scholarly journals Pepper CaMLO6 Negatively Regulates Ralstonia solanacearum Resistance and Positively Regulates High Temperature and High Humidity Responses

2020 ◽  
Vol 61 (7) ◽  
pp. 1223-1238
Author(s):  
Sheng Yang ◽  
Yuanyuan Shi ◽  
Longyun Zou ◽  
Jinfeng Huang ◽  
Lei Shen ◽  
...  
Keyword(s):  
High Temperature ◽  
Ralstonia Solanacearum ◽  
Negative Regulator ◽  
Marker Genes ◽  
Plant Responses ◽  
Resistance Locus ◽  
High Humidity ◽  
Virus Induced Gene Silencing ◽  
Transcriptional Responses ◽  
Pepper Plants

Abstract Plant mildew-resistance locus O (MLO) proteins influence susceptibility to powdery mildew. However, their roles in plant responses to other pathogens and heat stress remain unclear. Here, we showed that CaMLO6, a pepper (Capsicum annuum) member of MLO clade V, is a protein targeted to plasma membrane and probably endoplasmic reticulum. The transcript expression level of CaMLO6 was upregulated in the roots and leaves of pepper plants challenged with high temperature and high humidity (HTHH) and was upregulated in leaves but downregulated in roots of plants infected with the bacterial pathogen Ralstonia solanacearum. CaMLO6 was also directly upregulated by CaWRKY40 upon HTHH but downregulated by CaWRKY40 upon R. solanacearum infection. Virus-induced gene silencing of CaMLO6 significantly decreased pepper HTHH tolerance and R. solanacearum susceptibility. Moreover, CaMLO6 overexpression enhanced the susceptibility of Nicotiana benthamiana and pepper plants to R. solanacearum and their tolerance to HTHH, effects that were associated with the expression of immunity- and thermotolerance-associated marker genes, respectively. These results suggest that CaMLO6 acts as a positive regulator in response to HTHH but a negative regulator in response to R. solanacearum. Moreover, CaMLO6 is transcriptionally affected by R. solanacearum and HTHH; these transcriptional responses are at least partially regulated by CaWRKY40.

scholarly journals CaSK23, a Putative GSK3/SHAGGY-Like Kinase of Capsicum annuum, Acts as a Negative Regulator of Pepper’s Response to Ralstonia solanacearum Attack

2018 ◽  
Vol 19 (9) ◽  
pp. 2698 ◽  
Author(s):  
Ailian Qiu ◽  
Ji Wu ◽  
Yufen Lei ◽  
Yiting Cai ◽  
Song Wang ◽  
...  
Keyword(s):  
Ralstonia Solanacearum ◽  
Plant Immunity ◽  
Constitutive Expression ◽  
Negative Regulator ◽  
Virus Induced Gene Silencing ◽  
Pr Genes ◽  
Down Regulation ◽  
Genes Encoding ◽  
Transient Overexpression ◽  
Pepper Plants

GSK3-like kinases have been mainly implicated in the brassinosteroids (BR) pathway and, therefore, in plant growth, development, and responses to abiotic stresses; however, their roles in plant immunity remain poorly understood. Herein, we present evidence that CaSK23, a putative GSK3/SHAGGY-like kinase in pepper, acts as a negative regulator in pepper’s response to Ralstonia solanacearum (R. solanacearum) inoculation (RSI). Data from quantitative RT-PCR (qRT-PCR) showed that the constitutively-expressed CaSK23 in pepper leaves was down-regulated by RSI, as well as by exogenously-applied salicylic acid (SA) or methyl jasomonate (MeJA). Silencing of CaSK23 by virus-induced gene silencing (VIGS) decreased the susceptibility of pepper plants to RSI, coupled with up-regulation of the tested genes encoding SA-, JA-, and ethylene (ET)-dependent pathogenesis-related (PR) proteins. In contrast, ectopic overexpression (OE) of CaSK23 conferred a compromised resistance of tobacco plants to RSI, accompanied by down-regulation of the tested immunity-associated SA-, JA-, and ET-dependent PR genes. In addition, transient overexpression of CaSK23 in pepper plants consistently led to down-regulation of the tested SA-, JA-, and ET-dependent PR genes. We speculate that CaSK23 acts as a negative regulator in pepper immunity and its constitutive expression represses pepper immunity in the absence of pathogens. On the other hand, its decreased expression derepresses immunity when pepper plants are attacked by pathogens.

