This study aimed to evaluate the effects of specific LED light spectra on the growth and physiology of Changchuan No. 3 Capsicum annuum L. seedlings. The experimental design involved exposing pepper seedlings to six different spectral light combinations for 7, 14, and 21 days, with the treatments consisting of 2R1B1Y (red/blue/yellow = 2:1:1), 2R1B1FR (red/blue/far-red = 2:1:1), 2R1B1P (red/blue/purple = 2:1:1), 4R2B1G (red/blue/green = 4:2:1), 2R1B1G (red/blue/green = 2:1:1), and 2R1B (red/blue = 2:1). The results demonstrated distinct spectral regulation of seedling development: compared to the white light (CK), the 2R1B1FR (far-red light supplementation) treatment progressively stimulated stem elongation, increasing plant height and stem diameter by 81.6% and 25.9%, respectively, at day 21, but resulted in a more slender stem architecture. The 2R1B1G (balanced green light) treatment consistently promoted balanced growth, culminating in the highest seedling vigor index at the final stage. The 2R1B1P (purple light supplementation) treatment exhibited a strong promotive effect on root development, which became most pronounced at day 21 (126% increase in root dry weight), while concurrently enhancing soluble sugar content and reducing oxidative stress. Conversely, the 2R1B1Y (yellow light supplementation) treatment increased MDA content by 70% and led to a reduction in chlorophyll accumulation, while 2R1B (basic red–blue) resulted in lower biomass accumulation compared to the superior spectral treatments. The 4R2B1G (low green ratio) treatment showed context-dependent outcomes. This study elucidates how targeted spectral compositions, particularly involving far-red and green light, can optimize pepper seedling quality by modulating photomorphogenesis, carbon allocation, and stress physiology. The findings provide a mechanistic basis for designing efficient LED lighting protocols in controlled-environment agriculture to enhance pepper nursery production.

As important transcription factors, WRKYs play a vital role in the defense response of plants against the invasion of multiple pathogens. Though some WRKY members have been reported to participate in pepper immunity in response to Ralstonia solanacearum infection, the functions of the majority of WRKY members are still unknown. Herein, CaWRKY22b was cloned from the pepper genome and its function against R. solanacearum was analyzed. The transcript abundance of CaWRKY22b was significantly increased in response to the infection of R. solanacearum and the application of exogenous methyl jasmonate (MeJA). Subcellular localization assay in the leaves of Nicotiana benthamiana showed that CaWRKY22b protein was targeted to the nuclei. Agrobacterium-mediated transient expression in pepper leaves indicated that CaWRKY22b overexpression triggered intensive hypersensitive response-like cell death, H2O2 accumulation, and the up-regulation of defense- and JA-responsive genes, including CaHIR1, CaPO2, CaBPR1, and CaDEF1. Virus-induced gene silencing assay revealed that knock-down of CaWRKY22b attenuated pepper’s resistance against R. solanacearum and the up-regulation of the tested defense- and jasmonic acid (JA)-responsive genes. We further assessed the role of CaWRKY22b in modulating the expression of JA-responsive CaDEF1, and the result demonstrated that CaWRKY22b trans-activated CaDEF1 expression by directly binding to its upstream promoter. Collectively, our results suggest that CaWRKY22b positively regulated pepper immunity against R. solanacearum in a manner associated with JA signaling, probably by modulating the expression of JA-responsive CaDEF1.

