OsSTLs介导独脚金内酯运输调控水稻株型的遗传和分子机制

Strigoalctones (SLs) regulate plant development and morphogenesis, transporting distantly from biosynthesis sites into particular organs to exert related biological function, however, the mechanism (s) of how SLs distribution and transport in plants are still largely unknown, which has greatly restrained the application of SLs in promoting plant architecture. We have previously characterized a family of genes called OsSTLs (Oryza sativa Strigolactone transport like) potentially involving in rice SLs transport. Our results show that the expression of OsSTL1, 2, 3 is induced by both endogenous and exogenous SLs levels. We found that the OsSTL1, a plasma membrane-localized protein internalizing in response to SL treatment. To confirm OsSTLs acting as SL transporters in rice, we created OsSTLs knock-out mutants using CRISPR/CAS9-mediated genome editing system. In this research, we aim to validate gene function and genetic effects of OsSTL(s) through analyzing the influence on SLs transport and plant architecture that caused by OsSTL(s) knock-out in the genetic materials we already made. And by inducible and over expressing protein in plants, we aim to assess the effect of protein dosages on plant architecture. We will use live cell imaging system to characterize protein distribution and transport. Overall, our research will explore new gene resource for SL studying in rice, and start from transport aspect to gain further insights into mechanisms of SLs action in regulating rice architecture.

独脚金内酯（SLs）参与调控植物的生长发育和形态建成。SLs从合成部位经过长距离运输进入特定组织发挥生物学功能，但目前对于SLs的转运和分配机制仍不明确，制约了SLs在作物株型改良上的应用。前期我们在水稻中鉴定了一类可能的SLs运输基因OsSTLs。OsSTL1,2,3基因的表达响应外源和内源SLs含量变化；在SLs诱导下定位于细胞膜的OsSTL1蛋白出现内在化。为了证实OsSTLs具有SLs运输功能，我们利用CRISPR/CAS9基因编辑技术获得了OsSTLs基因敲除植株。本项目拟利用已获得的遗传材料，分析OsSTLs基因功能缺失对水稻SLs运输和株型的影响，明确基因功能和遗传效应；利用诱导和过量表达方法确定运输蛋白剂量对株型的影响；运用活体细胞成像技术研究OsSTLs蛋白分布和运输特征。本项目将为水稻SLs研究发掘新的基因资源，并从激素运输的角度阐述SLs调控水稻株型发育的作用机理。

独脚金内酯（SLs）参与调控植物的生长发育和形态建成。SLs从合成部位经过长距离运输进入特定组织发挥生物学功能，但目前对于SLs的转运机制仍不明确，制约了SLs在作物株型改良上的应用。本项目证实了OsSTLs基因参与调控水稻独脚金内酯运输。OsSTLs基因编码多向耐药性转运蛋白(Pleiotropic drug resistance, PDR)，与矮牵牛独脚金内酯运输蛋白PhPDR1高度同源。OsSTL1,2,3,5在GR24处理1小时后表达量显著上调能快速响应独脚金内酯激素含量的变化。OsSTLs同源基因在功能上存在冗余，我们利用CRISPR-Cas9系统对OsSTL家族的7个基因进行了敲除，创制了单基因和多基因敲除纯合株系。荧光独脚金内酯根部运输实验表明，OsSTLs基因敲除导致独脚金内酯在水稻根部运输受阻。OsSTLs基因敲除株系根长较野生型缩短，与独脚金内酯合成突变体d10相似。OsSTLs基因的组织表达模式和亚细胞定位出现了分化，暗示OsSTLs编码的转运蛋白运输的底物可能具有多样性。敲除株系的农艺性状分析发现，随着OsSTL家族基因敲除数量的增加，水稻生长发育受到了严重的抑制，类病斑坏死和早衰现象加剧。表明了OsSTL基因家族编码的PDR转运蛋白在参与独脚金内酯运输的同时，也协同调控其他物质的转运，从而维持水稻的正常生长。进一步的研究发现，OsSTL(s)家族基因表达量在不同程度上被茉莉酸甲酯、脱落酸、生长素诱导，暗示了OsSTL(s)基因编码的PDR转运蛋白可能在植物激素调控水稻生长发育过程中具有重要作用。本项目的研究明确了OsSTL基因在水稻独脚金内酯运输中的功能和遗传效应，初步揭示了一个多基因协同参与调控水稻独脚金内酯运输的新模式。