scholarly journals The Pepper Mitogen-Activated Protein Kinase CaMAPK7 Acts as a Positive Regulator in Response to Ralstonia solanacearum Infection

2021 ◽  
Vol 12 ◽  
Author(s):  
Lanping Shi ◽  
Kan Zhang ◽  
Linjing Xie ◽  
Mingxing Yang ◽  
Baixue Xie ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Ralstonia Solanacearum ◽  
Target Genes ◽  
Vital Role ◽  
Mitogen Activated Protein Kinase ◽  
Marker Genes ◽  
Virus Induced Gene Silencing ◽  
Mitogen Activated Protein ◽  
Pepper Genome ◽  
Pepper Plants

Mitogen-activated protein kinase (MAPK) pathways play a vital role in multiple plant processes, including growth, development, and stress signaling, but their involvement in response to Ralstonia solanacearum is poorly understood, particularly in pepper plants. Herein, CaMAPK7 was identified from the pepper genome and functionally analyzed. The accumulations of CaMAPK7 transcripts and promoter activities were both significantly induced in response to R. solanacearum strain FJC100301 infection, and exogenously applied phytohormones, including methyl jasmonate (MeJA), brassinolide (BR), salicylic acid (SA), and ethephon (ETN), were decreased by abscisic acid (ABA) treatment. Virus-induced gene silencing (VIGS) of CaMAPK7 significantly enhanced the susceptibility of pepper plants to infection by R. solanacearum and downregulated the defense-related marker genes, including CaDEF1, CaPO2, CaSAR82A, and CaWRKY40. In contrast, the ectopic overexpression of CaMAPK7 in transgenic tobacco enhanced resistance to R. solanacearum and upregulated the defense-associated marker genes, including NtHSR201, NtHSR203, NtPR4, PR1a/c, NtPR1b, NtCAT1, and NtACC. Furthermore, transient overexpression of CaMAPK7 in pepper leaves triggered intensive hypersensitive response (HR)-like cell death, H2O2 accumulation, and enriched CaWRKY40 at the promoters of its target genes and drove their transcript accumulations, including CaDEF1, CaPO2, and CaSAR82A. Taken together, these data indicate that R. solanacearum infection induced the expression of CaMAPK7, which indirectly modifies the binding of CaWRKY40 to its downstream targets, including CaDEF1, CaPO2, and CaSAR82A, ultimately leading to the activation of pepper immunity against R. solanacearum. The protein that responds to CaMAPK7 in pepper plants should be isolated in the future to build a signaling bridge between CaMAPK7 and CaWRKY40.

scholarly journals CaHDZ27, a Homeodomain-Leucine Zipper I Protein, Positively Regulates the Resistance to Ralstonia solanacearum Infection in Pepper

2017 ◽  
Vol 30 (12) ◽  
pp. 960-973 ◽  
Author(s):  
Shaoliang Mou ◽  
Zhiqin Liu ◽  
Feng Gao ◽  
Sheng Yang ◽  
Meixia Su ◽  
...  
Keyword(s):  
Ralstonia Solanacearum ◽  
Leucine Zipper ◽  
Fluorescent Protein ◽  
Plant Immunity ◽  
Bimolecular Fluorescence Complementation ◽  
Marker Genes ◽  
Plant Responses ◽  
Dependent Manner ◽  
Virus Induced Gene Silencing ◽  
Green Fluorescent

