Transcription factor CaHDZ15 promotes pepper basal thermotolerance by activating HEAT SHOCK FACTORA6a.

High temperature stress (HTS) is a serious threat to plant growth and development and to crop production in the context of global warming, and plant response to HTS is largely regulated at the transcriptional level by the actions of various transcription factors (TFs). However, whether and how homeodomain-leucine zipper (HD-Zip) TFs are involved in thermotolerance are unclear. Herein, we functionally characterized a pepper (Capsicum annuum) HD-Zip I TF CaHDZ15. CaHDZ15 expression was upregulated by HTS and abscisic acid in basal thermotolerance via loss- and gain-of-function assays by virus-induced gene silencing in pepper and overexpression in Nicotiana benthamiana plants. CaHDZ15 acted positively in pepper basal thermotolerance by directly targeting and activating HEAT SHOCK FACTORA6a (HSFA6a), which further activated CaHSFA2. In addition, CaHDZ15 interacted with HEAT SHOCK PROTEIN 70-2 (CaHsp70-2) and glyceraldehyde-3-phosphate dehydrogenase1 (CaGAPC1), both of which positively affected pepper thermotolerance. CaHsp70-2 and CaGAPC1 promoted CaHDZ15 binding to the promoter of CaHSFA6a, thus enhancing its transcription. Furthermore, CaHDZ15 and CaGAPC1 were protected from 26S proteasome-mediated degradation by CaHsp70-2 via physical interaction. These results collectively indicate that CaHDZ15, modulated by the interacting partners CaGAPC1 and CaHsp70-2, promotes basal thermotolerance by directly activating the transcript of CaHSFA6a. Thus, a molecular linkage is established among CaHsp70-2, CaGAPC1, and CaHDZ15 to transcriptionally modulate CaHSFA6a in pepper thermotolerance.

Differential CaKAN3-CaHSF8 associations underlie distinct immune and heat responses under high temperature and high humidity conditions.

High temperature and high humidity (HTHH) conditions increase plant susceptibility to a variety of diseases, including bacterial wilt in solanaceous plants. Some solanaceous plant cultivars have evolved mechanisms to activate HTHH-specific immunity to cope with bacterial wilt disease. However, the underlying mechanisms remain poorly understood. Here we find that CaKAN3 and CaHSF8 upregulate and physically interact with each other in nuclei under HTHH conditions without inoculation or early after inoculation with R. solanacearum in pepper. Consequently, CaKAN3 and CaHSF8 synergistically confer immunity against R. solanacearum via activating a subset of NLRs which initiates immune signaling upon perception of unidentified pathogen effectors. Intriguingly, when HTHH conditions are prolonged without pathogen attack or the temperature goes higher, CaHSF8 no longer interacts with CaKAN3. Instead, it directly upregulates a subset of HSP genes thus activating thermotolerance. Our findings highlight mechanisms controlling context-specific activation of high-temperature-specific pepper immunity and thermotolerance mediated by differential CaKAN3-CaHSF8 associations.

A cysteine-rich receptor-like protein kinase CaCKR5 modulates immune response against Ralstonia solanacearum infection in pepper.

BACKGROUND: Cysteine-rich receptor-like kinases (CRKs) represent a large subfamily of receptor-like kinases and play vital roles in diverse physiological processes in regulating plant growth and development. RESULTS: CaCRK5 transcripts were induced in pepper upon the infection of Ralstonia solanacearum and treatment with salicylic acid. The fusions between CaCRK5 and green fluorescence protein were targeted to the plasma membrane. Suppression of CaCRK5 via virus-induced gene silencing (VIGS) made pepper plants significantly susceptible to R. solanacearum infection, which was accompanied with decreased expression of defense related genes CaPR1, CaSAR8.2, CaDEF1 and CaACO1. Overexpression of CaCRK5 increased resistance against R. solanacearum in Nicotiana benthamiana. Furthermore, electrophoretic mobility shift assay and chromatin immunoprecipitation coupled with quantitative real-time PCR analysis revealed that a homeodomain zipper I protein CaHDZ27 can active the expression of CaCRK5 through directly binding to its promoter. Yeast two-hybrid and bimolecular fluorescence complementation (BiFC) analyses suggested that CaCRK5 heterodimerized with the homologous member CaCRK6 on the plasma membrane. CONCLUSIONS: Our data revealed that CaCRK5 played a positive role in regulating immune responses against R. solanacearum infection in pepper.

