Maize protein phosphatase gene family: identification and molecular characterization

Background

Protein phosphatases (PPs) play critical roles in various cellular processes through the reversible protein phosphorylation that dictates many signal transduction pathways among organisms. Recently, PPs in Arabidopsis and rice have been identified, while the whole complement of PPs in maize is yet to be reported.

Results

In this study, we have identified 159 PP-encoding genes in the maize genome. Phylogenetic analyses categorized the ZmPP gene family into 3 classes (PP2C, PTP, and PP2A) with considerable conservation among classes. Similar intron/exon structural patterns were observed in the same classes. Moreover, detailed gene structures and duplicative events were then researched. The expression profiles of ZmPPs under different developmental stages and abiotic stresses (including salt, drought, and cold) were analyzed using microarray and RNA-seq data. A total of 152 members were detected in 18 different tissues representing distinct stages of maize plant developments. Under salt stress, one gene was significantly up-expressed in seed root (SR) and one gene was down-expressed in primary root (PR) and crown root (CR), respectively. As for drought stress condition, 13 genes were found to be differentially expressed in leaf, out of which 10 were up-regulated and 3 exhibited down-regulation. Additionally, 13 up-regulated and 3 down-regulated genes were found in cold-tolerant line ETH-DH7. Furthermore, real-time PCR was used to confirm the expression patterns of ZmPPs.

Conclusions

Our results provide new insights into the phylogenetic relationships and characteristic functions of maize PPs and will be useful in studies aimed at revealing the global regulatory network in maize abiotic stress responses, thereby contributing to the maize molecular breeding with enhanced quality traits.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-773) contains supplementary material, which is available to authorized users.     

玉米苗期SR蛋白基因家族的干旱胁迫应答

基因表达的选择性剪接(Alternative splicing, AS)调控与植物对逆境胁迫应答密切相关, SR蛋白(Serine/ arginine-rich proteins)是其中关键的调节因子。文章对玉米B73参考基因组进行分析显示: 多数SR蛋白家族基因成员启动子区域含有3~8种与发育或胁迫相关的顺式调控元件; 27个基因成员编码碱性蛋白, 其中23个成员的编码蛋白依照其N′端的首个RRM(RNA recognition motif)结构域特征大体上可划分为5个亚组。利用双向分级聚类方法, 对三叶期干旱胁迫下玉米杂交种郑单958及其亲本郑58和昌7-2的SR蛋白基因家族的分析显示, 该基因家族的表达模式具有明显的组织表达特异性和基因型依赖性特征; 其中在干旱胁迫下地下组织以下调表达模式为主, 而地上组织中以上调表达模式为主。在重度干旱胁迫后的3个不同时段复水过程中, 地上和地下组织中SR蛋白基因家族的表达皆以下调表达模式为主。另外, 尽管不同基因成员的表达模式在干旱胁迫及其后的复水过程中存在明显差异, 但普遍存在自身选择性剪接现象。SR蛋白基因家族在玉米干旱胁迫的应答规律, 为从AS-network视角解析玉米的抗逆分子机制提供了新思路。     

System analysis of microRNAs in the development and aluminium stress responses of the maize root system

MicroRNAs (miRNAs) are a class of regulatory small RNAs (sRNAs) that down‐regulate target genes through mRNA cleavage or translational inhibition. miRNA is known to play an important role in the root development and environmental responses in both the Arabidopsis and rice. However, little information is available to form a complete view of miRNAs in the development of the maize root system and Al stress responses in maize. Four sRNA libraries were generated and sequenced from the early developmental stage of primary roots (PRY), the later developmental stage of maize primary roots (PRO), seminal roots (SR) and crown roots (CR). Through integrative analysis, we identified 278 miRNAs (246 conserved and 32 novel ones) and found that the expression patterns of miRNAs differed dramatically in different maize roots. The potential targets of the identified conserved and novel miRNAs were also predicted. In addition, our data showed that CR is more resistant to Al stress compared with PR and SR, and the differentially expressed miRNAs are likely to play significant roles in different roots in response to environmental stress such as Al stress. Here, we demonstrate that the expression patterns of miRNAs are highly diversified in different maize roots. The differentially expressed miRNAs are correlated with both the development and environmental responses in the maize root. This study not only improves our knowledge about the roles of miRNAs in maize root development but also reveals the potential role of miRNAs in the environmental responses of different maize roots.     

