大豆GmPIN2家族基因调控根系发育功能初探

Function of GmPIN2 family gene in regulating root development in soybean

生长素作为重要的植物激素之一。生长素的转运参与了植物各组织器官发育调控。在拟南芥中,生长素转运调控主要由PIN家族蛋白所介导,其中拟南芥AtPIN2主要通过介导生长素向基运输调控根的向重力性。大豆中PIN2家族蛋白及其功能研究尚未有报道。本研究通过构建系统进化树和蛋白质结构域分析发现,大豆GmPIN2a和GmPIN2b为AtPIN2的同源基因。组织表达分析研究发现,GmPIN2a和GmPIN2b在根、根瘤原基和根瘤等组织中高表达。GmPIN2a和GmPIN2b在根部主要表达在根尖表皮和外部皮层细胞,在根瘤中GmPIN2a、GmPIN2b均定位于根瘤基部维管束区,此外,GmPIN2a定位在根瘤顶部表皮及外皮层。利用基因编辑技术CRISPR/Cas9(Clustered Regularly Interspersed Short Palindromic Repeats/CRISPR associated 9)同时敲除GmPIN2a和GmPIN2b后发现,Gmpin2ab突变体根具有明显的重力缺失表型。与野生型相比,Gmpin2ab和35S::GmPIN2b的根面积和侧根长度显著降低,Gmpin2ab侧根夹角显著上升,而35S::GmPIN2b侧根夹角不变。综上所述,GmPIN2a和GmPIN2b通过介导生长素向基运输对大豆根型调控具有重要作用。本研究为深入探究大豆PIN蛋白介导生长素极性运输在根形态建成的作用机制奠定了一定研究基础。

Auxin is one of the important plant hormones.Auxin transport is involved in the regulation of plant tissue and organ development.In Arabidopsis thaliana,auxin transport is mainly regulated by PIN family proteins,among which AtPIN2 controls auxin basipetal transport and regulates root gravitation in Arabidopsis.However,GmPIN2 family proteins and their functions are still largely unknown in soybean.In this study,we identified two AtPIN2 homologous genes of GmPIN2a and GmPIN2b through phylogenetic and protein domain analysis.The relative expression level showed that GmPIN2a and GmPIN2b were highly expressed in root and nodule.Both genes were highly expressed in root epidermis and cortex,and the vascular bundles of the root nodule.Only GmPIN2a expressed in the epidermis and outer cortex at the tip of the root nodule.Furthermore,Gmpin2ab mutants were generated by CRISPR/Cas9(Clustered Regularly Interspersed Short Palindromic Repeats/CRISPR Associated 9)gene editing technology.Gmpin2ab mutants had a significant loss of root gravitropism.Moreover,the loss of Gmpin2 resulted in a significant decrease of root area and lateral root length,while the lateral root angle of Gmpin2ab significantly increased.Overexpression of GmPIN2b also resulted in a decrease in root area and lateral root length,but the lateral root angle unchanged.In conclusion,GmPIN2a and GmPIN2b play important roles in regulation of soybean root development via mediating auxin basipetal transport.This study laid a foundation for further analysis of the function and mechanism of GmPIN proteins in soybean root morphogenesis.