Exo70 Proteins Research Articles

The RING-Type E3 Ligase BOI Interacts with EXO70E2 and Mediates Its Ubiquitination in Arabidopsis.

The exocyst is a hetero-octameric complex that exhibits significant functional diversity in regulating biological processes and defense responses. In plants, the EXO70 proteins are important components of the exocyst complex and are involved in membrane trafficking, biotic and abiotic interactions, as well as cell wall formation. A previous study has indicated that a member of the EXO subfamily, EXO70E2, interacts with RIN4 to mediate plant immunity. In this study, we found that EXO70E2 interacts with the RING-type E3 ligase Botrytis susceptible1 interactor (BOI), and the C-terminal domain of BOI is necessary for its interaction with EXO70E2. Moreover, the protein level of EXO70E2 was degraded and ubiquitinated by BOI in vitro. Collectively, our study reveals a mechanism for regulating the stability of EXO70E2 by a RING-type E3 ligase BOI-mediated ubiquitination.

Zinc-finger (ZiF) fold secreted effectors form a functionally diverse family across lineages of the blast fungus Magnaporthe oryzae.

Filamentous plant pathogens deliver effector proteins into host cells to suppress host defence responses and manipulate metabolic processes to support colonization. Understanding the evolution and molecular function of these effectors provides knowledge about pathogenesis and can suggest novel strategies to reduce damage caused by pathogens. However, effector proteins are highly variable, share weak sequence similarity and, although they can be grouped according to their structure, only a few structurally conserved effector families have been functionally characterized to date. Here, we demonstrate that Zinc-finger fold (ZiF) secreted proteins form a functionally diverse effector family in the blast fungus Magnaporthe oryzae. This family relies on the Zinc-finger motif for protein stability and is ubiquitously present in blast fungus lineages infecting 13 different host species, forming different effector tribes. Homologs of the canonical ZiF effector, AVR-Pii, from rice infecting isolates are present in multiple M. oryzae lineages. Wheat infecting strains of the fungus also possess an AVR-Pii like allele that binds host Exo70 proteins and activates the immune receptor Pii. Furthermore, ZiF tribes may vary in the proteins they bind to, indicating functional diversification and an intricate effector/host interactome. Altogether, we uncovered a new effector family with a common protein fold that has functionally diversified in lineages of M. oryzae. This work expands our understanding of the diversity of M. oryzae effectors, the molecular basis of plant pathogenesis and may ultimately facilitate the development of new sources for pathogen resistance.

OsEXO70L2 is required for large lateral root formation and arbuscular mycorrhiza establishment in rice

As a major subunit of the exocyst complex, members of the EXO70 family have mainly been shown to play roles in cell polarity and morphogenesis in Arabidopsis, but their roles in plant endosymbiosis, such as with arbuscular mycorrhizal fungi (AMF), have rarely been reported. Here, using knockout and overexpression lines, we show that OsEXO70L2, which encodes a divergent EXO70 protein in rice, controls the number of primary roots and is essential for large lateral root formation. Furthermore, the OsEXO70L2 mutant sr1 displayed rare internal AMF hyphae and no arbuscules. We also found that AMF sporulation can occur in roots despite low colonization and that AMF colonization and sporulation are modulated by photoperiod and co-culture with clover. Finally, genes related to auxin homeostasis were found to be affected in the OsEXO70L2 knockout or overexpression lines, suggesting that auxin is at least partly responsible for the phenotypes. This study provides new perspectives on the role of the exocyst complex during root development and AM in rice.

Interplay of EXO70 and MLO proteins modulates trichome cell wall composition and susceptibility to powdery mildew.

