Molecular Basis of Gametophytic Self-Incompatibility in Nicotiana and Petunia

本课题旨在研究配子体自交不亲和中雌蕊s蛋白的功能及s基因的表达调控。高博士将以烟草和矮牵牛花为研究对象，1)通过测试将新的s基因引入与不同s等位基因纯合的植物中，或将反义s基因引入与同一s等位基因纯合的植物中，对自交不亲和行为的影响，建立s蛋白负责自交不亲和反应的直接证据；2)利用转基因植物中表达的嵌合s蛋白，鉴定s蛋白的s等位基因特异性结构域；3)检测突变S-蛋白的保守半胱氨酸残基对其功能的影响；4)通过将s基因的5′上游序列与报告基因GUS融合，确定s基因组织特异性和发育调控表达的序列，然后采用序列删除法检测对转基因植物中GUS表达的影响；5)利用一种独特的自相容花柱部分突变体，鉴定出与s位点花柱活性部分相关的蛋白质。这些实验的结果将提供自交不亲和分子基础上的重要信息。许多高等植物有一种称为自交不亲和的机制，可以防止来自特定植物的花粉与同一植物的雌性组织（雌蕊）自交受精。控制这种自交不亲和的基因已被确定，但不亲和发生的机制尚不清楚。除了植物生物学家对它的固有兴趣外，对这一过程的理解在农业上也有明显的应用。

The proposed project is to study the functional aspects of pistil S-proteins and regulation of S-gene expression in gametophytic self-incompatibility. Using Nicotiana alata and Petunia inflata, Dr. Kao will 1) establish direct evidence that S-proteins are responsible for the self-incompatibility reaction by testing the effect on self-incompatibility behavior of introducing either a new S-gene into plants homozygous for a different S-allele, or an antisense S-gene into plants homozygous for the same S-allele; 2) identify the s-allele specificity domain of the S-protein by using chimeric S-proteins expressed in transgenic plants; 3) test the effect of mutating the conserved cysteine residues of S- proteins on their function; 4) determine the sequences controlling tissue specific and developmentally regulated expression of S-genes by fusing their 5' upstream sequences to a reporter gene, GUS, and then using sequential deletion to test the effect on expression of GUS in transgenic plants; 5) identify a protein associated with the style activity part of the S-locus using a unique self-compatible style-part mutant of N. alata. The result of these experiment will yield important information on the molecular basis of self-incompatibility. %%% Many higher plants have a mechanism, termed self-incompatibility, with prevents the pollen from a given plant from self fertilizing the female tissue (pistil) of the same plant. Genes controlling this self- incompatibility have been identified but the mechanism whereby the incompatibility occurs is unknown. In addition to its inherent interest to plant biologists, there are clear applications to agriculture to an understanding of this process.