Biology subjects

Rikkerink, E. H. A.

Publications and source records attributed to Rikkerink, E. H. A..

5 recordsLinked to original sources

bioRxiv2025

Systematic Analysis of the EXO70 Gene Family in Kiwifruit Species: Evolutionary Selection and Potential Functions in Plant Immunity

BackgroundKiwifruit (Actinidia spp.) is a commercially and nutritionally valuable fruit crop that faces increasing challenges from pathogens, particularly Pseudomonas syringae pv. actinidiae (Psa). The exocyst complex, especially the EXO70 subunit, has been demonstrated to play a crucial role in vesicle trafficking and plant immune responses in model species such as Arabidopsis thaliana. However, the function and evolution of EXO70 genes in fruit crops remain largely unexplored. ResultsWe conducted a comprehensive genome-wide analysis of the EXO70 gene family across five Actinidia species using Arabidopsis EXO70 sequences as queries. A total of 217 EXO70 genes (23 to 54 paralogues per species) were identified and classified into three subfamilies and nine clades (EXO70A-EXO70I), consistent with previous classifications in other plant taxa. Phylogenetic reconstruction and microsynteny analyzes revealed lineage- and genus-specific expansions of EXO70C members, as well as species-specific expansion events, particularly within the EXO70E and EXO70H clades. To investigate potential immune functions, yeast two-hybrid and in planta co-immunoprecipitation assays confirmed that kiwifruit EXO70B1 physically interacts with the immune hub protein kiwifruit RIN4_1. These interactions support conservation of the EXO70-RIN4 module in plant immunity. ConclusionsThis study provides the first comprehensive characterization of the EXO70 gene family across multiple kiwifruit species and uncovers candidate genes potentially involved in plant immunity through RIN4-mediated defense signaling. Notably, the expansion and diversification of the EXO70E and EXO70H clades suggest possible adaptive evolution within these clades and neofunctionalization. These findings provide valuable genomic resources and novel insights into the evolution of vesicle trafficking components in fruit crops, laying the groundwork for future efforts to enhance disease resistance in kiwifruit through breeding or biotechnological approaches.

plant biology↗

bioRxiv2021

A conserved glutamate residue in RPM1-interacting protein4 is ADP-ribosylated by Pseudomonas effector AvrRpm2 to activate RPM1-mediated response

Gram-negative bacterial plant pathogens directly inject effectors into their hosts to hijack and manipulate metabolism, eluding the frontier surveillance at the cell surface. The effector AvrRpm1Pma from Pseudomonas syringae pv. maculicola functions as an ADP-ribosyl transferase, modifying RPM1-interacting protein4 (RIN4), leading to the activation of Arabidopsis resistance protein RPM1. We identified the ADP-ribosyl transferase activity of another bacterial effector AvrRpm2Psa from Pseudomonas syringae pv. actinidiae via infection using a Pseudomonas syringae pv. tomato strain following Agrobacterium-mediated transient expression of RIN4 in N. benthamiana. We conducted mutational analysis in combination with mass spectrometry to genetically locate the modified residue. We show that a conserved glutamate residue (E156) of AtRIN4 is the target site for AvrRpm2Psa by demonstrating the modified AtRIN4 with E156A substitution is no longer ADP-ribosylated. Accordingly, naturally occurring soybean and snap bean RIN4 homologs with no glutamate at the positions corresponding to the E156 of AtRIN4 are not ADP-ribosylated by AvrRpm2Psa. In contrast with another effector AvrB, modifications of potential phosphorylation sites including T166 in AtRIN4 affected neither ADP-ribosylation nor RPM1 activation by AvrRpm2Psa. This study suggests that separate biochemical reactions by different pathogen effectors may trigger the activation of the same resistance protein through distinct modifications of RIN4. One sentence summaryA conserved glutamate residue (E156) in the C-NOI domain of RPM1-interacting protein4 is ADP-ribosylated by Pseudomonas effector AvrRpm2 to activate RPM1-mediated defence response, independently of phosphorylation at T166.

