盐生植物海滨碱蓬Na+吸收和积累的研究

论文摘要

土壤中高浓度的盐分对大多数的高等植物,特别是农作物产生毒害效应,引起植物生长受抑、甚至死亡,对农牧业生产构成严重的威胁。由低亲和性吸收系统介导的毒性Na+的吸收是造成植物盐害的诱因,减少有害Na+进入植物体内是解决这一问题的关键。但目前国内外对在植物Na+吸收途径研究方面以拟南芥、水稻和小麦等盐敏感的甜土植物和盐芥等盐生植物为材料,它们均是叶中Na+积累量较低的拒盐植物,吸Na+能力十分有限,并不是研究Na+吸收途径的理想材料,因此,对高等植物低亲和性Na+吸收途径尚不明确。本研究以积盐型盐生植物海滨碱蓬（Suaeda maritima）为材料,利用22Na+同位素示踪技术,对海滨碱蓬Na+吸收与积累机制进行了系统分析,取得的主要结果如下:1.通过对Ca2+（非选择性阳离子通道与低亲和性阳离子转运蛋白抑制剂）、Li+（Na+吸收的竞争性抑制剂）以及TEA+、Cs+和Ba2+（K+通道抑制剂）等几种离子通道抑制剂影响海滨碱蓬22Na+内流的分析,发现无论是在轻度（25 mM NaCl）还是在重度（150 mM NaCl）盐生境下,Ca2+和Li+都没有影响Na+的吸收,表明目前被学术界广泛接受的非选择性阳离子通道（NSCCs）和低亲和性阳离子转运蛋白（LCT1）并非Na+进入海滨碱蓬根细胞的主要途径。鉴定出了两条低亲和性Na+吸收途径:途径1对Ba2+很敏感,对TEA+和Cs+不敏感,介导轻微盐生境（25 mM NaCl）下Na+的吸收,可能由HKT类转运蛋白完成;途径2对Ba2+、TEA+和Cs+都很敏感,介导重度盐生境（150 mM NaCl）下Na+的吸收,可能由AKT1类通道蛋白完成。2.在初步确定上述两条途径的外界NaCl浓度的拐点在75-100 mM NaCl的基础上,将其进一步细化在90-95 mM NaCl。我们进一步研究发现,K+影响Na+吸收的拐点也与TEA+影响Na+吸收的拐点一致。3.比较了TEA+和Ba2+对海滨碱蓬、小麦和水稻三种植物22Na+内流的影响,结果表明,在低盐生境下HKT类型的转运蛋白对小麦和水稻根Na+的吸收要远少于海滨碱蓬,且对小麦和水稻根的拒Na+有一定的贡献;在高盐生境下,HKT类型的转运蛋白在小麦和水稻中更多地行使了拒Na+的功能。4.分析了NH4+（5 mM）对TEA+和Ba2+抑制海滨碱蓬22Na+内流、Na+累积和Na+净吸收速率的影响,结果表明在加入Ba2+（5 mM）的条件下,同时加入NH4+来降低由H+-ATP酶建立的跨膜质子梯度显著地降低了海滨碱蓬根的拒Na+作用。据此,我们推测在海滨碱蓬根种存在至少两种HKT类转运蛋白:一种在低盐生境下行使Na+吸收的功能;另一种在低盐和高盐胁迫下行使拒Na+作用。另外,在高盐生境下ABC转运蛋白也可能是高等植物Na+外排泵蛋白的一个候选者。5.研究了阳离子-Cl-共转运蛋白（cation-Cl-cotransporter,CCC）抑制剂丁苯氧酸（bumetanide）对海滨碱蓬Na+累积和净吸收速率的影响,结果表明CCC转运蛋白在轻度盐生境下介导少量Na+的吸收,在高盐生境下介导大量Na+的吸收;另外,CCC转运蛋白也可能在调节高Na+和高K+环境下水分的传导中发挥作用。6.在高盐（100-200 mM NaCl）生境下,低浓度的K+（5 and 10 mM）促进了海滨碱蓬Na+的吸收,其可能的机制是激活了AKT1类型的K+通道。7.不同浓度NaCl（25、150和200 mM）处理192（8d）、240（10d）和360（10d）h时,海滨碱蓬根中Na+浓度均到达最高值（140、205和310 mM）,分别是外界生境中Na+浓度的5.6、1.4和1.5倍,而后随着处理时间的延长根中的Na+浓度逐渐下降;处理480 h（20 d）时,海滨碱蓬植株地上部Na+浓度均到达最高值（376、616和715mM）,分别是外界生境中Na+浓度的15、4.1和3.6倍;植株地上部Na+浓度在三种生境盐处理的6-12 h之间迅速增加,而后增加缓慢。根22Na+内流随时间进程的变化模式与根和植株地上部Na+浓度变化模式基本一致。以上结果同时也进一步表明,当盐处理达到一定阶段,植株中Na+的积累上升到一定水平且到达稳态平衡以后,即使在海滨碱蓬这样大量吸收Na+的盐生植物中也会产生拒Na+作用。8.不同浓度NaCl（25、150和200 mM）处N20 d后,植株地上部中K+浓度整体上分别显著下降了37%、64%和54%,根中的K+浓度变化起伏较大,整体上呈下降趋势;植株地上部中Na+/K+比分别增加至对照的47、156和147倍;根中的Na+/K+比分别显著增加为对照的41、38和47倍。

论文目录

• Acknowledgements
• List of abbreviation used
• 摘要
• Abstract
• CHAPTER 1:INTRODUCTION
• + UPTAKE AND Na⁺ TRANSPORT IN HIGHER PLANTS'>CHAPTER 2:PROGRESS ON Na⁺ UPTAKE AND Na⁺ TRANSPORT IN HIGHER PLANTS
• 2.1 Background
• 2.2 Negative effects of salinity on plant productivity
• 2.2.1 Osmotic stress
• 2.2.2 Ionic stress
• 2.2.3 Secondary stresses
• + uptake into roots'>2.3 Na⁺ uptake into roots
• + uptake in higher plants'>2.3.1 Physiological and electrophysiological mechanisms of Na⁺ uptake in higher plants
