POWRE: Characterization of a Novel ATPase

所有植物细胞都必须积极地从细胞质中输出钠。 钠在细胞质中的积累导致许多酶系统的抑制，并且是由于盐度（无论是自然的还是由于农田灌溉）而造成损害的主要原因。 因此，了解Na+运输的机制不仅对了解植物的功能很重要，而且还可以提高我们开发更耐盐碱化的作物的能力。 这个项目将使用两种轮藻，它们在耐盐条件的能力上不同。 Na ~+通过质膜输出而从细胞质中清除对于获得耐盐性是重要的，有证据表明ATP酶可能参与这种转运，包括Na ~+/H ~+反向转运不足以维持所测量的Na ~+梯度的生物物理证据，盐培藻类中不寻常的P型ATP酶的免疫学证据，以及耐盐藻类中不寻常的P型ATP酶的分子证据。 拟议研究的目标是 从耐盐植物长叶轮藻中分离一个不常见的ATP酶基因，并进行序列分析。 利用北方杂交技术检测该基因在C. longifolia适应于盐水和淡水介质，并经历对盐胁迫的适应。 为了确定盐敏感性物种，C。corpus a具有类似的基因，如果是这样，在什么条件下表达。长期目标包括通过在缺乏Na+或H+输出能力的酵母突变体中表达或通过在轮藻中通过将mRNA显微注射到节间来过表达来确定ATP酶基因的运输功能。 最终，基于这项研究的信息可以用来改变轮藻和其他植物的耐盐性，通过遗传操作来提高钠从细胞质中的输出。 这项研究将在POWRE资助计划下进行，因为它将帮助作者在植物盐度研究领域保持竞争力。 生物物理技术在过去已成功地用于研究这一领域的重要问题，但该项目的分子方面必须发展，以保持竞争力。

All plant cells must actively export sodium from the cytoplasm. Accumulation of sodium in the cytoplasm results in inhibition of a number of enzyme systems, and is a major cause of damage due to salinity, whether natural or due to irrigation of agricultural fields. A knowledge of the mechanisms of Na+ transport is therefore not only important in general understanding of the function of plants, but also enhances our ability to develop crop plants which are more resistant to salinization. This project will use two species of Charophyte algae that differ in their ability to tolerate saline conditions. Re-moval of Na+ from the cytoplasm by export across the plasma mem-brane is important in achieving salt tolerance, and there is evidence that an ATPase may be involved in this transport, including biophysical evidence that a Na+/H+ antiport is not sufficient to maintain the Na+ gradients measured, immunological evidence of an unusual P-type ATPase in salt-cultured algae, and molecular evidence for an unusual P-type ATPase in the salt-tolerant algae. The objectives of the proposed research are To isolate and sequence a full-length clone for an unusual ATPase from the salt-tolerant species, Chara longifolia. To use northern blot analysis to determine expression of this gene in C. longifolia adapted to saline and freshwater media, and undergoing adaptation to salt stress. To determine whether the salt-sensitive species, C. corallina, has a similar gene and, if so, under what conditions it is expressed. Long-term goals include determining the transport function of the ATPase gene either by expres-sion in yeast mutants lacking Na+ or H+ export capability or by overexpression in Chara by micro-injection of mRNA into the internode. Ultimately, information based on this research could be used to alter the salt tolerance of Chara and other plants by genetic manipulation to enhance sodium export from the cytoplasm. This research will be performed under the POWRE grant program because it will aid the author in remaining competitive in the field of salinity studies in plants. Bio-physical techniques have been successfully employed in the past to investigate important ques-tions in this field, but molecular aspects of the project must be developed in order to remain competitive.