芥菜AGL18家族成员与开花整合子SOC1的互作分析

Analysis of Interactions Between AGL18 Family Members and Flowering Time Integrator Factor SOC1 in Brassica juncea

为阐明芥菜开花抑制因子AGL18与开花整合子SOC1间的互作调控机制,在芥菜‘QJ’的开花期克隆了AGL18-1,幼苗期克隆了AGL18-2和AGL18-3,它们分别编码257、257和258个氨基酸,为AGL18家族的3个成员。序列比对表明,芥菜AGL18家族成员与十字花科芜菁和油菜同源性均高达90%。酵母双杂交和BiFC试验表明:芥菜AGL18-1、AGL18-2和AGL18-3蛋白与SOC1不会发生蛋白相互作用。酵母单杂交和Dual-Glo~ Luciferase试验表明:AGL18-1、AGL18-2和AGL18-3中仅有花期AGL18-1蛋白与SOC1启动子间存在互作。为进一步筛选AGL18-1/SOC1的互作区域,分别截取了AGL18-1蛋白的M域和IKC域,发现仅M域与SOC1启动子存在相互作用,说明M域是介导花期AGL18-1蛋白与SOC1启动子互作的关键区域。这为深入研究AGL18与SOC1互作的分子机制及其对开花时间的调控奠定了基础。

AGAMOUS-LIKE 18 （AGL18） is a key regulatory factor in flowering time control of Brassica juncea. In order to clarify the regulatory mechanism of AGL18 protein interact with SOC 1 in flowering pathways, AGL18-1 gene from reproductive shoots as well as AGL18-2 and AGL18-3 from vegetative shoots in ＇QJ＇ germplasm ofBrassicajuneea, were cloned respectively. AGL18-1, A GL18-2 and AGL18-3 were members of AGL18 family and respectively encoded 257, 257 and 258 amino acids. Sequences analysis showed that they were more than 90% similar to Brassiea rapa and Brassica napus. Both Yeast two hybrid and BiFC showed that SOC 1 protein could not interact with AGL 18-1, AGL 18-2 or AGL18-3. Yeast one hybrid and Dual-Glo~ Luciferase assays indicated that SOC1 promoter could interact with AGL18-1 but not AGL18-2 and AGL18-3. To screen the interaction regions of AGL18-1/SOC1, the M-domain and IKC-domain truncations of AGL18-1 protein were separated and found that only the M-domain truncation could interact with SOCI promoter. It suggested that M-domain was the key region that mediated the interaction between AGL18-1 and SOC1 promoter. The work provided valuable information for in-depth studies on the molecular mechanisms of AGL18 protein with SOC1 and the flowering time in Brassicajuncea.