• +,Na⁺ competition and selectivity'>2.3.1.1 K⁺,Na⁺ competition and selectivity
• +-Ca²⁺interactions under salt environments'>2.3.1.2 Na⁺-Ca²⁺interactions under salt environments
• + uptake'>2.3.1.3 Effects of driving force or electrochemical potential across the plasma on Na⁺uptake
• + uptake'>2.3.1.4 Effects of external salt concentration on Na⁺uptake
• + uptake'>2.3.1.5 Experimental methods in studying the physiological mechanisms of Na⁺uptake
• + uptake in higher plants'>2.3.2 Molecular biological mechanism of Na⁺ uptake in higher plants
• 2.3.2.1 NSCCs/VICs
• 2.3.2.2 LCT1
• 2.3.2.3 KUP/HAK/KT
• 2.3.2.4 HKT
• 2.3.2.5 AKT1
• 2.3.2.6 CCC
• + Transport in higher plants'>2.4 Na⁺ Transport in higher plants
• + xylem loading'>2.4.1 Control of Na⁺ xylem loading
• + retrieval from the xylem'>2.4.2 Na⁺ retrieval from the xylem
• + recirculation in the phloem'>2.4.3 Na⁺ recirculation in the phloem
• + extrusion or effiux from the root'>2.4.4 Na⁺ extrusion or effiux from the root
• + into the vacuoles'>2.4.5 Intracellular compartmentation of Na⁺ into the vacuoles
• + excretion'>2.4.6 Na⁺excretion
• CHAPTER 3:MATERIAL AND METHODS
• 3.1 Plant material and growth condition
• 3.2 Treatments
• + and K⁺ concentration determination'>3.3 Growth measurements,and Na⁺ and K⁺ concentration determination
• 22Na⁺ influx experiments'>3.4²²Na⁺ influx experiments
• 3.5 Statistical analysis
• CHAPTER 4:RESULTS
• 22Na⁺ influx in Suaeda maritima'>4.1 The bases of root ²²Na⁺ influx in Suaeda maritima
• 22Na⁺ influx'>4.1.1 Time course of root ²²Na⁺influx
• + concentration on root ²²Na⁺ influx'>4.1.2 Effects of external Na⁺ concentration on root ²²Na⁺influx
• 22Na⁺ influx and root Na⁺ concentration'>4.1.3 The correlation between root ²²Na⁺ influx and root Na⁺concentration
• 22Na⁺ influx and fresh weight of the root'>4.1.4 The correlation between ²²Na⁺ influx and fresh weight of the root
• 22Na⁺ loss during washing and blotting'>4.1.5 Root ²²Na⁺ loss during washing and blotting
• + uptake pathways in S.maritima by various inhibitors'>4.2 Differentiation of Na⁺ uptake pathways in S.maritima by various inhibitors
• 2+and Li⁺on²²Na⁺ influx of S.maritima under low（25 mM NaCl）and high（150 mM NaCl）salinity'>4.2.1 Effects of Ca²⁺and Li⁺on²²Na⁺ influx of S.maritima under low（25 mM NaCl）and high（150 mM NaCl）salinity
• +和Cs⁺on²²Na⁺ influx of S.maritima under low and high salinity'>4.2.2 Effects of TEA⁺和Cs⁺on²²Na⁺ influx of S.maritima under low and high salinity
• 2+blocked²²Na⁺ influx of S.maritima under both low and high salinity'>4.2.3 Ba²⁺blocked²²Na⁺ influx of S.maritima under both low and high salinity