As an important member of mitogen-activated protein kinase (MAPK) cascades, MAPKs play an important role in plant defense response against biotic and abiotic stresses; however, the involvement of the majority of the MAPK family members against Ralstonia solanacearum and heat stress (HS) remains poorly understood. In the present study, CaMAPK1 was identified from the genome of pepper and its function against R. solanacearum and HS was analyzed. The transcript accumulations of CaMAPK1 and the activities of its native promoter were both significantly induced by R. solanacearum inoculation, HS, and the application of exogenous hormones, including SA, MeJA, and ABA. Transient expression of CaMAPK1 showed that CaMAPK1 can be targeted throughout the whole cells in Nicotiana benthamiana and triggered chlorosis and hypersensitive response-like cell death in pepper leaves, accompanied by the accumulation of H2O2, and the up-regulations of hormones- and H2O2-associated marker genes. The knock-down of CaMAPK1 enhanced the susceptibility to R. solanacearum partially by down-regulating the expression of hormones- and H2O2-related genes and impairing the thermotolerance of pepper probably by attenuating CaHSFA2 and CaHSP70-1 transcripts. Taken together, our results revealed that CaMAPK1 is regulated by SA, JA, and ABA signaling and coordinates responses to R. solanacearum infection and HS in pepper.

Pepper (Capsicum annuum L.) suffers severe quality and yield loss from oomycete diseases caused by Phytophthora capsici. CaSGT1 was previously determined to positively regulate the immune response of pepper plants against P. capsici, but by which mechanism remains elusive. In the present study, the potential interacting proteins of CaSGT1 were isolated from pepper using a yeast two-hybrid system, among which CaARP1 was determined to interact with CaSGT1 via bimolecular fluorescence complementation (BiFC) and microscale thermophoresis (MST) assays. CaARP1 belongs to the auxin-repressed protein family, which is well-known to function in modulating plant growth. The transcriptional and protein levels of CaARP1 were both significantly induced by infection with P. capsici. Silencing of CaARP1 promotes the vegetative growth of pepper plants and attenuates its disease resistance to P. capsici, as well as compromising the hypersensitive response-like cell death in pepper leaves induced by PcINF1, a well-characterized typical PAMP from P. capsici. Chitin-induced transient expression of CaARP1 in pepper leaves enhanced its disease resistance to P. capsici, which is amplified by CaSGT1 co-expression as a positive regulator. Taken together, our result revealed that CaARP1 plays a dual role in the pepper, negatively regulating the vegetative growth and positively regulating plant immunity against P. capsici in a manner associated with CaSGT1.

Plants have evolved a sophisticated immune system to defend against invasion by pathogens. In response, pathogens deploy copious effectors to evade the immune responses. However, the molecular mechanisms used by pathogen effectors to suppress plant immunity remain unclear. Herein, we report that an effector secreted by Ralstonia solanacearum, RipAK, modulates the transcriptional activity of the ethylene-responsive factor ERF098 to suppress immunity and dehydration tolerance, which causes bacterial wilt in pepper (Capsicum annuum L.) plants. Silencing ERF098 enhances the resistance of pepper plants to R. solanacearum infection not only by inhibiting the host colonization of R. solanacearum but also by increasing the immunity and tolerance of pepper plants to dehydration and including the closure of stomata to reduce the loss of water in an abscisic acid signal-dependent manner. In contrast, the ectopic expression of ERF098 in Nicotiana benthamiana enhances wilt disease. We also show that RipAK targets and inhibits the ERF098 homodimerization to repress the expression of salicylic acid-dependent PR1 and dehydration tolerance-related OSR1 and OSM1 by cis-elements in their promoters. Taken together, our study reveals a regulatory mechanism used by the R. solanacearum effector RipAK to increase virulence by specifically inhibiting the homodimerization of ERF098 and reprogramming the transcription of PR1, OSR1, and OSM1 to boost susceptibility and dehydration sensitivity. Thus, our study sheds light on a previously unidentified strategy by which a pathogen simultaneously suppresses plant immunity and tolerance to dehydration by secreting an effector to interfere with the activity of a transcription factor and manipulate plant transcriptional programs.

