Jiaowen Pan

ZmMKK4, a novel group C mitogen‐activated protein kinase kinase in maize (Zea mays), confers salt and cold tolerance in transgenic Arabidopsis

Mitogen-activated protein kinase (MAPK) cascades are signalling modules that transduce extracellular signalling to a range of cellular responses. Plant MAPK cascades have been implicated in development and stress response. In this study, we isolated a novel group C MAPKK gene, ZmMKK4, from maize. Northern blotting analysis revealed that the ZmMKK4 transcript expression was up-regulated by cold, high salt and exogenous H(2)O(2,) but down-regulated by exogenous abscisic acid (ABA). Over-expression of ZmMKK4 in Arabidopsis conferred tolerance to cold and salt stresses by increased germination rate, lateral root numbers, plant survival rate, chlorophyll, proline and soluble sugar contents, and antioxidant enzyme [peroxidase (POD), catalase (CAT)] activities compared with control plants. Furthermore, ZmMKK4 enhanced a 37 kDa kinase activity after cold and salt stresses. RT-PCR analysis revealed that the transcript levels of stress-responsive transcription factors and functional genes were higher in ZmMKK4-over-expressing plants than in control plants. In addition, ZmMKK4 protein is localized in the nucleus. Taken together, these results indicate that ZmMKK4 is a positive regulator of salt and cold tolerance in plants.

ZmLEA3, a Multifunctional Group 3 LEA Protein from Maize (Zea mays L.), is Involved in Biotic and Abiotic Stresses

Late embryogenesis abundant (LEA) proteins accumulate to high levels during the late stage of seed maturation and in response to water deficit, and are involved in protecting higher plants from damage caused by environmental stresses, especially drought. In the present study, a novel maize (Zea mays L.) group 3 LEA gene, ZmLEA3, was identified and later characterized using transgenic tobacco plants to investigate its functions in abiotic and biotic stresses. Transcript accumulation demonstrated that ZmLEA3 was induced in leaves by high salinity, low temperature, osmotic and oxidative stress as well as by signaling molecules such as ABA, salicylic acid (SA) and methyl jasmonate (MeJA). The transcript of ZmLEA3 could also be induced by pathogens [Pseudomonas syringae pv. tomato DC3000 (pst dc3000)]. ZmLEA3 is located in the cytosol and the nucles. Further study indicated that the ZmLEA3 protein could bind Mn(2+), Fe(3+), Cu(2+) and Zn(2+). Overexpression of ZmLEA3 in transgenic tobacco (Nicotiana tabacum) and yeast (GS115) conferred tolerance to osmotic and oxidative stresses. Interestingly, we also found that overexpression of ZmLEA3 in transgenic tobacco increased the hypersensitive cell death triggered by pst dc3000 and enhanced the expression of PR1a, PR2 and PR4 when compared with the wild type. Thus, we proposed that the ZmLEA3 protein plays a role in protecting plants from damage by protecting protein structure and binding metals under osmotic and oxidative stresses. In addition, ZmLEA3 may also enhance transgenic plant tolerance to biotic stress.

Genome-wide identification and expression analysis of calcium-dependent protein kinase in maize

BackgroundCalcium-dependent protein kinases (CDPKs) have been shown to play important roles in various physiological processes, including plant growth and development, abiotic and biotic stress responses and plant hormone signaling in plants.ResultsIn this study, we performed a bioinformatics analysis of the entire maize genome and identified 40 CDPK genes. Phylogenetic analysis indicated that 40 ZmCPKs can be divided into four groups. Most maize CDPK genes exhibited different expression levels in different tissues and developmental stages. Twelve CDPK genes were selected to respond to various stimuli, including salt, drought and cold, as well as ABA and H2O2. Expression analyses suggested that maize CDPK genes are important components of maize development and multiple transduction pathways.ConclusionHere, we present a genome-wide analysis of the CDPK gene family in maize for the first time, and this genomic analysis of maize CDPK genes provides the first step towards a functional study of this gene family in maize.