Homeodomain-leucine zipper class I (HD-Zip I) transcription factors have been functionally characterized in plant responses to abiotic stresses, but their roles in plant immunity are poorly understood. Here, a HD-Zip I gene, CaHZ27, was isolated from pepper (Capsicum annum) and characterized for its role in pepper immunity. Quantitative real-time polymerase chain reaction showed that CaHDZ27 was transcriptionally induced by Ralstonia solanacearum inoculation and exogenous application of methyl jasmonate, salicylic acid, or ethephon. The CaHDZ27-green fluorescent protein fused protein was targeted exclusively to the nucleus. Chromatin immunoprecipitation demonstrated that CaHDZ27 bound to the 9-bp pseudopalindromic element (CAATAATTG) and triggered β-glucuronidase expression in a CAATAATTG-dependent manner. Virus-induced gene silencing of CaHDZ27 significantly attenuated the resistance of pepper plants against R. solanacearum and downregulated defense-related marker genes, including CaHIR1, CaACO1, CaPR1, CaPR4, CaPO2, and CaBPR1. By contrast, transient overexpression of CaHDZ27 triggered strong cell death mediated by the hypersensitive response and upregulated the tested immunity-associated marker genes. Ectopic CaHDZ27 expression in tobacco enhances its resistance against R. solanacearum. These results collectively suggest that CaHDZ27 functions as a positive regulator in pepper resistance against R. solanacearum. Bimolecular fluorescence complementation and coimmunoprecipitation assays indicate that CaHDZ27 monomers bind with each other, and this binding is enhanced significantly by R. solanacearum inoculation. We speculate that homodimerization of CaHZ27 might play a role in pepper response to R. solanacearum, further direct evidence is required to confirm it.

scholarly journals Ca14-3-3 Interacts With CaWRKY58 to Positively Modulate Pepper Response to Low-Phosphorus Starvation

2021 ◽  
Vol 11 ◽  
Author(s):  
Jinsen Cai ◽  
Weiwei Cai ◽  
Xueying Huang ◽  
Sheng Yang ◽  
Jiayu Wen ◽  
...  
Keyword(s):  
Ralstonia Solanacearum ◽  
Phosphorus Deficiency ◽  
Negative Regulator ◽  
Bimolecular Fluorescence Complementation ◽  
Virus Induced Gene Silencing ◽  
Mobility Shift ◽  
Natural Habitats ◽  
Low Phosphorus ◽  
Low Phosphorus Stress ◽  
Electrophoretic Mobility Shift Assays

Low-phosphorus stress (LPS) and pathogen attack are two important stresses frequently experienced by plants in their natural habitats, but how plant respond to them coordinately remains under-investigated. Here, we demonstrate that CaWRKY58, a known negative regulator of the pepper (Capsicum annuum) response to attack by Ralstonia solanacearum, is upregulated by LPS. Virus-induced gene silencing (VIGS) and overexpression of CaWRKY58 in Nicotiana benthamiana plants in combination with chromatin immunoprecipitation (ChIP) and electrophoretic mobility shift assays (EMSA) demonstrated that CaWRKY58 positively regulates the response of pepper to LPS by directly targeting and regulating genes related to phosphorus-deficiency tolerance, including PHOSPHATE STARVATION RESPONSE1 (PHR1). Yeast two-hybrid assays revealed that CaWRKY58 interacts with a 14-3-3 protein (Ca14-3-3); this interaction was confirmed by pull-down, bimolecular fluorescence complementation (BiFC), and microscale thermophoresis (MST) assays. The interaction between Ca14-3-3 and CaWRKY58 enhanced the activation of PHR1 expression by CaWRKY58, but did not affect the expression of the immunity-related genes CaNPR1 and CaDEF1, which are negatively regulated by CaWRKY58 in pepper upon Ralstonia solanacearum inoculation. Collectively, our data indicate that CaWRKY58 negatively regulates immunity against Ralstonia solanacearum, but positively regulates tolerance to LPS and that Ca14-3-3 transcriptionally activates CaWRKY58 in response to LPS.

scholarly journals The Grapevine Calmodulin-Like Protein Gene CML21 Is Regulated by Alternative Splicing and Involved in Abiotic Stress Response

2020 ◽  
Vol 21 (21) ◽  
pp. 7939
Author(s):  
Olga A. Aleynova ◽  
Konstantin V. Kiselev ◽  
Zlata V. Ogneva ◽  
Alexandra S. Dubrovina
Keyword(s):  
Abiotic Stress ◽  
Cold Stress ◽  
Splice Variants ◽  
Constitutive Expression ◽  
Freezing Stress ◽  
Marker Genes ◽  
Vitis Amurensis ◽  
Plant Responses ◽  
Abiotic Stress Response ◽  
Transcriptional Responses