Bacterial wilt, a severe disease involving vascular system blockade, is caused by Ralstonia solanacearum. Although both plant immunity and dehydration tolerance might contribute to disease resistance, whether and how they are related remains unclear. Herein, we showed that immunity against R. solanacearum and dehydration tolerance are coupled and regulated by the CaPti1-CaERF3 module. CaPti1 and CaERF3 are members of the serine/threonine protein kinase and ethylene-responsive factor families, respectively. Expression profiling revealed that CaPti1 and CaERF3 were upregulated by R. solanacearum inoculation, dehydration stress, and exogenously applied abscisic acid (ABA). They in turn phenocopied each other in promoting resistance of pepper (Capsicum annuum) to bacterial wilt not only by activating salicylic acid-dependent CaPR1, but also by activating dehydration tolerance-related CaOSM1 and CaOSR1 and inducing stomatal closure to reduce water loss in an ABA signaling-dependent manner. Our yeast two hybrid assay showed that CaERF3 interacted with CaPti1, which was confirmed using co-immunoprecipitation, bimolecular fluorescence complementation, and pull-down assays. Chromatin immunoprecipitation and electrophoretic mobility shift assays showed that upon R. solanacearum inoculation, CaPR1, CaOSM1, and CaOSR1 were directly targeted and positively regulated by CaERF3 and potentiated by CaPti1. Additionally, our data indicated that the CaPti1-CaERF3 complex might act downstream of ABA signaling, as exogenously applied ABA did not alter regulation of stomatal aperture by the CaPti1-CaERF3 module. Importantly, the CaPti1-CaERF3 module positively affected pepper growth and the response to dehydration stress. Collectively, the results suggested that immunity and dehydration tolerance are coupled and positively regulated by CaPti1-CaERF3 in pepper plants to enhance resistance against R. solanacearum.

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, H 2 O 2 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.

Bacterial wilt, a severe disease that affects over 250 plant species, is caused by Ralstonia solanacearum through vascular system blockade. Although both plant immunity and dehydration tolerance might contribute to disease resistance, whether and how they are related are still unclear. Herein, we provide evidence that immunity against R. solanacearum and dehydration tolerance are coupled and regulated by CaPti1-CaERF4 module. By expression profiling, virus-induced gene silencing in pepper and overexpression in Nicotiana benthamiana, both CaPti1 and CaERF4 were upregulated by R. solanacearum inoculation, dehydration stress and exogenously applied ABA. They in turn phenocopied with each other in promoting pepper resistance to bacterial wilt not only by activating HR cell death and SA-dependent CaPR1, but also by activating dehydration tolerance related CaOSM1 and CaOSR1, and stomata closure to reduce water loss in ABA signaling dependent manner. Yeast-two hybrid assay showed that CaERF4 interacts with CaPti1, which was confirmed by co-immunoprecipitation and pull-down assays. Chromatin immunoprecipitation and electrophoretic mobility shift assay showed that, upon R. solanacearum inoculation, CaPR1, CaOSM1 and CaOSR1 were directly targeted and positively regulated by CaERF4 via binding GCC-box or DRE-box, which was potentiated by CaPti1. In addition, our data indicate that CaPti1-CaERF4 complex might act downstream ABA signaling, since the exogenously applied ABA did not alter stomata aperture regulated by CaPti1-CaERF4 module. Importantly, CaPti1-CaERF4 module was found also acts positively in pepper growth and response to dehydration stress. Collectively, the results suggest that immunity and dehydration tolerance are coupled and positively regulated by CaPti1-CaERF4 in pepper plants to enhance resistance against R. solanacearum. Summary Pepper immunity and dehydration tolerance are coupled and regulated by the CaPti1-CaERF4 module partially in a way related to ABA signaling and stomata closure.

