmaA protein, suggesting that this modification does not occur in the cytoplasm. The glycan modification of EmaA appears to be required for collagen binding activity and protection of the protein against degradation by proteolytic enzymes.

scholarly journals Invadosome-Mediated Human Extracellular Matrix Degradation by Entamoeba histolytica

2018 ◽  
Vol 86 (9) ◽  
Author(s):  
Muhammad M. Hasan ◽  
Jose E. Teixeira ◽  
Christopher D. Huston
Keyword(s):  
Extracellular Matrix ◽  
Entamoeba Histolytica ◽  
Mammalian Cells ◽  
Protozoan Parasite ◽  
Human Colon ◽  
Colonic Tissue ◽  
Colon Tissue ◽  
Content Type ◽  
Tissue Invasion ◽  
And Function

ABSTRACT Entamoeba histolytica is a protozoan parasite that causes invasive amoebiasis when it invades the human colon. Tissue invasion requires a shift from an adhesive lifestyle in the colonic lumen to a motile and extracellular matrix (ECM) degradative lifestyle in the colonic tissue layers. How the parasite regulates these two lifestyles is largely unknown. Previously, we showed that silencing the E. histolytica surface metalloprotease EhMSP-1 results in parasites that are hyperadherent and less motile. To better understand the molecular mechanism of this phenotype, we now show that the parasites with EhMSP-1 silenced cannot efficiently form specialized dot-like polymerized actin (F actin) structures upon interaction with the human ECM component fibronectin. We characterized these F actin structures and found that they are very short-lived structures that are the sites of fibronectin degradation. Motile mammalian cells form F actin structures called invadosomes that are similar in stability and function to these amoebic actin dots. Therefore, we propose here that E. histolytica forms amoebic invadosomes to facilitate colonic tissue invasion.

scholarly journals Microtubule-bundling protein Spef1 enables mammalian ciliary central apparatus formation

2018 ◽  
Vol 11 (1) ◽  
pp. 67-77 ◽  
Author(s):  
Jianqun Zheng ◽  
Hao Liu ◽  
Lei Zhu ◽  
Yawen Chen ◽  
Huijie Zhao ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Coiled Coil ◽  
Cell Movement ◽  
Ependymal Cells ◽  
Coiled Coil Region ◽  
Evolutionarily Conserved ◽  
Flow Through ◽  
Ependymal Cilia ◽  
And Function ◽  
Signal Transductions

Abstract Cilia are cellular protrusions containing nine microtubule (MT) doublets and function to propel cell movement or extracellular liquid flow through beating or sense environmental stimuli through signal transductions. Cilia require the central pair (CP) apparatus, consisting of two CP MTs covered with projections of CP proteins, for planar strokes. How the CP MTs of such ‘9 + 2’ cilia are constructed, however, remains unknown. Here we identify Spef1, an evolutionarily conserved microtubule-bundling protein, as a core CP MT regulator in mammalian cilia. Spef1 was selectively expressed in mammalian cells with 9 + 2 cilia and specifically localized along the CP. Its depletion in multiciliated mouse ependymal cells by RNAi completely abolished the CP MTs and markedly attenuated ciliary localizations of CP proteins such as Hydin and Spag6, resulting in rotational beat of the ependymal cilia. Spef1, which binds to MTs through its N-terminal calponin-homologous domain, formed homodimers through its C-terminal coiled coil region to bundle and stabilize MTs. Disruption of either the MT-binding or the dimerization activity abolished the ability of exogenous Spef1 to restore the structure and functions of the CP apparatus. We propose that Spef1 bundles and stabilizes central MTs to enable the assembly and functions of the CP apparatus.

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