Exocyst component of 70-kDa (EXO70) proteins are constituents of the exocyst complex implicated in vesicle tethering during exocytosis. MILDEW RESISTANCE LOCUS O (MLO) proteins are plant-specific calcium channels and some MLO isoforms enable fungal powdery mildew pathogenesis. We here detected an unexpected phenotypic overlap of Arabidopsis thaliana exo70H4 and mlo2 mlo6 mlo12 triple mutant plants regarding the biogenesis of leaf trichome secondary cell walls. Biochemical and Fourier transform infrared spectroscopic analyses corroborated deficiencies in the composition of trichome cell walls in these mutants. Transgenic lines expressing fluorophore-tagged EXO70H4 and MLO exhibited extensive colocalization of these proteins. Furthermore, mCherry-EXO70H4 mislocalized in trichomes of the mlo triple mutant and, vice versa, MLO6-GFP mislocalized in trichomes of the exo70H4 mutant. Expression of GFP-marked PMR4 callose synthase, a known cargo of EXO70H4-dependent exocytosis, revealed reduced cell wall delivery of GFP-PMR4 in trichomes of mlo triple mutant plants. In vivo protein-protein interaction assays in plant and yeast cells uncovered isoform-preferential interactions between EXO70.2 subfamily members and MLO proteins. Finally, exo70H4 and mlo6 mutants, when combined, showed synergistically enhanced resistance to powdery mildew attack. Taken together, our data point to an isoform-specific interplay of EXO70 and MLO proteins in the modulation of trichome cell wall biogenesis and powdery mildew susceptibility.

A blast fungus zinc-finger fold effector binds to a hydrophobic pocket in host Exo70 proteins to modulate immune recognition in rice

Exocytosis plays an important role in plant-microbe interactions, in both pathogenesis and symbiosis. Exo70 proteins are integral components of the exocyst, an octameric complex that mediates tethering of vesicles to membranes in eukaryotes. Although plant Exo70s are known to be targeted by pathogen effectors, the underpinning molecular mechanisms and the impact of this interaction on infection are poorly understood. Here, we show the molecular basis of the association between the effector AVR-Pii of the blast fungus Maganaporthe oryzae and rice Exo70 alleles OsExo70F2 and OsExo70F3, which is sensed by the immune receptor pair Pii via an integrated RIN4/NOI domain. The crystal structure of AVR-Pii in complex with OsExo70F2 reveals that the effector binds to a conserved hydrophobic pocket in Exo70, defining an effector/target binding interface. Structure-guided and random mutagenesis validates the importance of AVR-Pii residues at the Exo70 binding interface to sustain protein association and disease resistance in rice when challenged with fungal strains expressing effector mutants. Furthermore, the structure of AVR-Pii defines a zinc-finger effector fold (ZiF) distinct from the MAX (Magnaporthe Avrs and ToxB-like) fold previously described for a majority of characterized M. oryzae effectors. Our data suggest that blast fungus ZiF effectors bind a conserved Exo70 interface to manipulate plant exocytosis and that these effectors are also baited by plant immune receptors, pointing to new opportunities for engineering disease resistance.

Deregulation of Exo70 Facilitates Innate and Acquired Cisplatin Resistance in Epithelial Ovarian Cancer by Promoting Cisplatin Efflux.