CaLRR-RLK1, a novel RD receptor-like kinase from Capsicum annuum and transcriptionally activated by CaHDZ27, act as positive regulator in Ralstonia solanacearum resistance.

BACKGROUND: Bacterial wilt caused by Ralstonia solanacearum is one of the most important diseases in pepper worldwide, however, the molecular mechanism underlying pepper resistance to bacterial wilt remains poorly understood. RESULTS: Herein, a novel RD leucine-rich repeat receptor-like kinase, CaLRR-RLK1, was functionally characterized in immunity against R. solanacearum. CaLRR-RLK1 was targeted exclusively to plasma membrane and was up-regulated by R. solanacearum inoculation (RSI) as well as by the exogenous application of salicylic acid (SA), methyl jasmonate (MeJA) or ethephon (ETH). The silencing of CaLRR-RLK1 led to enhanced susceptibility of pepper plants to RSI, accompanied by down-regulation of immunity-related genes including CaACO1, CaHIR1, CaPR4 and CaPO2. In contrast, transient overexpression of CaLRR-RLK1 triggered hypersensitive response (HR)-like cell death and H2O2 accumulation in pepper leaves, manifested by darker trypan blue and DAB staining respectively. In addition, the ectopic overexpression of CaLRR-RLK1 in tobacco plants enhanced resistance R. solanacearum, accompanied with the immunity associated marker genes including NtPR2, NtPR2, NtHSR203 and NtHSR515. Furthermore, it was found that CaHDZ27, a positive regulator in pepper response to RSI in our previous study, transcriptionally activated CaLRR-RLK1 by direct targeting its promoter probably in a CAATTATTG dependent manner. CONCLUSION: The study revealed that CaLRR-RLK1 confers pepper resistance to R. solanacearum as the direct targeting of CaHDZ27.

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

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 ß-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.

A novel leucine-rich repeat protein, CaLRR51, acts as a positive regulator in the response of pepper to Ralstonia solanacearum infection.

The leucine-rich repeat (LRR) proteins play important roles in the recognition of corresponding ligands and signal transduction networks in plant defence responses. Herein, a novel LRR protein from Capsicum annuum, CaLRR51, was identified and characterized. It was localized to the plasma membrane and transcriptionally up-regulated by Ralstonia solanacearum infection (RSI), as well as the exogenous application of salicylic acid (SA), jasmonic acid (JA) and ethephon (ETH). Virus-induced gene silencing of CaLRR51 significantly increased the susceptibility of pepper to RSI. By contrast, transient overexpression of CaLRR51 in pepper plants activated hypersensitive response (HR)-like cell death, and up-regulated the defence-related marker genes, including PO2, HIR1, PR1, DEF1 and ACO1. Moreover, ectopic overexpression of CaLRR51 in transgenic tobacco plants significantly enhanced the resistance to RSI. Transcriptional expression of the corresponding defence-related marker genes in transgenic tobacco plants was also found to be enhanced by the overexpression of CaLRR51, which was potentiated by RSI. These loss- and gain-of-function assays suggest that CaLRR51 acts as a positive regulator in the response of pepper to RSI. In addition, the putative signal peptide and transmembrane region were found to be required for plasma membrane targeting of CaLRR51, which is indispensable for the role of CaLRR51 in plant immunity.

CaCDPK15 positively regulates pepper responses to Ralstonia solanacearum inoculation and forms a positive-feedback loop with CaWRKY40 to amplify defense signaling.

CaWRKY40 is a positive regulator of pepper (Capsicum annum) response to Ralstonia solanacearum inoculation (RSI), but the underlying mechanism remains largely unknown. Here, we functionally characterize CaCDPK15 in the defense signaling mediated by CaWRKY40. Pathogen-responsive TGA, W, and ERE boxes were identified in the CaCDPK15 promoter (pCaCDPK15), and pCaCDPK15-driven GUS expression was significantly enhanced in response to RSI and exogenously applied salicylic acid, methyl jasmonate, abscisic acid, and ethephon. Virus-induced gene silencing (VIGS) of CaCDPK15 significantly increased the susceptibility of pepper to RSI and downregulated the immunity-associated markers CaNPR1, CaPR1, and CaDEF1. By contrast, transient CaCDPK15 overexpression significantly activated hypersensitive response associated cell death, upregulated the immunity-associated marker genes, upregulated CaWRKY40 expression, and enriched CaWRKY40 at the promoters of its targets genes. Although CaCDPK15 failed to interact with CaWRKY40, the direct binding of CaWRKY40 to pCaCDPK15 was detected by chromatin immunoprecipitation, which was significantly potentiated by RSI in pepper plants. These combined results suggest that RSI in pepper induces CaCDPK15 and indirectly activates downstream CaWRKY40, which in turn potentiates CaCDPK15 expression. This positive-feedback loop would amplify defense signaling against RSI and efficiently activate strong plant immunity.

