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盐生植物海滨碱蓬Na~+吸收和积累的研究

作　者: 张金林
导　师: 任继周；王锁民
学　校: 兰州大学
专　业: 草业科学
关键词: 低亲和性Na~+吸收途径 22Na+内流离子通道抑制剂 HKT AKT1 CCC Na~+积累拒钠海滨碱蓬
分类号: S184
类　型: 博士论文
年　份: 2008年
下　载: 274次
引　用: 2次
阅　读: 论文下载

内容摘要

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。

全文目录

Acknowledgements  5-6
List of abbreviation used  6-7
摘要  7-9
Abstract  9-19
CHAPTER 1:INTRODUCTION  19-22
CHAPTER 2:PROGRESS ON Na~+ UPTAKE AND Na~+ TRANSPORT IN HIGHER PLANTS  22-60
  2.1 Background  22-24
  2.2 Negative effects of salinity on plant productivity  24-27
    2.2.1 Osmotic stress  24-25
    2.2.2 Ionic stress  25-27
    2.2.3 Secondary stresses  27
  2.3 Na~+ uptake into roots  27-48
    2.3.1 Physiological and electrophysiological mechanisms of Na~+ uptake in higher plants  27-39
      2.3.1.1 K~+,Na~+ competition and selectivity  27-29
      2.3.1.2 Na~+-Ca~(2+)interactions under salt environments  29-32
      2.3.1.3 Effects of driving force or electrochemical potential across the plasma on Na~+ uptake  32-34
      2.3.1.4 Effects of external salt concentration on Na~+ uptake  34-37
      2.3.1.5 Experimental methods in studying the physiological mechanisms of Na~+ uptake  37-39
    2.3.2 Molecular biological mechanism of Na~+ uptake in higher plants  39-48
      2.3.2.1 NSCCs/VICs  39-41
      2.3.2.2 LCT1  41-42
      2.3.2.3 KUP/HAK/KT  42-43
      2.3.2.4 HKT  43-46
      2.3.2.5 AKT1  46-47
      2.3.2.6 CCC  47-48
  2.4 Na~+ Transport in higher plants  48-60
    2.4.1 Control of Na~+ xylem loading  48-50
    2.4.2 Na~+ retrieval from the xylem  50-51
    2.4.3 Na~+ recirculation in the phloem  51-52
    2.4.4 Na~+ extrusion or effiux from the root  52-54
    2.4.5 Intracellular compartmentation of Na~+ into the vacuoles  54-58
    2.4.6 Na~+ excretion  58-60
CHAPTER 3:MATERIAL AND METHODS  60-63
  3.1 Plant material and growth condition  60
  3.2 Treatments  60-61
  3.3 Growth measurements,and Na~+ and K~+ concentration determination  61
  3.4 ~(22)Na~+ influx experiments  61-62
  3.5 Statistical analysis  62-63
CHAPTER 4:RESULTS  63-107
  4.1 The bases of root ~(22)Na~+ influx in Suaeda maritima  63-68
    4.1.1 Time course of root ~(22)Na~+ influx  63-65
    4.1.2 Effects of external Na~+ concentration on root ~(22)Na~+ influx  65-66
    4.1.3 The correlation between root ~(22)Na~+ influx and root Na~+ concentration  66
    4.1.4 The correlation between ~(22)Na~+ influx and fresh weight of the root  66-67
    4.1.5 Root ~(22)Na~+ loss during washing and blotting  67-68
  4.2 Differentiation of Na~+ uptake pathways in S.maritima by various inhibitors  68-86
    4.2.1 Effects of Ca~(2+)and Li~+ on ~(22)Na~+ influx of S.maritima under low(25 mM NaCl)and high(150 mM NaCl)salinity  69-70
    4.2.2 Effects of TEA~+和Cs~+ on ~(22)Na~+ influx of S.maritima under low and high salinity  70-71
    4.2.3 Ba~(2+)blocked ~(22)Na~+ influx of S.maritima under both low and high salinity  71-72
    4.2.4 The turning-point of external NaCl concentrations for the inhibitory effects of TEA~+  72-73
    4.2.5 Comparison of the inhibitory effects of TEA~+ and Ba~(2+)on ~(22)Na~+ influx among S.maritima,wheat and rice  73-75
    4.2.6 NH_4~+ altered the effects of TEA~+ and Ba~(2+)on ~(22)Na~+ influx,Na~+ accumulation and net Na~+ uptake of S.maritima  75-78
    4.2.7 Effects of a CCC inhibitor on Na~+,K~+ accumulation and growth in S.maritima  78-86
      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  80-83
      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  83-86
  4.3 Effects of external K~+ on Na~+ uptake and accumulation of S.maritima  86-100
    4.3.1 Effects of K~+ on ~(22)Na~+ influx and Na~+ accumulation after 3 d of NaCl(2.5-200 mM)and KCl(10 or 50 mM)treatments  86-91
    4.3.2 Effects of K~+ on ~(22)Na~+ influx after 12 h of NaCl(2.5-200 mM)and KCl(10 or 50 mM)treatments  91-94
    4.3.3 Effects of K~+ on ~(22)Na~+ 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  94-100
  4.4 Time-course changes of Na~+,K~+ accumulation and ~(22)Na~+ influx in S.maritima cultured after salt stress treatment  100-107
    4.4.1 Time-course changes of Na~+ concentration in roots and shoots of S.maritima after salt stress treatment  100-103
    4.4.2 Time-course changes of ~(22)Na~+ influx into excised roots of S.maritima after salt stress treatment  103-104
    4.4.3 Time-course changes of K~+ concentration and Na~+/K~+ ratio in roots and shoots of S.maritima after salt stress treatment  104-107
CHAPTER 5:DISCUSSION  107-128
  5.1 S.maritima is a valuable plant material for characterizing Na~+ uptake and transport pathways in higher plants  107-110
  5.2 NSCCS and LCT1 were not supported to be the major pathways for Na~+ entry into root cells in plants  110-113
  5.3 Two different K~+ channel proteins might function in mediating low-affinity Na~+ uptake through two pathways  113-118
    5.3.1 HKT-type transporter might mediate low-affinity Na~+ uptake under low salinity condition  113-114
    5.3.2 AKT1-type channel might mediate low-affinity Na~+ uptake under high salinity condition  114-116
    5.3.3 The turning-point of external NaCl concentrations for the two pathways  116-118
  5.4 Comparison of the function of HKT-type transporter in Na~+ exclusion among S.maritima,wheat and rice  118-119
  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  119-122
  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  122-124
  5.7 Low concentration of K~+(5 and 10 mM)stimulate Na~+ uptake under high salinity(100-200 mM NaCl)  124-125
  5.8 Time-course changes of Na~+,K~+ accumulation and ~(22)Na~+ influx in S.maritima cultured after salt stress treatment  125-128
    5.8.1 Time-course changes of Na~+ concentration in roots and shoots and root ~(22)Na~+ influx of S.maritima after salt stress treatment  125-126
    5.8.2 Time-course changes of K~+ concentration and Na~+/K~+ ratio in roots and shoots of S.maritima after salt stress treatment  126-128
CHAPTER 6:CONCLUSION  128-130
References  130-165
Academic papers,accomplished and undertaken projects  165-168

盐生植物海滨碱蓬Na~+吸收和积累的研究

内容摘要

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