Simple SummaryInnate and acquired platinum resistance are the leading causes of epithelial ovarian cancer (EOC) mortality. However, the mechanisms remain elusive. Here we found that Exo70, a key subunit of the exocyst, is upregulated in EOC and promotes cisplatin efflux to facilitate innate resistance. More interestingly, cisplatin could downregulate Exo70 to sustain cell sensitivity. However, this function was hampered during prolonged cisplatin treatment, which in turn stabilized Exo70 to facilitate the acquired cisplatin resistance of EOC cells. Our study potentiates Exo70 as a promising target to overcome cisplatin resistance in EOC.Whilst researches elucidating a diversity of intracellular mechanisms, platinum-resistant epithelial ovarian cancer (EOC) remains a major challenge in the treatment of ovarian cancer. Here we report that Exo70, a key subunit of the exocyst complex, contributes to both innate and acquired cisplatin resistance of EOC. Upregulation of Exo70 is observed in EOC tissues and is related to platinum resistance and progression-free survival of EOC patients. Exo70 suppressed the cisplatin sensitivity of EOC cells through promoting exocytosis-mediated efflux of cisplatin. Moreover, cisplatin-induced autophagy-lysosomal degradation of Exo70 protein by modulating phosphorylation of AMPK and mTOR, thereby reducing the cellular resistance. However, the function was hampered during prolonged cisplatin treatment, which in turn stabilized Exo70 to facilitate the acquired cisplatin resistance of EOC cells. Knockdown of Exo70, or inhibiting exocytosis by Exo70 inhibitor Endosidin2, reversed the cisplatin resistance of EOC cells both in vitro and in vivo. Our results suggest that Exo70 overexpression and excessive stability contribute to innate and acquired cisplatin resistance through the increase in cisplatin efflux, and targeting Exo70 might be an approach to overcome cisplatin resistance in EOC treatment.

The resistance associated protein RIN4 promotes the extracellular transport of AtEXO70E2

RIN4 is an important immunomodulator in Arabidopsis, which is targeted by multiple pathogenic effectors, and consequently guarded by different immune receptors. Although RIN4 plays a significant role in plant immunity, its molecular function is not fully understood. We found that RIN4 interacts with the exocyst subunit EXO70E2. Transiently expressed RIN4 can recruits EXO70E2 vesicles to the plasma membrane, and promote the transport of the vesicles to the extracellular matrix. RIN4 also can decrease the protein level of EXO70E2. Base on the fact that EXO70 proteins positively mediates plant immunity, the function of RIN4 is to promote the extracellular export of defense related vesicles. Pathogens will secret effectors to modify or cleavage it to interfere this exocytosis.

GIV/Girdin and Exo70 Collaboratively Regulate the Mammalian Polarized Exocytic Machinery.

SummaryPolarized exocytosis is a fundamental process by which membranes and cargo proteins are delivered to the cell surface with precise spatial control. Although the need for the octameric exocyst complex is conserved from yeast to humans, what imparts spatial control is known only in yeast, i.e., a polarity scaffold called Bem1p. We demonstrate here that the mammalian scaffold protein, GIV/Girdin, fulfills the key criteria and functions of its yeast counterpart Bem1p; both bind Exo70 proteins via similar short-linear interaction motifs, and each prefers its evolutionary counterpart. Selective disruption of the GIV⋅Exo-70 interaction derails the delivery of the metalloprotease MT1-MMP to invadosomes and impairs collagen degradation and haptotaxis through basement membrane matrix. GIV's interacting partners reveal other components of polarized exocytosis in mammals. Findings expose how the exocytic functions aid GIV's pro-metastatic functions and how signal integration via GIV may represent an evolutionary advancement of the exocytic process in mammals.

The Pseudomonas Syringae Effector AvrPtoB Associates With and Ubiquitinates Arabidopsis Exocyst Subunit EXO70B1.

Many bacterial pathogens secret effectors into host cells to disable host defenses and thus promote infection. The exocyst complex functions in the transport and secretion of defense molecules, and loss of function of the EXO70B1 subunit leads to autoimmunity by activation of a truncated Toll/interleukin-1 receptor–nucleotide-binding sequence protein (TIR-NBS2; herein referred to as TN2). Here, we show that EXO70B1 is required for pathogen-associated molecular pattern-triggered immune responses in Arabidopsis thaliana. The effector AvrPtoB, an E3 ligase from Pseudomonas syringae pv. tomato (Pto) strain DC3000, associates with EXO70B1. AvrPtoB ubiquitinates EXO70B1 and mediates EXO70B1 degradation via the host’s 26S proteasome in a manner requiring E3 ligase activity. AvrPtoB enhances Pto DC3000 virulence by overcoming EXO70B1-mediated resistance. Moreover, overexpression of AvrPtoB in Arabidopsis leads to autoimmunity, which is partially dependent on TN2. Expression of TN2 in tobacco (Nicotiana tabacum and Nicotiana benthamiana) triggers strong and rapid cell death, which is suppressed by co-expression with EXO70B1 but reoccurs when co-expressed with AvrPtoB. Taken together, our data highlight that AvrPtoB targets the Arabidopsis thaliana EXO70 protein family member EXO70B1 to manipulate the defense molecule secretion machinery or immunity.