Calmodulin-like proteins (CMLs) represent a large family of plant calcium sensor proteins involved in the regulation of plant responses to environmental cues and developmental processes. In the present work, we identified four alternatively spliced mRNA forms of the grapevine CML21 gene that encoded proteins with distinct N-terminal regions. We studied the transcript abundance of CML21v1, CML21v2, CML21v3, and CML21v4 in wild-growing grapevine Vitis amurensis Rupr. in response to desiccation, heat, cold, high salinity, and high mannitol stress using quantitative real-time RT-PCR. The levels of all four splice variants of VaCML21 were highly induced in response to cold stress. In addition, VaCML21v1 and VaCML21v2 forms were highly modulated by all other abiotic stress treatments. Constitutive expression of VaCML21v2 and VaCML21v4 improved biomass accumulation of V. amurensis callus cell cultures under prolonged low temperature stress. Heterologous expression of the grapevine CML21v2 and VaCML21v4 splice variants in Arabidopsis improved survival rates of the transgenic plants after freezing. The VaCML21v2 overexpression enhanced activation of the cold stress-responsive marker genes AtDREB1A and AtDREB2A, while VaCML21v4 overexpression—AtCOR47, AtRD29A, AtRD29B, and AtKIN1 genes after freezing stress in the transgenic Arabidopsis. The results indicate that the grapevine CML21 gene acts as a positive regulator in the plant response to cold stress. The detected variety of CML21 transcripts and their distinct transcriptional responses suggested that this expansion of mRNA variants could contribute to the diversity of grapevine adaptive reactions.

scholarly journals CaCBL1 Acts as a Positive Regulator in Pepper Response to Ralstonia solanacearum

2020 ◽  
Vol 33 (7) ◽  
pp. 945-957 ◽  
Author(s):  
Lei Shen ◽  
Sheng Yang ◽  
Feng Yang ◽  
Deyi Guan ◽  
Shuilin He
Keyword(s):  
Bacterial Wilt ◽  
Ralstonia Solanacearum ◽  
Marker Genes ◽  
Virus Induced Gene Silencing ◽  
Mobility Shift ◽  
Positive Regulator ◽  
Positive Regulators ◽  
Mobility Shift Assay ◽  
Transient Overexpression ◽  
Important Disease

Bacterial wilt caused by Ralstonia solanacearum is an important disease of pepper (Capsicum annuum), an economically important solanaceous vegetable worldwide, in particular, under high temperature (HT) conditions. However, the molecular mechanism underlying pepper immunity against bacterial wilt remains poorly understood. Herein, CaCBL1, a putative calcineurin B-like protein, was functionally characterized in the pepper response to R. solanacearum inoculation (RSI) under HT (RSI/HT). CaCBL1 was significantly upregulated by RSI at room temperature (RSI/RT), HT, or RSI/HT. CaCBL1-GFP fused protein targeted to whole epidermal cells of Nicotiana benthamiana when transiently overexpressed. CaCBL1 silencing by virus-induced gene silencing significantly enhanced pepper susceptibility to RSI under RT or HT, while its transient overexpression triggered hypersensitive response mimic cell death and upregulation of immunity-associated marker genes, including CabZIP63, CaWRKY40, and CaCDPK15, the positive regulators in the pepper response to RSI or HT found in our previous studies. In addition, by chromatin immunoprecipitation PCR and electrophoretic mobility shift assay, CaCBL1 was found to be directly targeted by CaWRKY40, although not by CaWRKY27 or CaWRKY58, via the W-box-2 within its promoter, and its transcription was found to be downregulated by silencing of CaWRKY40 while it was enhanced by its transient overexpression. These results suggest that CaCBL1 acts as a positive regulator in pepper immunity against R. solanacearum infection, constituting a positive feedback loop with CaWRKY40.

scholarly journals TaDAD2, a Negative Regulator of Programmed Cell Death, Is Important for the Interaction Between Wheat and the Stripe Rust Fungus

2011 ◽  
Vol 24 (1) ◽  
pp. 79-90 ◽  
Author(s):  
Xiaojie Wang ◽  
Chunlei Tang ◽  
Hongchang Zhang ◽  
Jin-Rong Xu ◽  
Bo Liu ◽  
...  
Keyword(s):  
Cell Death ◽  
Stripe Rust ◽  
Rust Fungus ◽  
Control Level ◽  
Negative Regulator ◽  
Predicted Amino Acid Sequence ◽  
Initial Contact ◽  
Virus Induced Gene Silencing ◽  
Transcriptional Responses ◽  
Stripe Rust Fungus