Pepper CabZIP63 acts as a positive regulator during Ralstonia solanacearum or high temperature-high humidity challenge in a positive feedback loop with CaWRKY40.

CaWRKY40 is known to act as a positive regulator in the response of pepper (Capsicum annuum) to Ralstonia solanacearum inoculation (RSI) or high temperature-high humidity (HTHH), but the underlying mechanism remains elusive. Herein, we report that CabZIP63, a pepper bZIP family member, participates in this process by regulating the expression of CaWRKY40. CabZIP63 was found to localize in the nuclei, be up-regulated by RSI or HTHH, bind to promoters of both CabZIP63(pCabZIP63) and CaWRKY40(pCaWRKY40), and activate pCabZIP63- and pCaWRKY40-driven ß-glucuronidase expression in a C- or G-box-dependent manner. Silencing of CabZIP63 by virus-induced gene silencing (VIGS) in pepper plants significantly attenuated their resistance to RSI and tolerance to HTHH, accompanied by down-regulation of immunity- or thermotolerance-associated CaPR1, CaNPR1, CaDEF1, and CaHSP24. Hypersensitive response-mediated cell death and expression of the tested immunity- and thermotolerance-associated marker genes were induced by transient overexpression (TOE) of CabZIP63, but decreased by that of CabZIP63-SRDX. Additionally, binding of CabZIP63 to pCaWRKY40 was up-regulated by RSI or HTHH, and the transcript level of CaWRKY40 and binding of CaWRKY40 to the promoters of CaPR1, CaNPR1, CaDEF1 and CaHSP24 were up-regulated by TOE of CabZIP63. On the other hand, CabZIP63 was also up-regulated transcriptionally by TOE of CaWRKY40. The data suggest collectively that CabZIP63 directly or indirectly regulates the expression of CaWRKY40 at both the transcriptional and post-transcriptional level, forming a positive feedback loop with CaWRKY40 during pepper's response to RSI or HTHH. Altogether, our data will help to elucidate the underlying mechanism of crosstalk between pepper's response to RSI and HTHH.

Genome-wide identification and expression analysis of calcium-dependent protein kinase and its closely related kinase genes in Capsicum annuum.

As Ca2+ sensors and effectors, calcium-dependent protein kinases (CDPKs) play important roles in plant growth, development, and response to environmental cues. However, no CDPKs have been characterized in Capsicum annuum thus far. Herein, a genome wide comprehensive analysis of genes encoding CDPKs and CDPK-related protein kinases (CRKs) was performed in pepper, a total of 31 CDPK genes and five closely related kinase genes were identified, which were phylogenetically divided into four distinct subfamilies and unevenly distributed across nine chromosomes. Conserved sequence and exon-intron structures were found to be shared by pepper CDPKs within the same subfamily, and the expansion of the CDPK family in pepper was found to be due to segmental duplication events. Five CDPKs in the C. annuum variety CM334 were found to be mutated in the Chiltepin variety, and one CDPK present in CM334 was lost in Chiltepin. The majority of CDPK and CRK genes were expressed in different pepper tissues and developmental stages, and 10, 12, and 8 CDPK genes were transcriptionally modified by salt, heat, and Ralstonia solanacearum stresses, respectively. Furthermore, these genes were found to respond specifically to one stress as well as respond synergistically to two stresses or three stresses, suggesting that these CDPK genes might be involved in the specific or synergistic response of pepper to salt, heat, and R. solanacearum. Our results lay the foundation for future functional characterization of pepper CDPK and its closely related gene families.

Genome-wide identification and transcriptional expression analysis of mitogen-activated protein kinase and mitogen-activated protein kinase kinase genes in Capsicum annuum.

