The Function Of LeEBFs, A Component Of Ethylene A Signal Pathway, Which Interacted With LeEIN3

Ethylene, a gaseous phytohormone, regulates diverse developmental and physiological processes throughout the entire life cycle of plants, including seed germination, root initiation, flower and leaf senescence, abscission, fruit ripening, wounding response, and disease defense. Now the researches on ethylene signal transduction pathway and the regulation mechanism have received considerable attention in plant molecular biology. Potuschak T and Guo et al cloned two Arabidopsis F-box proteins, EIN3 binding F-box protein 1 (EBF1) and 2 (EBF2), which targeted the transcriptional activator, ethylene insensitive 3 (EIN3) protein for degradation. The mRNA of EIN3 is transcribed constitutively, but the EIN3 protein is unable to accumulate for being subjected to permanent proteolysis mediated by EBF1 and EBF2. The SCF complexes (SCF-EBF1/2) target EIN3 for ubiquitination and subsequent degradation. Double ebf1/ebf2 mutants display a severe growth inhibition after germination and have heightened levels of EIN3 even in the absence of exogenous ethylene. It suggests that EBF1 and EBF2 play an important role in regulating ethylene response and plant growth. Study on LeEBF1 and LeEBF2 will make us better understand the mechanism of ripening-related genes expression regulated by ethylene and further elucidate ethylene transduction pathway in tomato. The main results are as follows:â‘ Two full-length cDNAs of LeEBF1 and LeEBF2 were cloned from tomato (Solanum lycopersicum) by RT-PCR and rapid amplification of cDNA ends (RACE). They were 2866 and 2891 bp, with open reading frame (ORF) of 1911 bp and 1995 bp, encoding 637 and 665 amino acids, respectively. Putative LeEBF1 and LeEBF2 proteins, each containing a well-conserved F-box motif in the amino terminus and 14 tandem leucine-rich repeats (LRRs) in the carboxyl terminus, shared 58.6% of identity with each other. Identities with other species EBF proteins within F-box motif ranged from 24.4% to 73.2%.â‘¡RNA gel blot analysis results indicated that the expression levels of LeEBF1 and LeEBF2 in young leaves were higher than those in mature leaves of wild type and Nr. During the development of fruits, LeEBF1 and LeEBF2 expressed relatively weaker at Immature Green (IMG) stage than at other stages experimented.â‘¢Target genes LeEBF1 and LeEBF2 were cloned into RNAi expression vector, and then transformed to Nr tomato. The result showed that 10 transgenic lines were obtained. It provides a basis to approach the function of LeEBF and LeEBF2 in tomato.