Genome Wide Identification of C2H2-Type Zinc Finger Proteins of Tomato and Expression Analysis Under Different Abiotic Stresses

In plants, C2H2-type zinc finger proteins play important roles in multiple processes, including plant growth and development, as well as biotic and abiotic responses. In the present study, based on the presence of the C2H2 domain (CX2~4CX3FX5LX2HX3~5H), 112 C2H2-type zinc finger proteins were predicted in tomato. Through gene and protein structures analyses and phylogenetic analysis, the 112 C2H2-type zinc finger proteins were divided into five subfamilies. Members of the same subfamily shared similarities in gene and protein structures, while members of different subfamilies contained different numbers of the C2H2 domain. The tissue expression pattern analysis showed that 24 C2H2-type zinc finger proteins are constitutively expressed in all tissues, indicating that they may play important roles in the growth and development of all tissues. In addition, under chilling (4 °C), heat (42 °C), high salinity (200 Mm NaCl), and osmotic (20% PEG) stresses, members of C2H2-type zinc finger family were induced to varying degrees, which suggested that these genes were involved in multiple abiotic stress responses. This study will provide theoretical basis for further research of C2H2-type zinc finger proteins in tomato.

     

Arabidopsis SAP5 functions as a positive regulator of stress responses and exhibits E3 ubiquitin ligase activity

AtSAP5, one of approximately 14 members of the Stress Associated Protein gene family in Arabidopsis, was identified by its expression in response to salinity, osmotic, drought and cold stress. AtSAP5 shows strong homology to OSISAP1, an A20/AN1-type zinc finger protein implicated in stress tolerance in rice. To evaluate the function of AtSAP5 in the regulation of abiotic stress responses, transgenic Arabidopsis plants that over-express AtSAP5 (35S::AtSAP5) were characterized, along with wild-type and T-DNA knock-down plants. Plants that over-express AtSAP5 showed increased tolerance to environmental challenges including salt stress, osmotic stress and water deficit. Comparison of gene expression patterns between 35S::AtSAP5 transgenic plants and wild-type plants under normal conditions and water deficit stress indicated that over-expression of AtSAP5 correlates with up-regulation of drought stress responsive gene expression. Analysis of transgenic plants that express GFP-AtSAP5 showed that it is localized primarily in nuclei of root cells and recombinant AtSAP5 has E3 ubiquitin ligase activity in vitro. These results indicate that AtSAP5 has E3 ligase activity and acts as a positive regulator of stress responses in Arabidopsis.     

盐度胁迫下缢蛏渗透压变化及 V-ATPase H基因的表达分析

为研究V-ATPase H基因在缢蛏（Sinonovacula constricta）盐度胁迫中的功能,以缢蛏成体为实验材料,将缢蛏置于5、15、20、25、35盐度水体中进行胁迫实验,测定了不同胁迫时间缢蛏的血清渗透压、V-ATPase活性变化,克隆了V-ATPase H基因的开放阅读框（ORF）全长序列,并分析其mRNA表达特征。结果显示,低盐组（盐度5和盐度15）和高盐组（盐度25和盐度35）缢蛏血清渗透压变化明显,与对照组（盐度20）有极显著差异（P < 0.01）。随着时间的推移,实验组V-ATPase活力整体呈现先下降后上升的趋势,对照组（盐度20）无明显变化。V-ATPase H基因开放阅读框长度1 440 bp,编码479个氨基酸。qPCR结果显示,V-ATPase H基因在缢蛏鳃中的表达量极显著高于水管、外套膜、肾、肝胰腺、唇瓣、足6个组织（P < 0.01）;盐度胁迫下各个实验组V-ATPase H基因在鳃中的表达量持续上升,在24 h达到峰值,显著高于对照组（P < 0.05）。实验结果表明,缢蛏V-ATPase H基因在盐度适应过程中主要在低盐和高盐环境下起到维持自身血清渗透压与外界渗透压平衡的作用。     

Overexpression of dehydrin tas14 gene improves the osmotic stress imposed by drought and salinity in tomato