The tripartite mitogen-activated protein kinase (MAPK) signaling cascades have been implicated in plant growth, development, and environment adaptation, but a comprehensive understanding of MAPK signaling at genome-wide level is limited in Capsicum annuum. Herein, genome-wide identification and transcriptional expression analysis of MAPK and MAPK kinase (MAPKK) were performed in pepper. A total of 19 pepper MAPK (CaMAPKs) genes and five MAPKK (CaMAPKKs) genes were identified. Phylogenetic analysis indicated that CaMAPKs and CaMAPKKs could be classified into four groups and each group contains similar exon-intron structures. However, significant divergences were also found. Notably, five members of the pepper MAPKK family were much less conserved than those found in Arabidopsis, and 9 Arabidopsis MAPKs did not have orthologs in pepper. Additionally, 7 MAPKs in Arabidopsis had either two or three orthologs in the pepper genome, and six pepper MAPKs and one MAPKK differing in sequence were found in three pepper varieties. Quantitative real-time RT-PCR analysis showed that the majority of MAPK and MAPKK genes were ubiquitously expressed and transcriptionally modified in pepper leaves after treatments with heat, salt, and Ralstonia solanacearum inoculation as well as exogenously applied salicylic acid, methyl jasmonate, ethephon, and abscisic acid. The MAPKK-MAPK interactome was tested by yeast two-hybrid assay, the results showed that one MAPKK might interact with multiple MAPKs, one MAPK might also interact with more than one MAPKKs, constituting MAPK signaling networks which may collaborate in transmitting upstream signals into appropriate downstream cellular responses and processes. These results will facilitate future functional characterization of MAPK cascades in pepper.

Over-Expression of Rice CBS Domain Containing Protein, OsCBSX3, Confers Rice Resistance to Magnaporthe oryzae Inoculation.

Cystathionine ß-synthase (CBS) domain containing proteins (CDCPs) constitute a big family in plants and some members in this family have been implicated in a variety of biological processes, but the precise functions and the underlying mechanism of the majority of this family in plant immunity remain to be elucidated. In the present study, a CBS domain containing protein gene, OsCBSX3, is functionally characterized in rice resistance against Magnaporthe oryzae (M. oryzae). By quantitative real-time PCR, transcripts of OsCBSX3 are up-regulated significantly by inoculation of M. oryzae and the exogenously applied salicylic acid (SA) and methyl jasmonate (MeJA). OsCBSX3 is exclusively localized to the plasma membrane by transient expression of OsCBSX3 fused to green fluorescent protein (GFP) through approach of Agrobacterium infiltration in Nicotiana benthamiana leaves. The plants of homozygous T3 transgenic rice lines of over-expressing OsCBSX3 exhibit significant enhanced resistance to M. oryzae inoculation, manifested by decreased disease symptoms, and inhibition of pathogen growth detected in DNA. Consistently, the over-expression of OsCBSX3 enhances the transcript levels of immunity associated marker genes including PR1a, PR1b, PR5, AOS2, PAL, NH1, and OsWRKY13 in plants inoculated with M. oryzae. These results suggest that OsCBSX3 acts as a positive regulator in resistance of rice to M. oryzae regulated by SA and JA-mediated signaling pathways synergistically.

CaWRKY6 transcriptionally activates CaWRKY40, regulates Ralstonia solanacearum resistance, and confers high-temperature and high-humidity tolerance in pepper.

High temperature (HT), high humidity (HH), and pathogen infection often co-occur and negatively affect plant growth. However, these stress factors and plant responses are generally studied in isolation. The mechanisms of synergistic responses to combined stresses are poorly understood. We isolated the subgroup IIb WRKY family member CaWRKY6 from Capsicum annuum and performed quantitative real-time PCR analysis. CaWRKY6 expression was upregulated by individual or simultaneous treatment with HT, HH, combined HT and HH (HTHH), and Ralstonia solanacearum inoculation, and responded to exogenous application of jasmonic acid (JA), ethephon, and abscisic acid (ABA). Virus-induced gene silencing of CaWRKY6 enhanced pepper plant susceptibility to R. solanacearum and HTHH, and downregulated the hypersensitive response (HR), JA-, ethylene (ET)-, and ABA-induced marker gene expression, and thermotolerance-associated expression of CaHSP24, ER-small CaSHP, and Chl-small CaHSP. CaWRKY6 overexpression in pepper attenuated the HTHH-induced suppression of resistance to R. solanacearum infection. CaWRKY6 bound to and activated the CaWRKY40 promoter in planta, which is a pepper WRKY that regulates heat-stress tolerance and R. solanacearum resistance. CaWRKY40 silencing significantly blocked HR-induced cell death and reduced transcriptional expression of CaWRKY40. These data suggest that CaWRKY6 is a positive regulator of R. solanacearum resistance and heat-stress tolerance, which occurs in part by activating CaWRKY40.

