隐花色素GmCRY2与转录因子GmCIB3调控大豆开花的机理研究

Cryptochromes are blue light receptors found in all evolutionary lineages, which regulate photomorphogenesis and flowering time in plants. Soybean is a typical short-day plant, for which the photoperiodic regulation of development is not well studied. A previous study identified a BHLH transcription factor, CIB1 as the first protein which interacts with CRY2 in a blue light-dependent fashion and promotes flowering in Arabidopsis. Here we systematically analyzed 9 soybean CIB1-like proteins for their physical interaction with GmCRYs, and identified GmCIB3 as a GmCRY2-ineracting protein. Similar to the Arabidopsis proteins, GmCRY2 and GmCIB3 undergo blue light-specific interaction in yeast. Surprisingly, we found that GmCRY2 and GmCIB3 act as suppressors of floral initiation in soybean. Reduced expression of GmCRY2 caused accelerated flowering, whereas overexpression of GmCRY2 and GmCIB3 delayed flowering, indicating that GmCRY2 in soybeans may be involved in distiquish functions in compare to CRY2 in Arabidopsis. To this end, we will try to identify their downstream genes, analysis the DNA-binding activity of GmCRY2-GmCIB3 complex to their targeted DNA motif in response to blue light and further explore the function and action mechanism of GmCRY2 and its interaction partner GmCIB3 in the regulation of photoperiod-dependent flowering pathway. This research will provide a broad knowledge for the breeding of wide-suitable soybean varieties.

大豆属于短日植物，不同品种由于其生育期受光周期的影响而表现种植纬度偏好性，因此研究大豆开花相关基因对创制广适优良新品种具有重要指导意义。隐花色素是普遍存在于生物中的一类蓝光受体，其在植物中的主要功能是参与光形态建成和光周期介导的开花反应。CIB1是第一个被发现的与拟南芥隐花色素CRY2 在蓝光下特异作用的蛋白，被光激活的CRY2与CIB1结合并启动FT基因表达促进开花。我们分析了大豆中9个GmCIBs与GmCRY2蛋白相互作用，发现GmCIB3与GmCRY2在蓝光下特异结合。然而GmCRY2 RNAi转基因大豆表现早花，GmCRY2和GmCIB3过表达导致大豆晚花，表明这两个基因在大豆和拟南芥中具有不同的开花调控功能。本项目将通过遗传、生化和细胞生物学手段深入研究GmCRY2-GmCIB3介导光周期调控大豆开花的分子机制,并为培育广适大豆新品种提供理论参考。

豆是典型的短日农作物，对大豆光周期开花机理的研究对农业生产具有重要意义。我们已初步研究大豆隐花色素的功能，发现大豆隐花色素与拟南芥隐花色素间存在较大差别。本项目通过反向遗传学手段研究大豆中GmCRY2与其相互作用蛋白GmCIB1的生物学功能， 在连续光照条件下GmCRY2敲除的转基因大豆提前开花，而过表达GmCRY2、GmCIB1则导致大豆晚开花，表明了GmCRY2和GmCIB1抑制大豆开花。酵母双杂交和BiFC试验表明，GmCRY2和GmCIB1有蓝光特异的相互作用，且这种相互作用随着蓝光光强的增强和光照时间的增加而增强。GmCRY2的PHR结构域参与和GmCIB1蓝光依赖的相互作用。体内外Pull-down试验进一步验证了GmCRY2与GmCIB3间蓝光依赖的相互作用。GmCIB3体外可与E-box结合，染色体免疫共沉淀（ChIP）试验显示，GmCIB1与GmFT3和GmFT4 染色体区的E-box结合，但不能与开花抑制基因GmTFL1和GmTFL3的染色体片段结合。推测GmCIB1在大豆体内可能通过抑制GmFT的表达抑制大豆开花。