• +'>4.2.4 The turning-point of external NaCl concentrations for the inhibitory effects of TEA⁺
• + and Ba²⁺on²²Na⁺ influx among S.maritima,wheat and rice'>4.2.5 Comparison of the inhibitory effects of TEA⁺ and Ba²⁺on²²Na⁺ influx among S.maritima,wheat and rice
• 4⁺ altered the effects of TEA⁺ and Ba²⁺on²²Na⁺influx,Na⁺ accumulation and net Na⁺ uptake of S.maritima'>4.2.6 NH₄⁺ altered the effects of TEA⁺ and Ba²⁺on²²Na⁺influx,Na⁺ accumulation and net Na⁺ uptake of S.maritima
• +,K⁺ accumulation and growth in S.maritima'>4.2.7 Effects of a CCC inhibitor on Na⁺,K⁺ accumulation and growth in S.maritima
• + accumulation,growth and tissue water in S.maritima with increasing concentration of NaCl'>4.2.7.1 The effects of a CCC inhibitor on Na⁺ accumulation,growth and tissue water in S.maritima with increasing concentration of NaCl
• + accumulation,growth and tissue water in S.maritima with increasing concentration of KCl'>4.2.7.2 The effects of a CCC inhibitor on K⁺ accumulation,growth and tissue water in S.maritima with increasing concentration of KCl
• + on Na⁺ uptake and accumulation of S.maritima'>4.3 Effects of external K⁺ on Na⁺ uptake and accumulation of S.maritima
• +on²²Na⁺ influx and Na⁺ accumulation after 3 d of NaCl（2.5-200 mM）and KCl（10 or 50 mM）treatments'>4.3.1 Effects of K⁺on²²Na⁺ influx and Na⁺ accumulation after 3 d of NaCl（2.5-200 mM）and KCl（10 or 50 mM）treatments
• +on²²Na⁺ influx after 12 h of NaCl（2.5-200 mM）and KCl（10 or 50 mM）treatments'>4.3.2 Effects of K⁺on²²Na⁺ influx after 12 h of NaCl（2.5-200 mM）and KCl（10 or 50 mM）treatments
• +on²²Na⁺ influx and Na⁺ accumulation with 3 d of K⁺ starvation before12 h of NaCl（2.5-200 mM）and KCl（10 or 50 mM）treatments'>4.3.3 Effects of K⁺on²²Na⁺ influx and Na⁺ accumulation with 3 d of K⁺ starvation before12 h of NaCl（2.5-200 mM）and KCl（10 or 50 mM）treatments
• +,K⁺ accumulation and ²²Na⁺ influx in S.maritima cultured after salt stress treatment'>4.4 Time-course changes of Na⁺,K⁺ accumulation and ²²Na⁺ influx in S.maritima cultured after salt stress treatment
• + concentration in roots and shoots of S.maritima after salt stress treatment'>4.4.1 Time-course changes of Na⁺ concentration in roots and shoots of S.maritima after salt stress treatment
• 22Na⁺ influx into excised roots of S.maritima after salt stress treatment'>4.4.2 Time-course changes of ²²Na⁺ influx into excised roots of S.maritima after salt stress treatment