Overexpression of the dominant negative nbexo70d1 mutantionconfers tolerance to salt stress in transgenic tobacco

The vesicle trafficking process, which involves exocytotic and endocytotic pathways, has been reported to play a role in regulating plant responses to different environmental stresses. The Exo70 protein is important for the localization of the exocyst in the plasma membrane; however, its role in the physiology of stress tolerance is currently unclear. In this study, we characterized NbExo70D1, an Exo70 gene from tobacco (Nicotiana benthamiana). It was shown to have a role in the plant response to salt stress. More specifically, tolerance to salt stress is conferred by the overexpression of the dominant negative nbexo70d1 domain D mutation in transgenic tobacco. In addition, a reduced accumulation of reactive oxygen species (ROS) under salt treatment was observed in the transgenic lines compared to the wild type. Treatment with diphenylene iodonium, an NADPH oxidase inhibitor, resulted in a decrease in salt stress-triggered ROS production in the roots of both wild type tobacco and transgenic tobacco. Furthermore, there was a reduction in NADPH oxidase activity in the transgenic plants under salt treatment, which indicates NbExo70D1 is involved in NADPH oxidase-mediated ROS production. We also characterized the tissue-specific expression patterns of NbExo70D1 during salt stress response by expressing the ProNbExo70D1-β-glucuronidase reporter construct in plants. Importantly, the GFP-NbExo70D1 fusion protein was localized in both the plasma membrane and the cytoplasm; expressing the dominant negative mutation disrupted the interaction between NbExo70D1 protein and the plasma membrane. Overall, our study suggests that Exo70 plays an important role in regulating the production and transmission of ROS as part of a salt stress response in plants.

Characterization of the Interaction between ISTL1 and Exo70E2 in Arabidopsis

Internal and external stimuli at the plasma membrane are the cause of dynamic changes that affect the composition of the plasma membrane which is responsible for the proper responses, survival, and development of an organism. The Endosomal Sorting Complex Required for Transport, otherwise known as ESCRT, is a highly conserved pathway in all eukaryotes. This pathway allows for endocytosis, and endosomal trafficking which tags proteins for degradation. It is also responsible for the sorting of endocytic vesicles to the vacuole. This mediates the responses of plants. It has also been noted that there is a possible relationship between the ESCRT pathway and the exocyst. The exocyst is an octameric protein complex that directs vesicles after the Golgi complex to specific destinations of the the plasma membrane. It has recently been proposed that Exo70 proteins localize vesicle traffic.Support or Funding InformationNational Science FoundationNIH Minority Access to Research (MARC) 5 T34 GM 8395‐29 California State University NorthridgeOtegui LabThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Use Endosidin2 to Study Plant Exocytosis and Vacuolar Trafficking.

Membrane trafficking is essential for multiple aspects of plant growth and development. Some small molecules have been widely used to study the mechanisms of membrane trafficking in plants. For example, short-term treatment with small molecules combined with live cell imaging has been shown to be very useful in understanding the dynamic processes of membrane trafficking. Small molecule Endosidin2 (ES2) has been found to target Exo70 protein and inhibit exocytosis and promote vacuolar trafficking in plants. Here we describe the method of using short-term ES2 treatment combined with live cell imaging to study plant exocytosis and vacuolar trafficking in Arabidopsis seedlings expressing a cargo protein PIN2:GFP.

Expression and Functional Analysis of a Novel Group of Legume-specific WRKY and Exo70 Protein Variants from Soybean.

