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      Distinct expression patterns of two Arabidopsis phytocystatin genes, AtCYS1 and AtCYS2, during development and abiotic stresses

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          Abstract

          The phytocystatins of plants are members of the cystatin superfamily of proteins, which are potent inhibitors of cysteine proteases. The Arabidopsis genome encodes seven phytocystatin isoforms (AtCYSs) in two distantly related AtCYS gene clusters. We selected AtCYS1 and AtCYS2 as representatives for each cluster and then generated transgenic plants expressing the GUS reporter gene under the control of each gene promoter. These plants were used to examine AtCYS expression at various stages of plant development and in response to abiotic stresses. Histochemical analysis of AtCYS1 promoter- and AtCYS2 promoter-GUS transgenic plants revealed that these genes have similar but distinct spatial and temporal expression patterns during normal development. In particular, AtCYS1 was preferentially expressed in the vascular tissue of all organs, whereas AtCYS2 was expressed in trichomes and guard cells in young leaves, caps of roots, and in connecting regions of the immature anthers and filaments and the style and stigma in flowers. In addition, each AtCYS gene has a unique expression profile during abiotic stresses. High temperature and wounding stress enhanced the expression of both AtCYS1 and AtCYS2, but the temporal and spatial patterns of induction differed. From these data, we propose that these two AtCYS genes play important, but distinct, roles in plant development and stress responses.

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          Most cited references45

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          ABFs, a family of ABA-responsive element binding factors.

          Abscisic acid (ABA) plays an important role in environmental stress responses of higher plants during vegetative growth. One of the ABA-mediated responses is the induced expression of a large number of genes, which is mediated by cis-regulatory elements known as abscisic acid-responsive elements (ABREs). Although a number of ABRE binding transcription factors have been known, they are not specifically from vegetative tissues under induced conditions. Considering the tissue specificity of ABA signaling pathways, factors mediating ABA-dependent stress responses during vegetative growth phase may thus have been unidentified so far. Here, we report a family of ABRE binding factors isolated from young Arabidopsis plants under stress conditions. The factors, isolated by a yeast one-hybrid system using a prototypical ABRE and named as ABFs (ABRE binding factors) belong to a distinct subfamily of bZIP proteins. Binding site selection assay performed with one ABF showed that its preferred binding site is the strong ABRE, CACGTGGC. ABFs can transactivate an ABRE-containing reporter gene in yeast. Expression of ABFs is induced by ABA and various stress treatments, whereas their induction patterns are different from one another. Thus, a new family of ABRE binding factors indeed exists that have the potential to activate a large number of ABA/stress-responsive genes in Arabidopsis.
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            Protection against heat stress-induced oxidative damage in Arabidopsis involves calcium, abscisic acid, ethylene, and salicylic acid.

            Plants, in common with all organisms, have evolved mechanisms to cope with the problems caused by high temperatures. We examined specifically the involvement of calcium, abscisic acid (ABA), ethylene, and salicylic acid (SA) in the protection against heat-induced oxidative damage in Arabidopsis. Heat caused increased thiobarbituric acid reactive substance levels (an indicator of oxidative damage to membranes) and reduced survival. Both effects required light and were reduced in plants that had acquired thermotolerance through a mild heat pretreatment. Calcium channel blockers and calmodulin inhibitors increased these effects of heating and added calcium reversed them, implying that protection against heat-induced oxidative damage in Arabidopsis requires calcium and calmodulin. Similar to calcium, SA, 1-aminocyclopropane-1-carboxylic acid (a precursor to ethylene), and ABA added to plants protected them from heat-induced oxidative damage. In addition, the ethylene-insensitive mutant etr-1, the ABA-insensitive mutant abi-1, and a transgenic line expressing nahG (consequently inhibited in SA production) showed increased susceptibility to heat. These data suggest that protection against heat-induced oxidative damage in Arabidopsis also involves ethylene, ABA, and SA. Real time measurements of cytosolic calcium levels during heating in Arabidopsis detected no increases in response to heat per se, but showed transient elevations in response to recovery from heating. The magnitude of these calcium peaks was greater in thermotolerant plants, implying that these calcium signals might play a role in mediating the effects of acquired thermotolerance. Calcium channel blockers and calmodulin inhibitors added solely during the recovery phase suggest that this role for calcium is in protecting against oxidative damage specifically during/after recovery.
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              The cystatins: protein inhibitors of cysteine proteinases.

              The last decade has witnessed enormous progress of protein inhibitors of cysteine proteinases concerning their structures, functions and evolutionary relationships. Although they differ in their molecular properties and biological distribution, they are structurally related proteins. All three inhibitory families, the stefins, the cystatins and the kininogens, are members of the same superfamily. Recently determined crystal structures of chicken cystatin and human stefin B established a new mechanism of interaction between cysteine proteinases and their inhibitors which is fundamentally different from the standard mechanism for serine proteinases and their inhibitors.
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                Author and article information

                Contributors
                colim@gnu.ac.kr
                Journal
                Plant Cell Rep
                Plant Cell Reports
                Springer-Verlag (Berlin/Heidelberg )
                0721-7714
                1432-203X
                5 June 2010
                5 June 2010
                August 2010
                : 29
                : 8
                : 905-915
                Affiliations
                [1 ]Division of Applied Life Science (BK21 Program), Environmental Biotechnology National Core Research Center and PMBBRC, Graduate School of Gyeongsang National University, Jinju, 660-701 Korea
                [2 ]National Academy of Agricultural Science, Rural Development Administration, Suwon, 441-707 Korea
                [3 ]Department of Food and Nutrition, Silla University, Pusan, 617-736 Korea
                Author notes

                Communicated by R. Schmidt.

                Article
                876
                10.1007/s00299-010-0876-y
                2903682
                20526604
                0d5fd15c-02b2-4cc7-be9e-9cbb67b283a2
                © The Author(s) 2010
                History
                : 14 December 2009
                : 9 May 2010
                : 19 May 2010
                Categories
                Original Paper
                Custom metadata
                © Springer-Verlag 2010

                Plant science & Botany
                gene expression,promoter,abiotic stress,gus staining,cysteine protease inhibitor,transgenic plant

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