Self-Incompatibility in Nicotiana Alata

在配子体自交不亲和（GSI）的茄科植物中，s等位基因花粉识别和排斥是由单个遗传位点s控制的。如果花粉的单个s等位基因与雌蕊中的任何一个s等位基因匹配，则花粉被拒绝。传递了孢子体和配子体中s位点的等位基因构成信息。s位点的产物是这些信息的载体。在雌蕊中表达的s位点产物包括一系列等位核糖核酸酶（S-RNases）。研究表明，每个S-等位基因编码一个不同的S- rnase。然而，尚不清楚这些差异如何导致s等位基因特异性识别。在雄性配子体（即花粉s）中尚未发现s等位基因特异性产物。本研究旨在了解s等位基因特异性识别和排斥的分子基础。第一步将是开发一个系统，其中s等位基因特异性识别和排斥可以被操纵。从烟叶中克隆的S-RNases将被用于向转基因植物中引入新的等位基因特异性。利用该表达系统，将有可能在体外操作克隆的S-RNase序列，并评估其对体内传粉行为的影响。作为转基因受体的植物材料的选择非常重要。N. alata的变换效率太低，无法产生必要数量的变换子。因此，转基因将被引入植物中，如烟叶（Nicotiana plumbaginifolia），并杂交到白杨（n.a alata）或能够s等位基因特异性排斥花粉的可转化F1杂种。在任何一种情况下，SI反应的固有特异性提供了良好的控制。麦克卢尔博士将寻求在转基因植物中引入或修改一种新的特异性。鉴定花粉- s产物是本提案的主要目标。提出了一种遗传与分子生物学相结合的方法。产生了两个S等位基因分离的F2大群体。一个给定S-等位基因的纯合子将被合并。利用这些汇集的材料，减法杂交和差异mRNA显示将用于鉴定推定的花粉- s cDNA。GSI具有帮助理解植物细胞间通讯的潜力。为了实现其潜力，GSI研究必须进入分析阶段。本课题旨在通过开发S-RNases的等位基因特异性功能的直接分析系统，以及鉴定花粉中负责等位基因特异性识别的产物来实现这一目标。＊＊＊

9316152 McClure In solanaceous plants with gametophytic self-incompatibility (GSI), S-allele pollen recognition and rejection are controlled by a single genetic locus, the S-locus. Pollen is rejected if its single S-allele matches either S-allele in the pistil. Information is communicated about the allelic constitution of the S-locus in the sporophyte and in the gametophyte. The products of the S-locus are the carriers of this information. The S-locus products expressed in the pistil comprise a series of allelic ribonucleases, the S-RNases. It has been shown that each S- allele encodes a different S-RNase. However, it is not known how these differences lead to S-allele specific recognition. No S-allele specific product has yet been identified in the male gametophyte (i.e., pollen-S). This proposal seeks to develop an understanding of the molecular basis for S-allele specific recognition and rejection. The first step will be to develop a system in which S-allele specific recognition and rejection can be manipulated. Cloned S-RNases from Nicotiana alata will be used to introduce new allelic specificity into transgenic plants. Using the expression system, it will be possible to manipulate cloned S-RNase sequences in vitro and assess the effects on pollination behavior in vivo. The choice of plant materials for use as transgene recipient is very important. The transformation efficiency of N. alata is too low to generate the necessary number of transformants. Therefore, transgenes will be introduced into a plant such as Nicotiana plumbaginifolia and crossed into N. alata or into a transformable F1 hybrid capable of S-allele specific pollen rejection. In either case, the inherent specificity of the SI reaction provides excellent controls. Dr. McClure will look for the introduction of, or modification of, a new specificity in the transgenic plants. Identification of the pollen-S product is a major goal of this proposal. A combined genetic and molecul ar biological approach is proposed. A large F2 population segregating for two S- alleles has been generated. Plants homozygous for a given S- allele will be pooled. Using these pooled materials, subtractive hybridization and differential mRNA display will be used to identify a putative pollen-S cDNA. GSI has the potential to help understand plant cell-cell communication. To realize its potential, GSI research must move into an analytical phase. This proposal aims to accomplish this by developing a system for direct analysis of the allelic specificity function of S-RNases, and identifying the product responsible for allele specific recognition in pollen. ***