Defender against cell death (DAD) genes are known to function as negative regulators of cell death in animals. In plants, DAD orthologs are conserved but their role in cell death regulation is not well understood. Here, we report the characterization of the TaDAD2 gene in wheat. The predicted amino acid sequence of TaDAD2 contains typical structural features of DAD proteins, including a signal peptide, three transmembrane regions, and a subunit of oligosaccharyltransferase. Transcripts of TaDAD2 were detected in wheat leaves, culms, roots, florets, and spikelets. The expression level of TaDAD2 was reduced in the initial contact with the stripe rust fungus, subsequently induced and peaked at 18 h postinoculation (hpi), gradually reduced at 24 to 48 hpi, and restored to control level at 72 to 120 hpi. In addition, TaDAD2 exhibited positive transcriptional responses to abiotic stresses after the initial reduction at 1 hpi. Overexpression of TaDAD2 in tobacco leaves inhibited cell death. Furthermore, knocking down TaDAD2 expression by virus-induced gene silencing enhanced the susceptibility of wheat cv. Suwon11 to avirulent race CYR23 and reduced necrotic area at the infection sites. These results indicate that TaDAD2 may function as a suppressor of cell death in the early stages of wheat–stripe rust fungus interaction. However, it is dispensable for or plays an opposite role in hypersensitive response or cell death triggered by an avirulent race of stripe rust fungus at late-infection stages.

scholarly journals CaWRKY30 Positively Regulates Pepper Immunity by Targeting CaWRKY40 against Ralstonia solanacearum Inoculation through Modulating Defense-Related Genes

2021 ◽  
Vol 22 (21) ◽  
pp. 12091
Author(s):  
Ansar Hussain ◽  
Muhammad Ifnan Khan ◽  
Mohammed Albaqami ◽  
Shahzadi Mahpara ◽  
Ijaz Rasool Noorka ◽  
...  
Keyword(s):  
Cell Death ◽  
Ralstonia Solanacearum ◽  
Foliar Application ◽  
Critical Role ◽  
Underlying Mechanism ◽  
Marker Genes ◽  
Transcriptional Expression ◽  
Virus Induced Gene Silencing ◽  
Expression Levels ◽  
Over Expression

The WRKY transcription factors (TFs) network is composed of WRKY TFs’ subset, which performs a critical role in immunity regulation of plants. However, functions of WRKY TFs’ network remain unclear, particularly in non-model plants such as pepper (Capsicum annuum L.). This study functionally characterized CaWRKY30—a member of group III Pepper WRKY protein—for immunity of pepper against Ralstonia solanacearum infection. The CaWRKY30 was detected in nucleus, and its transcriptional expression levels were significantly upregulated by R. solanacearum inoculation (RSI), and foliar application ethylene (ET), abscisic acid (ABA), and salicylic acid (SA). Virus induced gene silencing (VIGS) of CaWRKY30 amplified pepper’s vulnerability to RSI. Additionally, the silencing of CaWRKY30 by VIGS compromised HR-like cell death triggered by RSI and downregulated defense-associated marker genes, like CaPR1, CaNPR1, CaDEF1, CaABR1, CaHIR1, and CaWRKY40. Conversely, transient over-expression of CaWRKY30 in pepper leaves instigated HR-like cell death and upregulated defense-related maker genes. Furthermore, transient over-expression of CaWRKY30 upregulated transcriptional levels of CaWRKY6, CaWRKY22, CaWRKY27, and CaWRKY40. On the other hand, transient over-expression of CaWRKY6, CaWRKY22, CaWRKY27, and CaWRKY40 upregulated transcriptional expression levels of CaWRKY30. The results recommend that newly characterized CaWRKY30 positively regulates pepper’s immunity against Ralstonia attack, which is governed by synergistically mediated signaling by phytohormones like ET, ABA, and SA, and transcriptionally assimilating into WRKY TFs networks, consisting of CaWRKY6, CaWRKY22, CaWRKY27, and CaWRKY40. Collectively, our data will facilitate to explicate the underlying mechanism of crosstalk between pepper’s immunity and response to RSI.

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