A Plant Protein Kinase and A Phosphatidylinositol 3-Kinase Involved in Vacuolar Protein Transport and Development

96-30108 DeWald 植物细胞的液泡在功能上与真菌的液泡和哺乳动物细胞的溶酶体相似。此外，植物细胞液泡还有其他功能;它们在调节胞质pH、渗透压休克和蛋白质储存中起作用。植物液泡可占细胞总体积的90%。液泡的维持需要蛋白质和脂质从其合成位点到液泡的特异性分选。蛋白质的识别、分类和运输到液泡的过程是复杂的。超过30个基因与液泡蛋白分选有关。拟进行的研究的目标是确定磷脂酰肌醇3-激酶（PI 3-激酶）和丝氨酸/苏氨酸蛋白激酶在植物细胞液泡蛋白分选中的作用。本研究的具体目的是：1）研究拟南芥（Arabidopsis thaliana）Vps 34 p PI 3-激酶活性在植物液泡蛋白分选中的作用; 2）研究拟南芥（Arabidopsis thaliana）Vsp 15 p蛋白激酶活性在植物液泡蛋白分选中的作用。thaliana.将克隆Vps 15 p的基因并测序。通过用A.拟南芥cDNA文库中，或通过基于PCR的基因片段扩增，所述基因片段将被用于探测cDNA文库以鉴定所述基因。Vps 15 p和Vps 34 p的作用将在A.通过检查野生型植物中的组织和时间基因表达模式，以及通过记录在转基因植物中过表达该基因的影响，来研究拟南芥。预计这些数据将为植物中蛋白质转运的调节提供新的见解。 植物细胞的液泡在功能上与真菌的液泡和哺乳动物细胞的溶酶体相似。此外，植物细胞液泡还有其他功能;它们在调节胞质pH、渗透压休克和蛋白质储存中起作用。植物液泡可占细胞体积的90%以上。液泡的维持需要蛋白质和脂质从其合成位点到液泡的特异性分选。蛋白质的识别、分类和运输到液泡的过程是复杂的。超过30个基因与液泡蛋白分选有关。拟进行的研究的目标是确定磷脂酰肌醇3-激酶（PI 3-激酶）和丝氨酸/苏氨酸蛋白激酶在植物细胞液泡蛋白分选中的作用。本研究的具体目的是：1）研究拟南芥（Arabidopsis thaliana）Vps 34 p PI 3-激酶活性在植物液泡蛋白分选中的作用; 2）研究拟南芥（Arabidopsis thaliana）Vsp 15 p蛋白激酶活性在植物液泡蛋白分选中的作用。thaliana.将克隆Vps 15 p的基因并测序。通过用A.拟南芥cDNA文库中，或通过基于PCR的基因片段扩增，所述基因片段将被用于探测cDNA文库以鉴定所述基因。Vps 15 p和Vps 34 p的作用将在A.通过检查野生型植物中的组织和时间基因表达模式，以及通过记录转基因植物中过表达该基因的影响， 植物预计这些数据将为植物中蛋白质转运的调节提供新的见解。 ***

96-30108 DeWald The plant cell vacuole is functionally similar to fungi vacuoles and to lysosome of mammalian cells. In addition, plant cell vacuoles have other functions; they function in regulating cytosolic pH, osmotic shock, and in the storage of proteins. Plant vacuole can occupy up to 90% of the total cellular volume. The maintenance of vacuoles requires the specific sorting of proteins and lipids from their sites of synthesis to the vacuoles. The process of recognition, sorting, and transport of proteins to the vacuole is complex. More than 30 genes have been implicated in vacuolar protein sorting. The goal of the proposed research is to determine the role of a phosphatidylinositol 3-kinase (PI 3- kinase) and that of a serine/threonine protein kinase in protein sorting to plant cell vacuole. The specific objectives are to study 1) the role of Vps 34p PI 3-kinase activity in protein sorting to plant vacuoles in Arabidopsis thaliana; 2) the role of Vsp 15p, a protein kinase, in protein sorting to plant vacuole in A. thaliana. The gene for Vps 15p will be cloned and sequence. Cloning of the gene will be done by functional complementation of a Vps 15p yeast mutant by transforming with an A. thaliana cDNA library, or by PCR-based amplification of gene fragments that will be used to probe the cDNA library to identify the gene. The role of Vps 15p and of Vps 34p will be assessed in A. thaliana by examining tissue and temporal gene expression patterns in wild type plants, and by documenting the effects of overexpressing the gene in transgenic plants. It is expected that this data will provide new insights into how protein transport is regulated in plants. %%% The plant cell vacuole is functionally similar to fungi vacuoles and to lysosome of mammalian cells. In addition, plant cell vacuoles have other functions; they function in regulating cytosolic pH, osmotic shock, and in the storage of proteins. Plant vacuole can occupy up to 90% of the total cellular vo lume. The maintenance of vacuoles requires the specific sorting of proteins and lipids from their sites of synthesis to the vacuoles. The process of recognition, sorting, and transport of proteins to the vacuole is complex. More than 30 genes have been implicated in vacuolar protein sorting. The goal of the proposed research is to determine the role of a phosphatidylinositol 3-kinase (PI 3- kinase) and that of a serine/threonine protein kinase in protein sorting to plant cell vacuole. The specific objectives are to study 1) the role of Vps 34p PI 3-kinase activity in protein sorting to plant vacuoles in Arabidopsis thaliana; 2) the role of Vsp 15p, a protein kinase, in protein sorting to plant vacuole in A. thaliana. The gene for Vps 15p will be cloned and sequence. Cloning of the gene will be done by functional complementation of a Vps 15p yeast mutant by transforming with an A. thaliana cDNA library, or by PCR-based amplification of gene fragments that will be used to probe the cDNA library to identify the gene. The role of Vps 15p and of Vps 34p will be assessed in A. thaliana by examining tissue and temporal gene expression patterns in wild type plants, and by documenting the effects of overexpressing the gene in transgenic plants. It is expected that this data will provide new insights into how protein transport is regulated in plants. ***