A novel model system to study the rapid diversification of R genes

植物利用抗病(R)基因来识别病原体，其中大部分基因编码核苷酸结合富亮氨酸重复序列（NB-LRR）蛋白。每个R蛋白识别特定的效应蛋白。为了对抗病原菌效应基因的快速多样化，认为R基因也在快速进化。这一观点得到了高度多态性在r基因中观察到的支持。然而，人们对r基因多样化的机制知之甚少。我们分析了拟南芥UNI - 1d突变体，该突变体含有UNI基因的一个半显性和功能获得等位基因，这是一个具有nb - lrr相关结构的R基因。uni-1D具有组成活性的R蛋白，可在没有任何病原体感染的情况下诱导抗性反应。此外，单- 1d杂合突变体（以下简称uni1D/+）在花序茎的茎尖分生组织中迅速消耗干细胞，形成极短的茎。有趣的是，在正常生长条件下，我们很少但重复地观察到，uni-1D/+产生的嵌合扇区表现出与野生型相似的长花序茎的形态。当我们检查这个嵌合茎中的核苷酸序列时，我们总是发现额外的突变，这些突变可能会破坏单维蛋白的功能。此回退事件发生的时间小于0。占总人口的5%用烷基化剂EMS进行试验，成功地将还原频率提高了30%左右。此外，当我们用导致DNA双链断裂的zeocin或通过耗尽脱氧核苷酸三磷酸池导致DNA修复和复制缺陷的羟基脲（HU）处理uni-1D/+时，逆转频率显著增加。这表明uni-1D系统可以轻松有效地检测出uni-1D基因中各种类型的核苷酸改变。目前，我们正在利用这一系统分析R基因快速多样化的分子机制，我们的初步结果表明DNA修复机制参与了这一现象。少

Plants use disease resistance(R) genes, most of which encode nucleotide-binding leucine-rich repeat(NB-LRR) protein, to recognize pathogens. Each R protein recognizes the specific effector protein. To counter the rapid diversification of pathogen effector genes, it thought that R genes also evolve rapidly. This idea is supported that high degree of polymorphism is observed in R-genes. However, little is known about the mechanisms underlying the R-gene diversification. We analyzed Arabidopsis uni-1D mutant, harbors a semi-dominant and gain-of-function allele of UNI gene, an R gene that has a NB-LRR-related structure. The uni-1D has a constitutively active R protein to induce resistance responses without any pathogen infection. Furthermore, uni-1D heterozygous mutant(hereafter uni1D/+) shows rapidly consume stem cells in the shoot apical meristem of the inflorescence stem, resulting in formation of very short stem. Interestingly, under normal growth condition, we infrequently but repeate … More dly observed that uni-1D/+produced chimeric sectors display the morphology of wild-type-like long inflorescence stem. When we checked nucleotide sequences in this chimeric stem, we always found additional mutations, which presumably disrupted the uni-1D protein function. This reversion event occurs less than 0. 5% of individuals among the population. When we tried with EMS, an alkylating agent, we succeeded to increase the reversion frequency about 30%. Furthermore, when we treated uni-1D/+with zeocin, which causes DNA double-strand breaks, or hydroxyurea(HU), which induces defects of DNA repair and replication by depletion by depleting deoxynucleotide triphosphate pools, the reversion frequency significantly increased. These suggest that the uni-1D systems can easily and efficiently detect various types of nucleotide alterations in the uni-1D gene. Currently, we are analyzing molecular mechanisms underlying the rapid diversification of R genes using this system and our preliminary results imply the involvement of DNA repair machinery in this phenomenon. Less