• + concentration and Na⁺/K⁺ ratio in roots and shoots of S.maritima after salt stress treatment'>4.4.3 Time-course changes of K⁺ concentration and Na⁺/K⁺ ratio in roots and shoots of S.maritima after salt stress treatment
• CHAPTER 5:DISCUSSION
• + uptake and transport pathways in higher plants'>5.1 S.maritima is a valuable plant material for characterizing Na⁺ uptake and transport pathways in higher plants
• + entry into root cells in plants'>5.2 NSCCS and LCT1 were not supported to be the major pathways for Na⁺ entry into root cells in plants
• + channel proteins might function in mediating low-affinity Na⁺ uptake through two pathways'>5.3 Two different K⁺ channel proteins might function in mediating low-affinity Na⁺ uptake through two pathways
• + uptake under low salinity condition'>5.3.1 HKT-type transporter might mediate low-affinity Na⁺ uptake under low salinity condition
• + uptake under high salinity condition'>5.3.2 AKT1-type channel might mediate low-affinity Na⁺ uptake under high salinity condition
• 5.3.3 The turning-point of external NaCl concentrations for the two pathways
• + exclusion among S.maritima,wheat and rice'>5.4 Comparison of the function of HKT-type transporter in Na⁺ exclusion among S.maritima,wheat and rice
• + exclusion in S.maritima under high salinity condition'>5.5 HKT-type protein and ABC transporter involved in plasma membrane ATPase activity may play roles in root Na⁺ exclusion in S.maritima under high salinity condition
• + uptake,especially under high salinity condition,but no significant effects on K⁺ uptake'>5.6 CCC transporter also might be a candidate for mediating low-affinity Na⁺ uptake,especially under high salinity condition,but no significant effects on K⁺uptake
• +（5 and 10 mM）stimulate Na⁺ uptake under high salinity（100-200 mM NaCl）'>5.7 Low concentration of K⁺（5 and 10 mM）stimulate Na⁺ uptake under high salinity（100-200 mM NaCl）
• +,K⁺ accumulation and ²²Na⁺ influx in S.maritima cultured after salt stress treatment'>5.8 Time-course changes of Na⁺,K⁺ accumulation and ²²Na⁺ influx in S.maritima cultured after salt stress treatment
• + concentration in roots and shoots and root ²²Na⁺ influx of S.maritima after salt stress treatment'>5.8.1 Time-course changes of Na⁺ concentration in roots and shoots and root ²²Na⁺ influx of S.maritima after salt stress treatment
• + concentration and Na⁺/K⁺ ratio in roots and shoots of S.maritima after salt stress treatment'>5.8.2 Time-course changes of K⁺ concentration and Na⁺/K⁺ ratio in roots and shoots of S.maritima after salt stress treatment
• CHAPTER 6:CONCLUSION
• References
• Academic papers,accomplished and undertaken projects