Legumes fix atmospheric nitrogen through symbiosis with microorganisms and contain special traits in nitrogen assimilation and associated processes. Recently, we have reported a novel WRKY-related protein (GmWRP1) and a new clade of Exo70 proteins (GmExo70J) from soybean with homologs found only in legumes. GmWRP1 and some of the GmExo70J proteins are localized to Golgi apparatus through a novel N-terminal transmembrane domain. Here, we report further analysis of expression and functions of the novel GmWRP1 and GmExo70J genes. Promoter-GUS analysis in Arabidopsis revealed distinct tissue-specific expression patterns of the GmExo70J genes not only in vegetative but also in reproductive organs including mature tissues, where expression of previously characterized Exo70 genes is usually absent. Furthermore, expression of some GmExo70J genes including GmExo70J1, GmExo70J6 and GmExo70J7 increases greatly in floral organ-supporting receptacles during the development and maturation of siliques, indicating a possible role in seed development. More importantly, suppression of GmWRP1, GmExo70J7, GmExo70J8 and GmExo70J9 expression in soybean using virus- or artificial microRNA-mediated gene silencing resulted in accelerated leaf senescence and reduced nodule formation. These results strongly suggest that legume-specific GmWRP1 and GmExo70J proteins play important roles not only in legume symbiosis but also in other processes critical for legume growth and development.

Rice Exo70 interacts with a fungal effector, AVR-Pii, and is required for AVR-Pii-triggered immunity.

Vesicle trafficking including the exocytosis pathway is intimately associated with host immunity against pathogens. However, we still have insufficient knowledge about how it contributes to immunity, and how pathogen factors affect it. In this study, we explore host factors that interact with the Magnaporthe oryzae effector AVR-Pii. Gel filtration chromatography and co-immunoprecipitation assays identified a 150kDa complex of proteins in the soluble fraction comprising AVR-Pii and OsExo70-F2 and OsExo70-F3, two rice Exo70 proteins presumably involved in exocytosis. Simultaneous knockdown of OsExo70-F2 and F3 totally abrogated Pii immune receptor-dependent resistance, but had no effect on Pia- and Pik-dependent resistance. Knockdown levels of OsExo70-F3 but not OsExo70-F2 correlated with reduction of Pii function, suggesting that OsExo70-F3 is specifically involved in Pii-dependent resistance. Under our current experimental conditions, over-expression of AVR-Pii or knockdown of OsExo70-F2 and -F3 genes in rice did not affect the virulence of compatible isolates of M.oryzae. AVR-Pii interaction with OsExo70-F3 appears to play a crucial role in immunity triggered by Pii, suggesting a role for OsExo70 as a decoy or helper in Pii/AVR-Pii interactions.

Identification and characterization of a novel group of legume-specific, Golgi apparatus-localized WRKY and Exo70 proteins from soybean.

Many plant genes belong to families that arise from extensive proliferation and diversification allowing the evolution of functionally new proteins. Here we report the characterization of a group of proteins evolved from WRKY and exocyst complex subunit Exo70 proteins through fusion with a novel transmembrane (TM) domain in soybean (Glycine max). From the soybean genome, we identified a novel WRKY-related protein (GmWRP1) that contains a WRKY domain with no binding activity for W-box sequences. GFP fusion revealed that GmWRP1 was targeted to the Golgi apparatus through its N-terminal TM domain. Similar Golgi-targeting TM domains were also identified in members of a new subfamily of Exo70J proteins involved in vesicle trafficking. The novel TM domains are structurally most similar to the endosomal cytochrome b561 from birds and close homologues of GmWRP1 and GmEx070J proteins with the novel TM domain have only been identified in legumes. Transient expression of some GmExo70J proteins or the Golgi-targeting TM domain in tobacco altered the subcellular structures labelled by a fluorescent Golgi marker. GmWRP1 transcripts were detected at high levels in roots, flowers, pods, and seeds, and the expression levels of GmWRP1 and GmExo70J genes were elevated with increased age in leaves. The legume-specific, Golgi apparatus-localized GmWRP1 and GmExo70J proteins are probably involved in Golgi-mediated vesicle trafficking of biological molecules that are uniquely important to legumes.