AbstractCaWRKY40 was previously found to be transcriptionally up‐regulated by Ralstonia solanacearum inoculation (RSI) or heat stress (HS), but the underlying mechanism remains unknown. Herein, we report that a double‐W box‐element (DWE) in the promoter of CaWRKY40 is critical for these responses. The upstream W box unit WI of this composite element is crucial for preferential binding by CaWRKY40 and responsiveness to RSI or HS. DWE‐driven CaWRKY40 can be transcriptionally and nonspecifically regulated by itself and by CaWRKY58 and CaWRKY27. The DWE was also found in the promoters of CaWRKY40 orthologs, including AtWRKY40, VvWRKY40, GmWRKY40, CplWRKY40, SaWRKY40, SpWRKY40, NtWRKY40, and NaWRKY40. DWEAtWRKY40 was analogous to DWECaWRKY40 by responding to RSI or HS and AtWRKY40 expression. These data suggest that a conserved response of plants to pathogen infection or HS is probably mediated by binding of the DWE by WRKY40.

ChiIV3, a chitinase of pepper (Capsicum annuum), stimulates cell death in pepper plants. However, there are only scarce reports on its role in resistance against bacterial wilt disease such as that caused by Ralstonia solanacearum and their transcriptional regulation. In this study, the silencing of ChiIV3 in pepper plants significantly reduced the resistance to R. solanacearum. The transcript of ChiIV3 was induced by R. solanacearum inoculation (RSI) as well as exogenous application of methyl jasmonate and abscisic acid. The bioinformatics analysis revealed that the ChiIV3 promoter consists of multiple stress-related cis elements, including six W-boxes and one MYB1AT. With the 5' deletion assay in the ChiIV3 promoter, the W4-box located from -640 to -635 bp was identified as the cis element that is required for the response to RSI. In addition, the W4-box element was shown to be essential for the binding of the ChiIV3 promoter by the WRKY40 transcription factor, which is known to positively regulate the defense response to R. solanacearum. Site-directed mutagenesis in the W4-box sequence impaired the binding of WRKY40 to the ChiIV3 promoter. Subsequently, the transcription of ChiIV3 decreased in WRKY40-silenced pepper plants. These results demonstrated that the expression of the defense gene ChiIV3 is controlled through multiple modes of regulation, and WRKY40 directly binds to the W4-box element of the ChiIV3 promoter region for its transcriptional regulation.

Despite the involvement of many members of the chitinase family in plant immunity, the precise functions of the majority of the members remain poorly understood. Herein, the gene ChiIV3 in Capsicum annuum encoding a chitinase protein containing a chitin binding domain and targeting to the plasma membrane was found to be induced by Phytophthora capsici inoculation (PCI) and applied chitin treatment. Besides its direct inhibitory effect on growth of Phytophthora capsici (P. capsici), ChiIV3 was also found by virus-induced gene silencing (VIGS) and transient overexpression (TOE) in pepper plants to act as a positive regulator of plant cell death and in triggering defense signaling and upregulation of PR (pathogenesis related) genes against PCI. A 5′ deletion assay revealed that pChiIV3−712 to −459 bp was found to be sufficient for ChiIV3’ response to PCI. Furthermore, a mutation assay indicated that W-box−466 to −461 bp in pChiIV3−712 to −459 bp was noted to be the PCI-responsible element. These results collectively suggest that ChiIV3 acts as a likely antifungal protein and as a receptor for unidentified chitin in planta to trigger cell death and defense signaling against PCI.

