PbrS-RNase介导PbrPPA5乙酰化调控梨自交不亲和性反应的分子机制

梨是典型的配子体型自交不亲和性果树，其自我花粉管在花柱中段停止生长，不能完成受精。申请人前期研究表明，在梨自我花粉管内PbrS-RNase能够诱导无机焦磷酸酶PbrPPA5乙酰化，促使PbrPPA5向细胞核转移，与转录因子PbrbZIP1形成抑制复合体后阻止果胶甲基酯酶的表达，导致花粉管尖端膨大，但PbrPPA5如何响应PbrS-RNase尚不清楚。因此，本项目以‘砀山酥梨’为试材，通过反义寡核苷酸等技术，鉴定PbrPPA5调控梨花粉管生长特性；构建仿乙酰化突变、蛋白质免疫印迹和原位免疫荧光染色技术，分析乙酰化修饰对PbrPPA5蛋白功能及其定位的影响；通过酵母双杂交、免疫共沉淀与串联质谱等技术，探明赖氨酸乙酰转移酶在调控PbrPPA5乙酰化中的功能机制。相关研究将进一步完善梨自交不亲和性反应的信号转导路径，为培育梨自交亲和新品种提供理论基础和基因资源。

Pear is a fruit tree with typical gametophytic self-incompatibility, in which the growth of self-pollen stops in the middle of the style, resulting in the unsuccessful fertilization. Previous study of our group showed that the incompatible PbrS-RNase resulted in the acetylation of PbrPPA5 and induced its translocation into the pollen tube nucleus. In the nucleus, PbrPPA5 directly binds to the transcription factor PbrbZIP1 to form a functional transcriptional repressor complex, which resulted in the swelling of pollen tube tip. However, the underlying mechanism of PbrPPA5 reponding to PbrS-RNase remains unclear. In this project, ‘Dangshansuli’ is selected as the plant material. Oligo deoxynucleotide is applied to identify the biological function of PbrPPA5 in the regulation of pear pollen tube growth. Acetyl-mimetic mutants, western blot and immunohistochemical analysis are performed to analyze the regulation of acetylation on both function and subcellular localization of PbrPPA5. Through yeast two hybrid, co-immunoprecipitation and liquid chromatography-tandem mass spectrometry, we intend to clarify the regulatory mechanism of lysine acetyl transferase on modifying PbrPPA5. Overall, our results will further elucidate the signaling pathway of self-incompatibility in pear and provide both genetic resource and theoretical foundation for the breeding of self-compatible pear cultivar.