Comparison of <italic>EXO70</italic> duplication between <italic>Arabidopsis thaliana</italic> and <italic>Oryza sativa</italic>

EXO70, an important component of the exocyst complex, is encoded by a single gene in humans, mice and yeast. To understand EXO70 gene duplication in plant genomes, we identified and analyzed all OsEXO70 and AtEXO70 genes in Arabidopsis thaliana and Oryza sativa . A total of 23 different EXO70 genes were identified in the A. thaliana genome, of which 15 were present on the ‘-’ strand. We detected 27 different mature mRNAs of EXO70 arising from alternative splicing. In O. sativa , 38 OsEXO70 genes coding for proteins comprising more than 412 amino acids were found to be distributed across the 12 rice chromosomes. Of these genes, 29 were located on the ‘+’ strand. Interestingly, a product of large-scale genome duplication—an approximately 2-Mbp-long fragment including both OsEXO70H3 and OsEXO70H4 , was highly conserved between chromosomes 11 and 12. The gene duplication mode of OsEXO70 is more complex than that of AtEXO70 . Although synonymous codon bias exists in both AtEXO70 and OsEXO70 , codon preferences differ between them. All EXO70 proteins contain a pfam03081 domain, indicating that these proteins may have the same or similar functions. We conclude that EXO70 has been duplicated multiple times during the evolution of both A. thaliana and O. sativa , with more duplications occurring in O. sativa than in A. thaliana . Additionally, a greater number of gene duplication mechanisms are operative for OsEXO70 than for AtEXO70 . We also propose that the pfam03081 domain may play an essential role in EXO70 functional determination. Finally, many differences in synonymous codon preference exist between AtEXO70 and OsEXO70 .

Cellular Scale Biophysical Models of Membrane Sculpting by the Proteins During Endocytosis and Exocytosis

We employ coarse-grained molecular dynamics (CGMD) simulations and continuum mechanics methods to quantify the membrane-bending action of exo70. CGMD simulations show that exo70 produces the strongest curvature when bound to a key lipid PIP2, and when dimerized. To relate these simulations to experiments, we first quantify the deformation field generated by exo70 using CGMD simulation, and then investigate the resulting cellular morphologies using a continuum mechanics model for membrane bending. In the first step, we characterize the deformation field by directly analyzing curved membrane conformations generated by CGMD simulations of mutant and wild-type exo70 on mixed lipid bilayers. Monte Carlo simulations yield membrane morphologies which we relate to cryo-transmission electron microscopy images of exo70 remodeling a GUV. This model agrees with experiments which show that PIP2 binding and dimerization are necessary for generating membrane tubulues and protrusions. We show that this multi-scale model allows us to relate the structural as well as thermodynamic details of exo70 membrane-binding and self-association to the induced deformation field and resulting cellular morphologies. We also extend our studies to ENTH, BAR, and ESCRT family proteins. We show that thermal undulations in the membrane and cooperativity in the curvature fields, collectively drive the membrane into different morphological states (buds, tubules, etc.) that resemble those in cellular experiments in vivo and vesicle experiments in vitro. We determine the relative stability of the above mentioned shapes based on the free energy of these membrane configurations, determined using the methods of Thermodynamic Integration (TI) and Bennett Acceptance (BA), and Widom Insertion techniques. Results are shown for the case of Exo70 protein and ENTH, and N-BAR domains and also compared against measurements determined from experiments.

