Members of the “stress-associated protein” (SAP) gene family have been frequently implicated in the plant stress response. The genes' sequence most distinctive feature is the presence of two zinc finger domains (A20 and AN1), which have been long recognized as key elements in the animal immune system. The A20 domain was first identified in a human TNF-α inducible protein, and AN1 in an ubiquitin-like fusion protein in the Xenopus laevis egg and early embryo. The number of SAP genes present in plant genomes ranges from three in Chlamydomonas reinhardtii to 19 in Populus trichocarpa; two types of SAP gene product have been recognized: type I proteins harbor the motif Cx2Cx9-12Cx1-2Cx4Cx2Hx5HxC, while type II ones harbor the more expansive motif Cx4Cx9-12Cx1-2Cx4Cx2Hx5HxC (where “x” represents any amino acid).
The xerophytic grass species Leymus chinensis (Trin.) Tzvel. is adapted to alkaline-sodic soils in northern China. It therefore serves both as a useful model for understanding the basis of abiotic stress tolerance in the monocotyledons, and as a potential donor of tolerance genes to the cereals. Here, a SAP gene was isolated from a L. chinensis leaf and root cDNA library, and its inducibility by salinity stress was characterized. The gene was also heterologously expressed in brewers' yeast (Saccharomyces cerevisiae) with a view to testing its potential as a salinity tolerance enhancing transgenehttp://www.cusabio.com/.
After research, we get these findings bellow:
The full length LcSAP cDNA featured an 889 nt open reading frame, a 211 nt 5’-UTR and a 192 nt 3’-UTR. Its predicted product was a 161 residue protein of molecular mass 17.6 kDa and pI 8.56. The product, like other type I SAPs, harbored a A20 domain (Cx2-4Cx11Cx2C) at its N terminus and an AN1 domain (Cx2Cx9-12Cx1-2Cx4Cx2Hx5HxC) at its C terminus. The phylogenetic analysis grouped it with other plant SAPs (Fig. 1B). The sequence shared a high level of identity at the peptide level with that of a protein from Aeluropus littoralis, but its homology level with the other plant SAPs tested was no higher than 45.1%.
Under control conditions, the abundance of LcSAP transcript was clearly higher in the L. chinensis leaf than in its root. Other plant SAP genes have been found to also behave in this way. When exposed to 400 mM NaCl, the level of LcSAP transcription began to rise after 6 h, and had not fallen by the end of the sampling period (24 h). The effect of exposure to 100 mM Na 2 CO 3 was also to induce LcSAP transcription, but the extent of the induction lessened after 6 h. Salinity stress has been observed to up-regulate SAP genes in Arabidopsis thaliana, sorghum, tea, sugar cane, tomato, A. littoralis, rice, Medicago truncatula and Festuca arundinacea. However, in a few cases, it has been shown to down-regulate them: for example, MusaSAP1 in Suaeda salsa, SlSAP1/11 in tomato and ZFP177 in rice. It is presumed that this observed variation in the direction of salinity-induced regulation of the various SAP genes reflects the variety of roles which the genes play in the salinity response.
The successful transcription of the LcSAP transgene in yeast was confirmed by northern blotting. Under non-stressed conditions, the growth rate of the transgenic yeast cells was indistinguishable from that of wild type cells. However, in the presence of 1.4 M NaCl, the transgenic cells were better able to maintain their growth than were the wild type cells. An equivalent experience has been reported for a number of other SAP genes. Thus, for example, the constitutive expression of the rice AN/A20 gene OsiSAP1 succeeded in enhancing the tolerance of tobacco plants to a range of abiotic stresses, including salinity. Similarly, the over-expression of OsiSAP8 in rice and its constitutive expression in tobacco improved the plants' capacity to tolerate salinity, drought and low temperature stress. The constitutive expression of OsSAP11 in A. thaliana conferred tolerance to both salinity and moisture-deficit. When the rice gene ZFP177 was constitutively expressed in tobacco, the plants displayed increased tolerance to both high and low temperature and to peroxide, but became more sensitive to both dehydration and salinity. The over-expression of AtSAP5 led to an improved level of tolerance to both salinity and water deficiency. The expression of the A. littoralis gene AlSAP in both yeast and tobacco raised the tolerance of the host to both salinity, ionic and osmotic stress. Similarly, the heterologous expression of sorghum SbSAP14 in rice helped to protect the plants against the oxidative damage caused by salinity. Transgenic tobacco plants expressing the sugar cane gene ShSAP1 were found to be more tolerant to both salinity and drought than the wild type controls.
Finally, the constitutive expression in tobacco of the M. truncatula gene MtSAP1 conferred tolerance to various abiotic stresses. While SAPs are clearly involved in the plant response to salinity, the mechanistic basis of their contribution is not as yet known. A future research goal will be to identify the proteins which interact with LcSAP.
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