plant biology↗

bioRxiv2020

GAI MoRFs Regulate Cleft and Channel Binding Pathways for Gibberellin in GID1A

AbstractThe hormone gibberellin (GA) promotes arabidopsis growth by enhancing binding between GA Insensitive DELLA transcriptional repressors and GA Insensitive Dwarf 1 (GID1) receptors to regulate DELLA degradation. The binding mechanism for GA was elucidated by employing a computational study of dissociations of the N-terminus of the DELLA family member GAI (GA Insensitive transcriptional repressor) from the GID1A receptor in the presence and absence of bound GA, and of GA from GID1A in the presence and absence of GAI. The tRAMD method was employed to deduce egression pathways for a diverse set of GA molecules (GA(x)). Two pathways in the form of a newly identified cleft and a previously identified channel are prevalent. The cleft pathway is open in the absence of GAI. Upon GAI binding, the cleft route is blocked, resulting in a slower process for GA(x) to exit and enter the binding pocket through the channel. Several binding pocket residues are identified as gate-keepers to the channel. Molecular recognition features (MoRFs) found in the disordered signaling protein GAI affect GA(x) binding and GID1A dynamics. A three-step synergistic binding cycle is proposed where GAI MoRFs regulate the process. Rapid binding takes place through the cleft where little to no distinctions are made between major and less active forms of GA(x). After GAI is bound to the GA(x) {middle dot} GID1A complex, the channel supports a rectification process that increases the retention of major active forms of GA within the binding pocket. Both the cleft and channel contact residues to GA(x) are markedly conserved in a GID1 phylogeny, suggesting this binding process in the GID1 {middle dot} DELLA GA-receptor complex represents a general paradigm for GA binding. Non-specific GA binding assists binding of GAI, which then helps to select the major active forms of the hormone and induce a downstream signalling cascade in response to bioactive GA. Non-expert Summary StatementGibberellins are plant hormones essential for growth and development. The DELLA proteins are a disordered family of repressors that transcriptionally repress GA responsive genes. Degradation of DELLA proteins in response to GA results in GA-responsive genes being upregulated. Binding of GA to the GA-Insensitive Dwarf 1 receptor (GID1) facilitates binding of DELLA to the GA {middle dot} GID1 complex. Through computational modelling and phylogenetic analyses, we identified a new GA binding cleft that is blocked by DELLA binding and a three-step mechanism for the GA {middle dot} DELLA {middle dot} GID1 complex that also involves the known GA binding channel. We propose a dual (cleft/channel) pathway that allows access to the binding pocket as a paradigm for selection of specific GA forms among a mixture of major active and inactive forms. The cleft is less selective, but preference for active GA in the binding pocket of GID1A is amplified by expunging inactive GA forms, followed by recruiting active forms through the more selective channel. This mechanism allows plants to sense concentration changes of GA with high specificity to enable certain GA variants to trigger specific signalling events. These novel insights into the receptor mechanism in part may explain the large number of different GA forms that exist in nature. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/422840v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@a7b6e5org.highwire.dtl.DTLVardef@d89799org.highwire.dtl.DTLVardef@1c8ee74org.highwire.dtl.DTLVardef@9f31c6_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2018

Re-programming of Pseudomonas syringae pv. actinidiae gene expression during early stages of infection of kiwifruit

BackgroundPseudomonas syringae is a widespread bacterial species complex that includes a number of significant plant pathogens. Amongst these, P. syringae pv. actinidiae (Psa) initiated a worldwide pandemic in 2008 on cultivars of Actinidia chinensis var. chinensis. To gain information about the expression of genes involved in pathogenicity we have carried out transcriptome analysis of Psa during the early stages of kiwifruit infection.\n\nResultsGene expression in Psa was investigated during the first five days after infection of kiwifruit plantlets, using RNA-seq. Principal component and heatmap analyses showed distinct phases of gene expression during the time course of infection. The first phase was an immediate transient peak of induction around three hours post inoculation (HPI) that included genes that code for a Type VI Secretion System and nutrient acquisition (particularly phosphate). This was followed by a significant commitment, between 3 and 24 HPI, to the induction of genes encoding the Type III Secretion System (T3SS) and Type III Secreted Effectors (T3SE). Expression of these genes collectively accounted for 6.3% of the bacterial transcriptome at this stage. There was considerable variation in the expression levels of individual T3SEs but all followed the same temporal expression pattern, with the exception of HopAS1, which peaked later in expression at 48 HPI. As infection progressed over the time course of five days, there was an increase in the expression of genes with roles in sugar, amino acid and sulfur transport and the production of alginate and colanic acid. These are both polymers that are major constituents of extracellular polysaccharide substances (EPS) and are involved in biofilm production. Reverse transcription-quantitative PCR (RT-qPCR) on an independent infection time course experiment showed that the expression profile of selected bacterial genes at each infection phase correlated well with the RNA-seq data.\n\nConclusionsThe results from this study indicate that there is a complex remodeling of the transcriptome during the early stages of infection, with at least three distinct phases of coordinated gene expression. These include genes induced during the immediate contact with the host, those involved in the initiation of infection, and finally those responsible for nutrient acquisition.

microbiology↗

bioRxiv2018

NRG1-mediated recognition of HopQ1 reveals a link between PAMP and Effector-triggered Immunity.

O_LIMembers of the hopQ1/XopQ family of effectors are conserved amongst pathogenic bacteria suggesting an important function in virulence. Therefore, the identification of R proteins recognising members of this family is potentially of high value to achieve broad-spectrum resistance in crops.\nC_LIO_LIRecent studies demonstrated that HopQ1/XopQ recognition is EDS1-dependant and is mediated by the TIR-NB-LRR protein Roq1. Using our recently described R gene RNAi library, we have investigated the mechanism of HopQ1 recognition and the other components of its signalling network.\nC_LIO_LIWe show that silencing of the CCR-NB-LRR helper protein N Requirement Gene 1 (NRG1) prevents HopQ1 recognition in N. benthamiana and that NRG1 expression confers recognition of HopQ1 and restricts Pto DC3000 proliferation in the otherwise susceptible host Arabidopsis. Moreover NRG1 expression is briefly but strongly induced at a very early stage of the plant innate response, establishing a strong dependency link between PAMP-triggered immunity (PTI) and effector-triggered immunity (ETI).\nC_LIO_LIOverall we demonstrate that in addition to Roq1, HopQ1 recognition in N. benthamiana requires the CCR-NB-LRR protein NRG1 and propose a model based on the dual requirement of a CCR-NB-LRR and a TIR-NB-LRR that could extend beyond HopQ1 detection and possibly be used to recognize other pathogen effectors.\nC_LI

plant biology↗