SRC2-1 is required in PcINF1-induced pepper immunity by acting as an interacting partner of PcINF1.

Elicitins are elicitors that can trigger hypersensitive cell death in most Nicotiana spp., but their underlying molecular mechanism is not well understood. The gene Phytophthora capsici INF1 (PcINF1) coding for an elicitin from P. capsici was characterized in this study. Transient overexpression of PcINF1 triggered cell death in pepper (Capsicum annuum L.) and was accompanied by upregulation of the hypersensitive response marker, Hypersensitive Induced Reaction gene 1 (HIR1), and the pathogenesis-related genes SAR82, DEF1, BPR1, and PO2. A putative PcINF1-interacting protein, SRC2-1, was isolated from a pepper cDNA library by yeast two-hybrid screening and was observed to target the plasma membrane. The interaction between PcINF1 and SRC2-1 was confirmed by bimolecular fluorescence complementation and co-immunoprecipitation. Simultaneous transient overexpression of SRC2-1 and PcINF1 in pepper plants triggered intensive cell death, whereas silencing of SRC2-1 by virus-induced gene silencing blocked the cell death induction of PcINF1 and increased the susceptibility of pepper plants to P. capsici infection. Additionally, membrane targeting of the PcINF1-SRC2-1 complex was required for cell death induction. The C2 domain of SRC2-1 was crucial for SRC2-1 plasma membrane targeting and the PcINF1-SRC2-1 interaction. These results suggest that SRC2-1 interacts with PcINF1 and is required in PcINF1-induced pepper immunity.

Overexpression of a pepper CaERF5 gene in tobacco plants enhances resistance to Ralstonia solanacearum infection.

ETHYLENE RESPONSE FACTORs (ERF) transcription factors (TFs) constitute a large transcriptional regulator family belonging to the AP2/ERF superfamily and are implicated in a range of biological processes. However, the specific roles of individual ERF family members in biotic or abiotic stress responses and the underlying molecular mechanism still need to be elucidated. In the present study, a cDNA encoding a member of ethylene response factor (ERF) transcription factor, CaERF5, was isolated from pepper. Sequence analysis showed that CaERF5 contains a typical 59 amino acid AP2/ERF DNA-binding domain, two highly conserved amino acid residues (14th alanine (A) and 19th aspartic acid (D)), a putative nuclear localisation signal (NLS), a CMIX-2 motif in the N-terminal region and two putative MAP kinase phosphorylation site CMIX-5 and CMIX-6 motifs. It belongs to group IXb of the ERF subfamily. A CaERF5-green fluorescence protein (GFP) fusion transiently expressed in onion epidermal cells localised to the nucleus. CaERF5 transcripts were induced by Ralstonia solanacearum infection, salicylic acid (SA), methyl jasmonate (MeJA) and ethephon (ETH) treatments. Constitutive expression of the CaERF5 gene in tobacco plants upregulated transcript levels of a set of defence- related genes and enhanced resistance to R. solanacearum infection. Our results suggest that CaERF5 acts as a positive regulator in plant resistance to R. solanacearum infection and show that overexpression of this transcription factor can be used as a tool to enhance disease resistance in crop species.

Functional analysis and expressional characterization of rice ankyrin repeat-containing protein, OsPIANK1, in basal defense against Magnaporthe oryzae attack.

The ankyrin repeat-containing protein gene OsPIANK1 (AK068021) in rice (Oryza sativa L.) was previously shown to be upregulated following infection with the rice leaf blight pathogen Xanthomonas oryzae pv oryzae (Xoo). In this study, we further characterized the role of OsPIANK1 in basal defense against Magnaporthe oryzae (M.oryzae) by 5' deletion analysis of its promoter and overexpression of the gene. The promoter of OsPIANK1 with 1,985 bps in length was sufficient to induce the OsPIANK1 response to inoculation with M.oryzae and to exogenous application of methyl jasmonate (MeJA) or salicylic acid (SA), but not to exogenous application of abscisic acid (ABA). A TCA-element present in the region between -563 bp and -249 bp may be responsible for the OsPIANK1 response to both M.oryzae infection and exogenous SA application. The JERE box, CGTCA-box, and two MYB binding sites locating in the region between -1985 bp and -907 bp may be responsible for the response of OsPIANK1 to exogenous MeJA. OsPIANK1 expression was upregulated after inoculation with M.oryzae and after treatment with exogenous SA and MeJA. Overexpression of OsPIANK1 enhanced resistance of rice to M.oryzae, although it did not confer complete resistance. The enhanced resistance to M.oryzae was accompanied by enhanced transcriptional expression of SA- and JA-dependent genes such as NH1, WKRY13, PAL, AOS2, PR1b, and PR5. This evidence suggests that OsPIANK1 acted as a positive regulator in rice basal defense mediated by SA- and JA-signaling pathways.