Membrane Remodelling and Protein Interactions - A Free Energy Perspective

We investigate the interplay between cell membrane curvature induced at the atomic scale, due to specialized peripheral membrane proteins, and the resulting membrane morphologies, of varying complexity, observed at the mesoscale. The biological membrane, in our approach, is represented by a dynamically triangulated surface while the proteins are modeled as curvature fields on the membrane, that are either isotropic or anisotropic. In order to compare with experiments, we have focused on the ENTH domain containing EPSIN whose curvature field is modeled as isotropic, and on the BAR, Exo70 and ESCRT family proteins whose curvature fields are determined to be anisotropic, both in experiments and in molecular simulations. Thermal undulations in the membrane and cooperativity in the curvature fields, due to the stabilization of a nematic phase, collectively drive the membrane into different morphological states (buds, tubules, etc.) that resemble those in cellular experiments in vivo and vesicle experiments in vitro. The relative stabilities of these self-organized shapes are examined by two approaches to compute the free energy of the system: (i) the Widom insertion technique to compute excess chemical potentials and (ii) thermodynamic integration using the Kirkwood coupling parameter to compute free energies. Building on these methods, we propose a hybrid scheme that couples both the approaches for computing free energies in membrane systems with heterogeneous and phase-segregated protein field - examples being the endoplasmic reticulum (ER) membrane discs with α- calreticulon protein confined to the rim and the vesicular bud formed due to the constriction by ESCRT proteins at its neck.

The role for the exocyst complex subunits Exo70B2 and Exo70H1 in the plant-pathogen interaction.

Recently, the octameric vesicle-tethering complex exocyst was found in plants and its importance for Arabidopsis morphogenesis was demonstrated. Exo70 exocyst subunits in plants, unlike in yeasts and mammals, are represented by a multigene family, comprising 23 members in Arabidopsis. For Exo70B2 and Exo70H1 paralogues, transcriptional up-regulation was confirmed on treatment with an elicitor peptide, elf18, derived from the bacterial elongation factor. Their ability to participate in the exocyst complex formation was inferred by the interaction of both the Exo70s with several other exocyst subunits using the yeast two-hybrid system. Arabidopsis plants mutated in these two genes were used to analyse their local reaction upon inoculation with Pseudomonas syringae pv. maculicola and the fungal pathogen Blumeria graminis f. sp. hordei. The Pseudomonas sensitivity test revealed enhanced susceptibility for the two exo70B2 and one H1 mutant lines. After Blumeria inoculation, an increase in the proportion of abnormal papilla formation, with an unusual wide halo made of vesicle-like structures, was found in exo70B2 mutants. Intracellular localization of both Exo70 proteins was studied following a GFP fusion assay and Agrobacterium-mediated transient expression of the constructs in Nicotiana benthamiana leaf epidermis. GFP-Exo70H1 localizes in the vesicle-like structures, while GFP-Exo70B2 is localized mainly in the cytoplasm. It is concluded that both Exo70B2 and Exo70H1 are involved in the response to pathogens, with Exo70B2 having a more important role in cell wall apposition formation related to plant defence.

EXO70 protein influences dengue virus secretion

The involvement of host proteins in assisting the exocytosis of flaviviruses is largely unknown. In this study, we aimed to investigate if dengue virus (DENV) utilizes the exocyst components to aid the exocytosis of virus particles. This study identified that EXO70 protein, a member of the exocyst complex influenced DENV infection. Dengue virus production was significantly attenuated in EXO70 knock-down cells. EXO70 did not influence viral transcription and translation. It influenced virus egression/secretion from DENV-infected cells. We also showed that EXO70 expression was up-regulated from 18 h post-infection following DENV infection. Although the envelope protein of DENV influenced EXO70 expression, the co-expression of pre-membrane and envelope proteins significantly increased the expression levels of EXO70 during DENV infection. When pre-membrane protein was expressed alone, there was no significant difference in the expression levels of EXO70. This indicated that the presence of pre-membrane protein might help in the proper folding of envelope protein. Increased expression levels of EXO70 might help in the exocytosis process of virus or subviral particles.