拟南芥转录因子HB22与CRY1相互作用研究

Research on the Interaction Between CRY1 and Transcription Factor HB22 in Arabidopsis

通过酵母双杂交的方法,从拟南芥转录因子库中筛选出了6个与CRY1相互作用的转录因子.为了测定其中的HB22与CRY1相互作用的强度,采用了ONPG与CPRG两种方法对其β-半乳糖苷酶活性进行了分析.结果显示在蓝光光强为50μmol/m2s,孵育时间为4 h的情况下,蓝光与暗处理情况下的β-半乳糖苷酶活性比值分别为1.668和2.18.进一步设置蓝光处理时间及光强梯度实验数据显示,在蓝光光强为50μmol/m2s孵育时间为3 h时,二者相互作用强度达到最高.说明HB22与CRY1的相互作用具有蓝光响应.对蓝光处理不同时间的野生型col-4与cry1缺失突变体的材料进行HB22基因的定量PCR分析,发现拟南芥cry1缺失突变体中该基因的表达量比野生型中高,在蓝光处理2 h时,缺失突变体中表达量为野生型中的6倍左右.说明CRY1可能介导蓝光抑制HB22基因表达.

Yeast two-hybrid system was performed to screen CRY1 interaction proteins in A rabidopsis tran- scription factors library, and six proteins were obtained. The interaction intensity between CRY1 and HB22, which is one of the six proteins, were measured by using ONPG and CPRG assay with/3-galactosidase The result showed that the ratio of/3-galactosidase activity in blue light or in the dark is 1.668 or 2.18 respec- tively when irradiated with blue light （50 txmol/m2s ） for 4 h. Their interaction irradiated with blue light for different time or with different fluence rates were further analyzed by CPRG. It was found that their interac- tion was the strongest when incubated under 50 Ixmol/m2s blue light for 3 h . The mRNA level of HB22 was then detected by real-time fluorescent quantitative PCR. It expressed higher in cryl mutant than in col-4, and 6 times higher than in col-4 when treated with blue light for 2 h. The results demonstrated that CRY1 may mediate blue light inhibition of HB22 gene expression.