Overexpression of a Chinese cabbage BrERF11 transcription factor enhances disease resistance to Ralstonia solanacearum in tobacco.

Ethylene-responsive factors (ERFs) play diverse roles in plant growth, developmental processes and stress responses. However, the roles and underlying mechanism of ERFs remain poorly understood, especially in non-model plants. In this study, a full length cDNA of ERF gene was isolated from the cDNA library of Chinese cabbage. According to sequence alignment, we found a highly conservative AP2/ERF domain, two nuclear localization signals, and an ERF-associated Amphiphilic Repression (EAR) motif in its C-terminal region. It belonged to VIIIa group ERFs sharing the highest sequence identity with AtERF11 in all of the ERFs in Arabidopsis and designated BrERF11. BrERF11-green fluorescence protein (GFP) transient expressed in onion epidermis cells localized to the nucleus. The transcript levels of BrERF11 were induced by exogenous salicylic acid (SA), methyl jasmonate (MeJA), ethephon (ETH), and hydrogen peroxide (H(2)O(2)). Constitutive expression of BrERF11 enhanced tolerance to Ralstonia solanacearum infection in transgenic tobacco plants, which was coupled with hypersensitive response (HR), burst of H(2)O(2) and upregulation of defense-related genes including HR marker genes, SA-, JA-dependent pathogen-related genes and ET biosynthesis associated genes and downregulation of CAT1, suggesting BrERF11 may participate in pathogen-associated molecular pattern (PAMP)- and effector-triggered immunity (PTI and ETI) mediated by SA-, JA- and ET-dependent signaling mechanisms.

CaWRKY40, a WRKY protein of pepper, plays an important role in the regulation of tolerance to heat stress and resistance to Ralstonia solanacearum infection.

WRKY proteins form a large family of plant transcription factors implicated in the modulation of numerous biological processes, such as growth, development and responses to various environmental stresses. However, the roles of the majority WRKY family members, especially in non-model plants, remain poorly understood. We identified CaWRKY40 from pepper. Transient expression in onion epidermal cells showed that CaWRKY40 can be targeted to nuclei and activates expression of a W-box-containing reporter gene. CaWRKY40 transcripts are induced in pepper by Ralstonia solanacearum and heat shock. To assess roles of CaWRKY40 in plant stress responses we performed gain- and loss-of-function experiments. Overexpression of CaWRKY40 enhanced resistance to R. solanacearum and tolerance to heat shock in tobacco. In contrast, silencing of CaWRKY40 enhanced susceptibility to R. solanacearum and impaired thermotolerance in pepper. Consistent with its role in multiple stress responses, we found CaWRKY40 transcripts to be induced by signalling mechanisms mediated by the stress hormones salicylic acid (SA), jasmonic acid (JA) and ethylene (ET). Overexpression of CaWRKY40 in tobacco modified the expression of hypersensitive response (HR)-associated and pathogenesis-related genes. Collectively, our results suggest that CaWRKY40 orthologs are regulated by SA, JA and ET signalling and coordinate responses to R. solanacearum attacks and heat stress in pepper and tobacco.

Differential gene expression in pepper (Capsicum annuum) exposed to UV-B.

In the present paper, complementary DNA-amplified fragment length polymorphism (cDNA-AFLP) was used to examine and identify differentially expressed genes in Capsicum annuum exposed to UV-B irradiation. Around 4000 transcript derived fragments (TDFs) were visualized and in total 183 TDFs were isolated, sequenced and analyzed by Blast 2 go. Among these TDFs, 84 of them showed homology to known genes. There were 43 TDFs showing up-regulated expression, 24 TDFs showing down-regulated expression and 29 TDFs showing both up-regulated and down-regulated expression, respectively. Some of these TDFs were found to be in response/related to UV-B stress, including carbonic anhydrase, calcium-dependent protein, thionin-like protein, bzip protein and so on. In particular, chlorophyll a/b binding protein (Capcab) responding to UV-B stress was cloned. It was concluded that Capcab could play a protective role in plant anti-UV-B and maintaining photosynthetic rate under UV-B stress.