Aspergillus glaucus Aquaglyceroporin Gene glpF Confers High Osmosis Tolerance in Heterologous Organisms
Aspergillus glaucus Aquaglyceroporin Gene glpF Confers High Osmosis Tolerance in Heterologous Organisms
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青曲霉水甘油孔蛋白基因 glpF 赋予异源生物高渗透耐受性
DOI：
10.1128/aem.02127-15
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发表时间：
2015-10-01
影响因子：
4.4
通讯作者：
Zhang，Shi-Hong
中科院分区：
文献类型：
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作者：
Liu，Xiao-Dan;Wei，Yi;Zhang，Shi-Hong
水甘油孔蛋白（GlpF）是一种与水和其他不带电溶质沿着转运甘油的蛋白质，参与了许多物种的代谢调节。真菌种类构成了一大类真核生物，它们的GlpF可能是多样的，表现出各种活性。然而，很少有丝状真菌GlpF的生物学研究。在这里，glpF基因从嗜盐真菌灰绿曲霉（AgglpF）被证实是一个通道的水或甘油在非洲爪蟾卵母细胞，并进一步在三个异源系统的功能分析。在酿酒酵母中，过表达AgglpF的细胞对干旱、盐和某些金属离子具有显著的耐受性。然后在粗糙脉孢菌的丝状真菌中表征AgglpF。基于N.本研究构建了一系列含有Crassa水通道蛋白基因（NcAQP）缺失突变体（Δaqp突变体）、NcAQP和AgglpF互补突变体以及三个缺失天冬酰胺-脯氨酸-丙氨酸基因（NPA）片段的AgglpF互补突变体。耐盐性分析表明，AgglpF互补菌株具有最高的耐盐性。此外，在AgglpF互补菌株的胞内甘油含量显着高于其他菌株。AgGlpF-绿色荧光蛋白（GFP）融合蛋白定位于洋葱表皮细胞的质膜上，表明AgGlpF在植物中具有功能。事实上，当AgglpF在拟南芥中表达时，转基因株系在高渗透胁迫条件下，特别是在干旱胁迫条件下存活。总的来说，我们的研究结果表明，AgGlpF作为一个水/甘油转运蛋白是必需的真菌和植物在高渗透胁迫条件下的生存，并可能在基因工程中产生高盐和抗旱性的应用价值。
ABSTRACT Aquaglyceroporins (GlpFs) that transport glycerol along with water and other uncharged solutes are involved in osmoregulation in myriad species. Fungal species form a large group of eukaryotic organisms, and their GlpFs may be diverse, exhibiting various activities. However, few filamentous fungal GlpFs have been biologically investigated. Here, a glpF gene from the halophilic fungus Aspergillus glaucus (AgglpF) was verified to be a channel of water or glycerol in Xenopus laevis oocytes and was further functionally analyzed in three heterologous systems. In Saccharomyces cerevisiae, cells overexpressing AgglpF possessed significant tolerance of drought, salt, and certain metal ions. AgglpF was then characterized in the filamentous fungus of Neurospora crassa. Based on the N. crassa aquaporin gene (NcAQP) disruption mutant (the Δaqp mutant), a series of complementary strains carrying NcAQP and AgglpF and three asparagine-proline-alanine-gene (NPA)-deleted AgglpF fragments were created. As revealed by salt resistance analysis, the AgglpF complementary strain possessed the highest salt resistance among the tested strains. In addition, the intracellular glycerol content in the AgglpF complementary strain was markedly higher than that in the other strains. The AgGlpF-green fluorescent protein (GFP) fusion protein was subcellularly localized in the plasma membrane of onion epidermal cells, suggesting that AgglpF functions in plants. Indeed, when AgglpF was expressed in Arabidopsis thaliana, transgenic lines survived under conditions of high osmotic stress and under conditions of drought stress in particular. Overall, our results revealed that AgGlpF as a water/glycerol transporter is required for survival of both fungi and plants under conditions of high osmotic stress and may have value in applications in genetic engineering for generating high salt and drought resistance.