ZmMPK17, a novel maize group D MAP kinase gene, is involved in multiple stress responses

Plant mitogen-activated protein kinase (MAPK) cascades play a pivotal role in a range of biotic and abiotic stress responses. In this study, we isolated a novel group D MAPK gene, ZmMPK17, from maize (Zea mays L.). ZmMPK17 is localized mainly to the nucleus and its C-terminal domain extension is believed to be essential for this. Northern-blot analysis indicated that ZmMPK17 transcription is involved in response to exogenous signaling molecules such as abscisic acid, hydrogen peroxide, salicylic acid, jasmonic acid and ethylene and induced by low temperature and osmotic stress. Hydrogen peroxide and Ca2+ mediate PEG-induced downregulation of ZmMPK17 at transcription level and Ca2+ also mediates low temperature-induced expression of ZmMPK17. Overexpression of ZmMPK17 in tobacco (Nicotoniatobaccum) accumulated less reactive oxygen species under osmotic stress by affecting antioxidant defense systems. Transgenic tobacco exhibited enhanced tolerance to cold by means of an increased germination rate, and increased proline and soluble sugar levels relative to control plants. The transcription levels of NtERD10 genes were higher in ZmMPK17-overexpressing lines than in control plants under cold and osmotic stress conditions. ZmMPK17-overexpressing plants displayed enhanced resistance to viral pathogens, and the expression of the pathogenesis-related gene PR1a was significantly increased, indicating that ZmMPK17 might be involved in SA-mediated pathogen defense-signaling pathways.

ZmMKK1, a novel group A mitogen-activated protein kinase kinase gene in maize, conferred chilling stress tolerance and was involved in pathogen defense in transgenic tobacco

As an important intracellular signaling module, the mitogen-activated protein kinase (MAPK) cascades have been previously implicated in signal transduction during plants responsing to various environmental stresses as well as pathogen attack. The mitogen-activated protein kinase kinase acts as the convergent point of MAPK cascades during a variety of stress signaling. In this study, a novel MAPKK gene, ZmMKK1, in maize (Zea mays L.) belonging to group A MAPKK was isolated and functionally characterized. ZmMKK1 was mainly localized in the cytoplasm and its constitutive kinase-active form ZmMKK1DD was localized in both cytoplasm and nucleus. QRT-PCR analysis uncovered that ZmMKK1 expression was triggered by abiotic and biotic stresses and exogenous signaling molecules. Moreover, hydrogen peroxide (H2O2) and Ca(2+) mediated 12°C-induced up-regulated expressing of ZmMKK1 at mRNA level. Ectopic expression of ZmMKK1 in tobacco (Nicotiana tabacum) conferred tolerance to chilling stress by higher antioxidant enzyme activities, more accumulation of osmoregulatory substances and more significantly up-expression of ROS-related and stress-responsive genes compared with empty vector control plants. Furthermore, ZmMKK1 played differential functions in biotrophic versus necrotrophic pathogen-induced responses. These results suggested ZmMKK1 played a crucial role in chilling stress and pathogen defense in plants.

A maize calcium-dependent protein kinase gene, ZmCPK4, positively regulated abscisic acid signaling and enhanced drought stress tolerance in transgenic Arabidopsis

Calcium-dependent protein kinases (CDPKs) play essential roles in calcium-mediated signal transductions in plant response to abiotic stress. Several members have been identified to be regulators for plants response to abscisic acid (ABA) signaling. Here, we isolated a subgroup I CDPK gene, ZmCPK4, from maize. Quantitative real time PCR (qRT-PCR) analysis revealed that the ZmCPK4 transcripts were induced by various stresses and signal molecules. Transient and stable expression of the ZmCPK4-GFP fusion proteins revealed ZmCPK4 localized to the membrane. Moreover, overexpression of ZmCPK4 in the transgenic Arabidopsis enhanced ABA sensitivity in seed germination, seedling growth and stomatal movement. The transgenic plants also enhanced drought stress tolerance. Taken together, the results suggest that ZmCPK4 might be involved in ABA-mediated regulation of stomatal closure in response to drought stress.

