水稻3-磷脂酰肌醇激酶FAB1/PIKfyve基因家族的鉴定及功能分析  

Identification and Functional Analysis of 3-Phosphatidylinositol Kinase FAB1/PIKfyve Gene Family in Rice(Oryza sativa)

作　　者：徐忆纯 梁婉琪[1] 王灿华[1] 余婧[1] Xu Yichun;Liang Wanqi;Wang Canhua;Yu Jing(School of Life Sciences and Biotechnology,Shanghai Jiao Tong University,Shanghai,200240)

机构地区：[1]上海交通大学生命科学技术学院,上海200240

出　　处：《分子植物育种》2021年第22期7309-7318,共10页Molecular Plant Breeding

基　　金：国家重点研发项目(2016YFD0100902);国家自然科学基金项目(31700276)。

摘　　要：FAB1/PIKfyve是催化3-磷酸磷脂酰肌醇(PtdIns3P)形成3,5-二磷酸磷脂酰肌醇(PtdIns(3,5)P_(2))过程的关键酶,其产物PtdIns(3,5)P_(2)在真核细胞发育中发挥重要功能。为探寻PtdIns(3,5)P_(2)在水稻生殖发育过程中的功能,本研究结合生物信息学和遗传学方法,对水稻FAB1/PIKfyve基因进行鉴定,分析其理化性质、基因结构、保守结构域、系统进化、顺式作用元件和组织表达模式并利用CRISPR/Cas9基因编辑技术获得osfab1b突变体。生物信息学分析结果显示,水稻基因组中共鉴定到9个FAB1基因家族成员;基因结构分析显示FAB1家族基因结构存在差异,外显子数量为8~12个;保守结构域分析表明仅Os FAB1A和Os FAB1B含有N端FYVE结构域,其余成员具有Cpn60_(T)CP1结构域和PIPKc激酶结构域;系统进化分析提示FAB1家族功能在单双子叶植物中具有高度保守性;FAB1基因上游调控区域顺式元件预测发现多种生长发育相关、光响应以及激素和胁迫响应顺式元件;组织表达模式分析显示大多数FAB1基因为泛表达,其中FAB1C亚类基因在内外稃中的高表达提示其可能参与花器官发育。最后通过CRISPR/Cas9系统得到osfab1b突变体,经碘染观察花粉活力无明显异常,暗示FAB1家族在水稻生殖发育调控中存在功能冗余。本研究结果为单子叶模式植物水稻中磷脂酰肌醇调控网络及其生物学功能的研究提供了理论参考。FAB1/PIKfyve is a key enzyme that catalyzes phosphatidylinositol 3-phosphate(PtdIns3P)to form phosphatidylinositol 3,5-bisphosphate(PtdIns(3,5)P_(2)).Its product PtdIns(3,5)P_(2)plays an important role in the development of eukaryotic cells.In order to figure out the function of PtdIns(3,5)P_(2)in rice reproductive development,this study combined bioinformatics and genetics methods to identify the rice FAB1/PIKfyve genes,analyzed the physical and chemical properties,gene structures,conserved domains,phylogenetic tree,cis-acting elements tissue expression profiles and used CRISPR/Cas9 gene editing technology to obtain osfab1b mutants.Bioinformatics analysis results showed that nine FAB1 gene family members were identified from the whole genome of Oryza sativa.Gene structure analysis indicated the differences existed in the gene structure of FAB1 family,the number of exons are 8~12.Conserve domain analysis showed that Os FAB1A and Os FAB1B contained the N-terminal FYVE domain,and the remaining members contained the Cpn60_TCP1 domain and PIPKc kinase domain.Phylogenetic analysis indicated that the functions of FAB1 family were highly conserved in mono-and dicotyledonous plants.Prediction of cis-element in the upstream regulatory region of FAB1 genes revealed a variety of growth-related,light-responsive,and hormone and stress-responsive cis-elements.Tissue expression profiles showed that most of FAB1 genes were global-expressed,and the high expression of FAB1C sub-cluster genes in lemma and palea suggested that they might be involved in floral organ development.Finally,the osfab1b mutants were obtained through the CRISPR/Cas9 system,and the pollen vitality of the osfab1b mutants showed no significantly abnormality,implied that FAB1 family had functional redundancy in the reproductive development of rice.The present results provide a theoretical reference for biological function studies of the phosphatidylinositol regulatory network in rice.

关 键 词：水稻(Oryza sativa) FAB1/PIKfyve基因家族 3 5-二磷酸磷脂酰肌醇 CRISPR/Cas9 

分 类 号：S511[农业科学—作物学]