Characterization Of JA Signal Receptor HbCoi1 In Hevea Brasiliensis

Hevea brasiliensis is the main source of natural rubber. Laticifer is important organ of latex synthesis and storage in rubber trees. Mechanical damage or treatment of exogenous JA can induce differenciation of laticifer, suggesting that JA is important signal molecules regulating the differentiation of Hevea laticifer. Coi1 is JA receptor and important regulators in jasmonic acid related signaling pathway in plant. HbCoi1 has been cloned from Hevea brasiliensis, but the function of HbCoi1 is still unknown. HbCoi1 was characterized in the following setions:(1) The expression vector was first constructed for HbCoi1 and transformed into Arabidopsis coil mutant. Totally,123 T1 transgenic plants were obtained.9 T2 tramsgenic lines with homozygous coil backgroud were generated through self-crossing and verified by PCR and restriction digestion of PCR product. The fertility phenotype was restored in the most T1 transformed with HbCoi1 in homozygous coil backgroud, suggesting that HbCoi1 complemented the function of Arabidopsis coil.(2) The bait vector of HbCoi1 (pGBKT7-HbCoi1) was then constructed. The interactions between HbCoi1 and JAZ proteins of Hevea brasiliensis (HbJAZs) were analysed by yeast two hybrid (Y2H). Y2H results showed that HbCoi1 interacted with HbJAZ8 and HbJAZ9 when coronatine existed, suggesting that HbCoi1 regulated the downstream signal through interation with HbJAZ proteins.(3) The promoter of HbCoi1 gene was cloned by PCR and inserted into pHIS2.1 to construct a bait vector. The interaction of The promoter of HbCoi1 with MYC transcription factors of jasmonic acid signaling pathway in Hevea brasiliensis, HbMYC2, HbMYC3, and HbMYC4, was analysed using yeast one hybrid (Y1H). No interaction between MYC transcription factors and the promoter of HbCoi1 gene was observed, it was therefore proposed that HbCOI1 indirectly control the gene expression of MYC transcription factors, possibly by degradation of HbJAZs repressors through SCF-COIl protein complex.(4) Using real time quantitative PCR, the gene expression patterns of HbCoi1 in latex, leaves and bark, or tapping treatment, was analysed. It has been found that HbCoi1 gene was prominently induced by tapping. The expression of HbCoi1 in rubber latex is much higher than that in leaves and bark. The results showed the gene expression pattern of tisuue specicity and induction by mechanical wounding.The study first proved the functional similarity of HbCoi1 with Arabidopsis COIl gene. HbCoi1 was further characterized using yeast two-hybrid system and real time quantitative PCR. The results lay a good foundation for revealing the molecular mechanism of wounding response and latex biosynthesis in Hevea brasiliensis.