ZmMKK3, a novel maize group B mitogen-activated protein kinase kinase gene, mediates osmotic stress and ABA signal responses

Mitogen-activated protein kinase (MAPK) cascades are important intracellular signaling modules and function as a convergent point for crosstalk during abiotic stress signaling. In this article, we isolated a novel group B MAPKK gene, ZmMKK3, from Zea mays. ZmMKK3 protein might be localized in both the cytoplasm and the nucleus. RNA blot analysis indicated that the ZmMKK3 transcription was up-regulated by abscisic acid (ABA), hydrogen peroxide (H(2)O(2)) and PEG, and that H(2)O(2) mediated PEG-induced expression of ZmMKK3. Constitutive expression of ZmMKK3 in Nicotiana tabacum reduced H(2)O(2) accumulation under osmotic stress by affecting antioxidant defense systems and alleviated reactive oxygen species-mediated injury under oxidative stress. Transgenic tobacco exhibited attenuated ABA sensitivity by means of an increased germination rate and main root growth. Taken together, these results indicate that ZmMKK3 is a positive regulator of osmotic tolerance and ABA signaling in plants.

ZmMKK4 regulates osmotic stress through reactive oxygen species scavenging in transgenic tobacco

Mitogen-activated protein kinase kinase (MAPKKs) are important components of MAPK cascades, which are universal signal transduction modules and play important role in regulating both plant development and biotic or abiotic stress responses. In this study, we identified the group C MAPKK gene, ZmMKK4, in maize (Zea mays L.). Overexpression of ZmMKK4 in tobacco enhanced tolerance to osmotic stress by increased proline content and antioxidant enzyme (POD) activities compared with wild-type plants. RT-PCR revealed that one peroxidase (POX) gene, NtPOX1, was higher in ZmMKK4-overexpressing plants than in the wild-type plants. In addition, the accumulation of reactive oxygen species (ROS) in ZmMKK4-overexpressing plants is much less than that of wild-type plants. These results suggest that ZmMKK4 may be involved in ROS signaling. Taken together, these results indicate that ZmMKK4 is a positive regulator of osmotic stress by regulating scavenging of ROS in plants.

Identification of mitogen-activated protein kinase kinase gene family and MKK-MAPK interaction network in maize.

Plant mitogen-activated protein kinases (MAPK) are involved in important processes, including stress signaling and development. MAPK kinases (MAPKK, MKK) have been investigated in several plant species including Arabidopsis thaliana, Oryza sativa, Populus trichocarpa, and Brachypodium distachyon. In the present study, nine putative maize MKK genes have been identified. Analysis of the conserved protein motifs, exon-intron junctions and intron phase has revealed high levels of conservation within the phylogenetic groups. Next, we defined four new ZmMKK-ZmMPK interactions using yeast two-hybrid. Finally, we examined the biological functions of the ZmMKK4 gene. Overexpression of ZmMKK4 in Arabidopsis conferred tolerance to oxidative stress by increased germination rate and early seedling growth compared with WT plants. Taken together, we provide a comprehensive bioinformatics analysis of the MKK gene family in maize genome and our data provide an important foundation for further functional study of MAPK and MKK families in maize.

The overexpression of a maize mitogen‐activated protein kinase gene (ZmMPK5) confers salt stress tolerance and induces defence responses in tobacco

As sessile organisms, plants are exposed to potential dangers, including multiple biotic and abiotic stresses. The mitogen-activated protein kinase (MAPK) is a universal signalling pathways involved in these processes. A previous study showed that maize ZmMPK5 is induced by various stimuli at transcriptional and post-translational levels. In this study, ZmMPK5 was overexpressed in tobacco to further analyse its biological functions. Under salt and oxidative stresses, ZmMPK5-overexpressing lines displayed less severe damage and stronger growth phenotypes corresponding to a series of physiological changes. In addition, the transgenic lines accumulated less reactive oxygen species (ROS) and had higher levels of antioxidant enzyme activity and metabolites than wild-type (WT) plants following NaCl treatment. Quantitative RT-PCR revealed that the expression of ROS-related and stress-responsive genes was higher in transgenic plants than in WT plants. Furthermore, transgenic lines exhibited enhanced resistance to viral pathogens, and expressed constitutively higher transcript levels of pathogenesis-related genes, such as PR1a, PR4, PR5 and EREBP. Taken together, these results demonstrated that ZmMPK5 is involved in salt stress, oxidative stress and pathogen defence signalling pathways, and its function may be at least partly devoted to efficiently eliminating ROS accumulation under salt stress.