One strategy to increase the level of drought and salinity tolerance is the transfer of genes codifying different types of proteins functionally related to macromolecules protection, such as group 2 of late embryogenesis abundant (LEA) proteins or dehydrins. The TAS14 dehydrin was isolated and characterized in tomato and its expression was induced by osmotic stress (NaCl and mannitol) and abscisic acid (ABA) [Godoy et al., Plant Mol Biol 1994;26:1921-1934], yet its function in drought and salinity tolerance of tomato remains elusive. In this study, transgenic tomato plants overexpressing tas14 gene under the control of the 35SCaMV promoter were generated to assess the function of tas14 gene in drought and salinity tolerance. The plants overexpressing tas14 gene achieved improved long-term drought and salinity tolerance without affecting plant growth under non-stress conditions. A mechanism of osmotic stress tolerance via osmotic potential reduction and solutes accumulation, such as sugars and K(+) is operating in tas14 overexpressing plants in drought conditions. A similar mechanism of osmotic stress tolerance was observed under salinity. Moreover, the overexpression of tas14 gene increased Na(+) accumulation only in adult leaves, whereas in young leaves, the accumulated solutes were K(+) and sugars, suggesting that plants overexpressing tas14 gene are able to distribute the Na(+) accumulation between young and adult leaves over a prolonged period in stressful conditions. Measurement of ABA showed that the action mechanism of tas14 gene is associated with an earlier and greater accumulation of ABA in leaves during short-term periods. A good feature for the application of this gene in improving drought and salt stress tolerance is the fact that its constitutive expression does not affect plant growth under non-stress conditions, and tolerance induced by overexpression of tas14 gene was observed at the different stress degrees applied to the long term.     

Rapid accumulation of phosphatidylinositol 4,5-bisphosphate and inositol 1,4,5-trisphosphate correlates with calcium mobilization in salt-stressed arabidopsis

The phosphoinositide phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P(2)] is a key signaling molecule in animal cells. It can be hydrolyzed to release 1,2-diacyglycerol and inositol 1,4,5-trisphosphate (IP(3)), which in animal cells lead to protein kinase C activation and cellular calcium mobilization, respectively. In addition to its critical roles in constitutive and regulated secretion of proteins, PtdIns(4,5)P(2) binds to proteins that modify cytoskeletal architecture and phospholipid constituents. Herein, we report that Arabidopsis plants grown in liquid media rapidly increase PtdIns(4,5)P(2) synthesis in response to treatment with sodium chloride, potassium chloride, and sorbitol. These results demonstrate that when challenged with salinity and osmotic stress, terrestrial plants respond differently than algae, yeasts, and animal cells that accumulate different species of phosphoinositides. We also show data demonstrating that whole-plant IP(3) levels increase significantly within 1 min of stress initiation, and that IP(3) levels continue to increase for more than 30 min during stress application. Furthermore, using the calcium indicators Fura-2 and Fluo-3 we show that root intracellular calcium concentrations increase in response to stress treatments. Taken together, these results suggest that in response to salt and osmotic stress, Arabidopsis uses a signaling pathway in which a small but significant portion of PtdIns(4,5)P(2) is hydrolyzed to IP(3). The accumulation of IP(3) occurs during a time frame similar to that observed for stress-induced calcium mobilization. These data also suggest that the majority of the PtdIns(4,5)P(2) synthesized in response to salt and osmotic stress may be utilized for cellular signaling events distinct from the canonical IP(3) signaling pathway.     

Salicylic acid treatment via the rooting medium interferes with stomatal response, CO2 fixation rate and carbohydrate metabolism in tomato, and decreases harmful effects of subsequent salt stress

Salicylic acid (SA) applied at 10(-3) m in hydroponic culture decreased stomatal conductance (g(s)), maximal CO(2) fixation rate (A(max) ) and initial slopes of the CO(2) (A/C(i)) and light response (A/PPFD) curves, carboxylation efficiency of Rubisco (CE) and photosynthetic quantum efficiency (Q), resulting in the death of tomato plants. However, plants could acclimate to lower concentrations of SA (10(-7) -10(-4) m) and, after 3 weeks, returned to control levels of g(s), photosynthetic performance and soluble sugar content. In response to high salinity (100 mm NaCl), the pre-treated plants exhibited higher A(max) as a function of internal CO(2) concentration (C(i) ) or photosynthetic photon flux density (PPFD), and higher CE and Q values than salt-treated controls, suggesting more effective photosynthesis after SA treatment. Growth in 10(-7) or 10(-4) m SA-containing solution led to accumulation of soluble sugars in both leaf and root tissues, which remained higher in both plant parts during salt stress at 10(-4) m SA. The activity of hexokinase (HXK) with glucose, but not fructose, as substrate was reduced by SA treatment in leaf and root samples, leading to accumulation of glucose and fructose in leaf tissues. HXK activity decreased further under high salinity in both plant organs. The accumulation of soluble sugars and sucrose in roots of plants growing in the presence of 10(-4) m SA contributed to osmotic adjustment and improved tolerance to subsequent salt stress. Apart from its putative role in delaying senescence, decreased HXK activity may divert hexoses from catabolic reactions to osmotic adaptation.