Aquaporin （AQP） belongs to a highly conserved group of membrane proteins considered as major intrinsic proteins, which facilitate water transport across biological membranes. The discovery of AQPs in plants has resu...Aquaporin （AQP） belongs to a highly conserved group of membrane proteins considered as major intrinsic proteins, which facilitate water transport across biological membranes. The discovery of AQPs in plants has resulted in a paradigm shift in the understanding of plant-water relations, however, the potential relationship between the role of aquaporins in regulating plant water balance and drought tolerance still remains elusive. In this study, the gene encoding potato AQP cDNA, StPIP1 （GenBank accession no. DQ999080）, was cloned from the leaf of potato cultivar Gannongshu 2 by reverse transcription-PCR （RT-PCR）. Sequence alignment was made by BLASTn in GenBank, the phylogenetic analysis was conducted using PHYLIPWY, the 3D structure was predicted in Swiss-Model server. Subcellular localization of StPIP1 was performed by constructing CaMV35S-StPIP1-GFP and rd29A-StPIP1-GFP fusion proteins and transient expression in onion epidermis. To understand StPIP1 physiological functions in potato under various stress conditions, the StPIP1 gene in a reverse orientation was transformed into tobacco driven by the Cauliflower mosaic virus （CMV） 35S promoter. The expression levels of transgenic and wild-type plants were assessed under various abiotic stress conditions using semi-quantitative RT-PCR, and the morphological and physiological responses of transgenic plants to different stress conditions were investigated. The expression of StPIP1 mRNA decreased in transgenic plants under non-stress and stress conditions, however, the reduction was more severer under drought stress. In both non-stress and stress conditions, StPIP1 was expressed predominantly in root. The morphological and physiological investigation showed no significant differences in growth rate, germination rate, and root fresh weight （FW） between transgenic and wild-type plants when grown under favorable conditions. In contrast, under drought stress, the reduction in StPIPI expression leads to a delay in seed germination and seedling growth, accelerated seedling wilt, and leaf morphological abnormity. Under ＂enough＂ water conditions （i.e., water culture）, the aerial parts of anti-sense plants showed no differences. However, for the aerial parts to accumulate the same amount of biomass, transgenic plants needed about 3 times more abundant root system to transport water for plant growth than wild-type plants. Morphological investigation showed that the reduction in StPIP1 expression increased the root system in transgenic plants under drought stress. As a result, the increase of root mass might compensate the reduced cellular water permeability in order to ensure a sufficient water supply for the plant. Results demonstrated that StPIP1 plays an important role for water transportation in potato, especially under drought stress conditions. The reduced expression of StPIP1 decreases the cellular water transport and influences the expression of endogenous AQPs genes and thereby, has impacts on seed germination, seedling growth, and stress responses of potato to drought conditions.展开更多
基金supported by the National 973 Program of China (2006CB708200)Gansu Province Key Technologies R&D Program (2GS054-A41-00501),Chinathe President Youth Fund of Academy of Agri-Sciences Anhui Province, China (200933)
文摘Aquaporin （AQP） belongs to a highly conserved group of membrane proteins considered as major intrinsic proteins, which facilitate water transport across biological membranes. The discovery of AQPs in plants has resulted in a paradigm shift in the understanding of plant-water relations, however, the potential relationship between the role of aquaporins in regulating plant water balance and drought tolerance still remains elusive. In this study, the gene encoding potato AQP cDNA, StPIP1 （GenBank accession no. DQ999080）, was cloned from the leaf of potato cultivar Gannongshu 2 by reverse transcription-PCR （RT-PCR）. Sequence alignment was made by BLASTn in GenBank, the phylogenetic analysis was conducted using PHYLIPWY, the 3D structure was predicted in Swiss-Model server. Subcellular localization of StPIP1 was performed by constructing CaMV35S-StPIP1-GFP and rd29A-StPIP1-GFP fusion proteins and transient expression in onion epidermis. To understand StPIP1 physiological functions in potato under various stress conditions, the StPIP1 gene in a reverse orientation was transformed into tobacco driven by the Cauliflower mosaic virus （CMV） 35S promoter. The expression levels of transgenic and wild-type plants were assessed under various abiotic stress conditions using semi-quantitative RT-PCR, and the morphological and physiological responses of transgenic plants to different stress conditions were investigated. The expression of StPIP1 mRNA decreased in transgenic plants under non-stress and stress conditions, however, the reduction was more severer under drought stress. In both non-stress and stress conditions, StPIP1 was expressed predominantly in root. The morphological and physiological investigation showed no significant differences in growth rate, germination rate, and root fresh weight （FW） between transgenic and wild-type plants when grown under favorable conditions. In contrast, under drought stress, the reduction in StPIPI expression leads to a delay in seed germination and seedling growth, accelerated seedling wilt, and leaf morphological abnormity. Under ＂enough＂ water conditions （i.e., water culture）, the aerial parts of anti-sense plants showed no differences. However, for the aerial parts to accumulate the same amount of biomass, transgenic plants needed about 3 times more abundant root system to transport water for plant growth than wild-type plants. Morphological investigation showed that the reduction in StPIP1 expression increased the root system in transgenic plants under drought stress. As a result, the increase of root mass might compensate the reduced cellular water permeability in order to ensure a sufficient water supply for the plant. Results demonstrated that StPIP1 plays an important role for water transportation in potato, especially under drought stress conditions. The reduced expression of StPIP1 decreases the cellular water transport and influences the expression of endogenous AQPs genes and thereby, has impacts on seed germination, seedling growth, and stress responses of potato to drought conditions.