Transcriptomic profiling of maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.) seedlings in response to <Emphasis Type="Italic">Pseudomonas putida</Emphasis> stain FBKV2 inoculation under drought stress

Several mechanisms have been proposed for plant growth-promoting rhizobacteria (PGPR)-mediated drought stress tolerance in plants, but little is known about the molecular pathways involved in the drought tolerance promoted by PGPR. We, therefore, aim to study the differential gene response between Pseudomonas putida strain FBKV2 and maize interaction under drought stress using Illumina sequencing. RNA Seq libraries were generated from leaf tissue of maize seedlings with and without strain FBKV2 subjected to drought stress. The libraries were mapped with maize genome database for the identification of differentially expressed genes (DEGs). The expression studies confirmed the downregulation of ethylene biosynthesis (ET), abscisic acid (ABA) and auxin signaling, superoxide dismutase, catalase, and peroxidase in FBKV2-inoculated seedlings. On the other hand, genes involved in β-alanine and choline biosynthesis, heat shock proteins, and late embryogenesis abundant (LEA) proteins were upregulated, which could act as key elements in the drought tolerance conferred by P. putida strain FBKV2. Another remarkable expression was observed in genes encoding benzoxazinoid (BX) biosynthesis which act as the chemoattractant, which was further confirmed by gfp-labeled P. putida strain FBKV2 root colonization studies. Overall, these results indicate that secretion of BXs attracted P. putida strain FBKV2 resulted in root colonization and mediated drought tolerance by modulating metabolic, signaling, and stress-responsive genes.     

GsZFP1, a new Cys2/His2-type zinc-finger protein, is a positive regulator of plant tolerance to cold and drought stress

Plant acclimation to environmental stress is controlled by a complex network of regulatory genes that compose distinct stress-response regulons. The C2H2-type zinc-finger proteins (ZFPs) have been implicated in different cellular processes involved in plant development and stress responses. Through microarray analysis, an alkaline (NaHCO(3))-responsive ZFP gene GsZFP1 was identified and subsequently cloned from Glyycine soja. GsZFP1 encodes a 35.14?kDa protein with one C2H2-type zinc-finger motif. The QALGGH domain, conserved in most plant C2H2-type ZFPs, is absent in the GsZFP1 protein sequence. A subcellular localization study using a GFP fusion protein indicated that GsZFP1 is localized to the nucleus. Real-time RT-PCR analysis showed that GsZFP1 was induced in the leaf by ABA (100?μM), salt (200?mM NaCl), and cold (4°C), and in the root by ABA (100?μM), cold (4°C), and drought (30% PEG 6000). Over-expression of GsZFP1 in transgenic Arabidopsis resulted in a greater tolerance to cold and drought stress, a decreased water loss rate, and an increase in proline irrespective of environmental conditions. The over-expression of GsZFP1 also increased the expression of a number of stress-response marker genes, including CBF1, CBF2, CBF3, NCED3, COR47, and RD29A in response to cold stress and RAB18, NCED3, P5CS, RD22, and RD29A in response to drought stress, especially early during stress treatments. Our studies suggest that GsZFP1 plays a crucial role in the plant response to cold and drought stress.     

Photosynthetic CO<Subscript>2</Subscript>/H<Subscript>2</Subscript>O gas exchange and dynamics of carbohydrates content in maize leaves under drought