PcINF1 was previously found to induce pepper defense response by interacting with SRC2-1, but the underlying mechanism remains uninvestigated. Herein, we describe the involvement of SGT1 in the PcINF1/SRC2-1-induced immunity. SGT1 was observed to be up-regulated by Phytophthora capsici inoculation and synergistically transient overexpression of PcINF1/SRC2-1 in pepper plants. SGT1-silencing compromised HR cell death, blocked H2O2 accumulation, and downregulated HR-associated and hormones-dependent marker genes' expression triggered by PcINF1/SRC2-1 co-overexpression. The interaction between SRC2-1 and SGT1 was found by the yeast two hybrid system and was further confirmed by bimolecular fluorescence complementation and co-immunoprecipitation analyses. The SGT1/SRC2-1 interaction was enhanced by transient overexpression of PcINF1 and Phytophthora capsici inoculation, and SGT1-silencing attenuated PcINF1/SRC2-1 interaction. Additionally, by modulating subcellular localizations of SRC2-1, SGT1, and the interacting complex of SGT1/SRC2-1, it was revealed that exclusive nuclear targeting of the SGT1/SRC2-1 complex blocks immunity triggered by formation of SGT1/SRC2-1, and a translocation of the SGT1/SRC2-1 complex from the plasma membrane and cytoplasm to the nuclei upon the inoculation of P. capsici. Our data demonstrate that the SGT1/SRC2-1 interaction, and its nucleocytoplasmic partitioning, is involved in pepper's immunity against P. capsici, thus providing a molecular link between Ca(2+) signaling associated SRC2-1 and SGT1-mediated defense signaling.

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 β-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.

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 (p CaCDPK15 ), and p CaCDPK15 -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 p CaCDPK15 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.

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.

Cystathionine β-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.

辣椒CaWRKY40的转录表达受到病原菌和高温的诱导并在辣椒抗青枯病以及耐高温中起着重要的作用,但其应答青枯菌侵染和高温胁迫表达的调节机制还不清楚.为了进一步分离调节CaWRKY40表达的转录因子,建立辣椒酵母单杂交cDNA文库,以CaWRKY40启动子的青枯病原菌及高温应答区域CaWRKY40p(W40p)为诱饵,筛选cDNA文库,共获得56个阳性克隆.序列分析结果表明,有27个克隆为非重复序列,对其推定氨基酸序列进行同源比对和功能结构域分析,发现有1个推定转录因子,即ARF(Auxin response factor),以及其他蛋白如eIF(Eukaryotic initiation factor,真核起始因子),EF(延伸因子,Elongation factor)等.进一步通过染色质免疫共沉淀验证这些转录因子与W40p的结合.剖析CaWRKY40表达的分子机制有利于深入解析CaWRKY40介导的辣椒抗病及耐高温分子机制.

促分裂原活化蛋白激酶(MAPK)在植物生长、发育和适应环境中起着十分重要的作用,但对该家族多数成员的表达调控机制认识仍十分有限.前期实验已分离获得辣椒一个MAPK家族成员CaMA PK7,并发现该基因的表达受病原菌、高温等逆境的诱导,但其表达调控机制仍不清楚.本研究进一步分离了CaMA PK7的启动子序列(pCaMAPK7),发现其TATA框位于-165 bp与-170 bp之间,此外还发现W-box,HSE,TCA,LTR,ERE等多种与生物及非生物逆境胁迫应答相关的顺式作用元件.通过构建该启动子与报告基因GUS融合表达载体pCaMA PK7::GUS,利用农杆菌介导的本氏烟草叶片瞬间表达系统分析发现pCaMA PK7可应答青枯菌接种及水杨酸(SA)、茉莉酸甲酯(MeJA)、油菜素内酯(BR)、脱落酸(ABA)和乙烯(ET)等外源激素处理,表明CaMA PK7应答青枯菌、外源激素处理主要是通过该启动子介导的转录调节实现的.

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.

A full length cDNA of CaMAPK7 was amplified by PCR with specific primers in normalized cDNA library of Capsicum annuum as template. It contains an intact open reading frame encoding a protein of 370 amino acid in length sharing 84% to 97% sequence identities to MAPKs genes from other plant species. The results of real time PCR showed that CaMAPK7 exhibited a constitutive transcriptional expression level in different organs of pepper plants,and the transcriptional expression of CaMAPK7 in pepper leaves can be induced by Ralstonia solanacearum infection,heat shock treatment and exogenous application of salicylic acid(SA),methyljasmonate(MeJA),ethephon(ETH) treatments,but was down-regulated by abscisic acid(ABA). It is concluded that CaMAPK7 may play vital role in pepper response to pathogen inoculation and heat shock.