铝胁迫对拟南芥根尖ＡｔＰＩＮ２ 蛋白表达活性的影响

Effects of aluminum stress on the activity of PIN2 protein in Arabidopsis thaliana apical roots

关键词:  铝胁迫, 拟南芥, AtPIN2, 表达活性

Key words:  aluminum stress, Arabidopsis thaliana, AtPIN2, expression activity

摘要:

铝胁迫能影响根尖生长素的运输,这与生长素运输载体密切相关,PIN2作为根尖生长素的运输蛋白,其独特的组织定位可能诱导PIN2蛋白参与了铝调节生长素的运输过程。该研究以拟南芥PIN2缺失突变体(pin2)、PIN2□∷□GFP融合体及其野生型(WT)为材料,应用激光扫描共聚焦显微技术,研究铝处理对拟南芥根尖生长素运输蛋白PIN2的表达活性、蛋白在组织及亚细胞水平分布及其对铝内置化作用的影响。结果表明:短期铝处理或低铝浓度能明显增加拟南芥根尖细胞PIN2蛋白表达活性,而长期铝处理或高铝浓度抑制其表达活性; 以100 μmol·L^-1 AlCl₃处理4 h的蛋白表达活性最高。蛋白印迹反应发现,铝处理促进PIN2蛋白在细胞膜上累积,减少胞内囊泡中PIN2蛋白的含量; 囊泡运输抑制剂(BFA)能抑制铝诱导PIN2蛋白的分配。铝胁迫增加拟南芥根尖细胞H₂O₂累积,pin2的H₂O₂累积量大于WT,而相对根长小于WT。Morin染色结果显示,pin2的铝内置化显著小于WT。上述研究表明,PIN2蛋白在100 μmol·L^-1 AlCl₃处理条件下活性最高,细胞膜累积程度加强,铝内置化能力增强,从而调节根系的生长发育。该研究结果进一步为铝抑制生长素的运输机制提供了理论基础。

Abstract:

There is closely relationship between Al-influenced auxin transport and auxin transporter. PIN2 is an auxin efflux carrier protein, it unique localization may responses to Al-influenced auxin transport. In this study, using pin2, PIN2□∷□GFP and WT as experimental materials, the effects of Al stress on PIN2 protein activity,distribution and Al internalization were investigated with confocal laser scanning microscope. The results indicated that short-term Al treatment or low Al level increased the activity of PIN2 protein in apical cells of Arabidopsis seedlings, while long-term Al treatment or high Al levels inhibited it. The activity of PIN2 protein reached the maximal value in response to 100 μmol·L^-1 Al for 4 h. The results from Western-blotting analysis indicated that Al enhanced the accumulation of PIN2 protein on cell membrane, while decreased it in vesicles. Brefeldin A(BFA),an inhibitor of vesicle transport suppressed Al-induced allocation of PIN2 protein. On the other hand, Al stress could increase the accumulation of H₂O₂ in apices. pin2 had a higher H₂O₂ accumulation and a lower relative root elongation than WT. And the results from Al-Morin staining analysis indicated that pin2 had a lower Al internalization than WT. Taken together, the above results suggested that 100 μmol·L^-1 Al induced the highest activity of PIN2 protein, and enhanced its accumulation in horizontal direction of plasma membrane and Al internalization,thus mediated root growth. The results would provide scientific basis for elucidating Al-influenced auxin transport.