We studied the temporal sequence of changes in the photosynthetic CO₂/H₂O gas exchange intensity, as well as leaf water status, contents of soluble carbohydrates, starch, proline, pigments, and MDA, in maize seedlings (Zea mays L., cv. Luchistaya) under adaptation to increasing water deficit. The duration of drought was 2, 3, 5, and 6 days. Withholding water from maize plants caused gradual increase in the intensity of water deficit: from mild (2 or 3 days) to moderate (5 days) and nearly severe (6 days) water stress. After 6 days, relative leaf water content decreased by 19.8% as compared to the control. On the second day after the onset of drought, slight reduction in the photosynthetic CO₂/H₂O gas exchange intensity of the treated plants was observed. After 6 days, photosynthesis and transpiration of leaves synchronously reduced almost threefold due to stomatal closure. The progressive soil drought had substantial impact on the carbohydrate metabolism. After 2 days of water deficit, the content of reducing sugars and sucrose increased slightly, whereas after 6 days, it increased ten and four times, respectively. After 2, 3, and 5 days of drought, the starch content declined slightly; however, under severe drought (6 days), it increased by 30% as compared to the control. Simultaneously with the increase in the content of soluble sugars, proline content increased significantly and it was the highest on the sixth day of drought. At all stages of water deficit, the proline content increased more significantly than the content of reducing carbohydrates and sucrose. Under increasing water deficit (5 and 6 days), the content of MDA was found to rise. At the initial drought stage (2 or 3 days) and under severe water deficit (6 days), no significant changes in the pigment content were observed. Thus, at the initial stages of progressive drought, in the leaves of this maize cultivar, a decline in photosynthetic activity proceeded simultaneously with accumulation of reducing sugars, sucrose, and proline. The results obtained showed that, at the first stages of adaptation of maize seedlings to drought, the changes in carbohydrate and proline metabolism have been observed, which have increased upon further plant dehydration.     

Proteomic approach: Identification of <Emphasis Type="Italic">Medicago truncatula</Emphasis> proteins induced in roots after infection with the pathogenic oomycete <Emphasis Type="Italic">Aphanomyces euteiches</Emphasis>

The legume root rot disease caused by the oomycete pathogen Aphanomyces euteiches is one major yield reducing factor in legume crop production. A comparative proteomic approach was carried out in order to identify proteins of the model legume Medicago truncatula which are regulated after an infection with A. euteiches. Several proteins were identified by two dimensional gel electrophoresis to be differentially expressed after pathogen challenge. Densitometric evaluation of expression values showed different regulation during the time-course analysed. Proteins regulated during the infection were identified by matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS). Among the differentially expressed proteins, two encoded putative cell wall proteins and two were designated as small heat shock proteins. Furthermore, an isoform of the chalcone-O-methyltransferase was found to be increased in infected roots. The majority of induced proteins belonged to the family of class 10 of pathogenesis related proteins (PR10). Previously, various PR10-like proteins have been shown to be regulated by general stress or abscisic acid (ABA). Therefore, these proteins were further investigated concerning their regulation in response to drought stress and exogenous ABA-application. Complex regulation patterns were identified: three of the A. euteiches-induced PR10-like proteins were also induced by exogenous ABA- but none of them is induced after drought stress. In contrast, three of these proteins are down-regulated by drought stress. Hence, the strong expression of different PR10-family members and their regulation profiles indicates that this set of proteins plays a major role during root adaptations to various stress conditions.     

水稻病程相关蛋白质在逆境胁迫下的表达研究

植物病程相关(PR)基因一般在病原物侵染过程中受诱导发生转录上调．目前有证据提示植物PR基因在非生物逆境胁迫下也发生转录变化,但其蛋白质的表达变化情况还鲜有报道．为了解水稻PR蛋白质在逆境胁迫下的表达特征,本文采用免疫印迹技术(Western blotting,WB)调查了8个PR蛋白质在冷、热、旱、淹和盐等5种胁迫下的表达谱．结果表明：在冷胁迫下PR8表达上调,在热胁迫下PR1a、PR3、PR5和PR16表达下调;在旱胁迫下PR1a、PR2和PR8表达上调,而PR5 和PR16表达下调,在淹胁迫下PR1、PR2和PR15表达上调,PR1a、PR3、PR5和PR8表达下调;在盐胁迫下PR2和PR3表达上调,而PR1a、PR5、PR8和PR16表达下调．另外,对这些PR 基因的上游启动子区进行分析,发现存在与胁迫响应相关的调控元件,其中脱落酸反应元件(ABRE)、TC-rich repeats和HSE的出现频率较高．这些蛋白质表达数据进一步佐证了PR蛋白在逆境胁迫反应中发挥着重要且不尽相同的作用．