| Ubiquitin/proteasome system plays an important role in plant development,morphogenesis and disease resistance.Ubiquitin-conjugating enzyme is the second enzyme participates in the ubiquitination process and mediated the transfer of ubiquitin from E1 to substrate or E3.E2 determines the topology of ubiquitin chain,so as the destiny of the ubiquitinated protein.In this study,bioinformatics,qRT-PCR,yeast twohybrid were used to analyze the expression pattern of rice ubiquitin-conjugating enzyme genes during rice-Magnaporthe oryzae interaction and screened out the interacting proteins of OsUBC26 in rice.Phylogenetic tree analysis showed that 48 rice ubiquitin-conjugating enzyme genes can divided into 3 group,9 sub-group.OsUBC26 and OsUBC25 have highest homology,which may produced by duplication during the evolutionary process.Protein domain analysis showed that OsUBC26 mainly consist of an ubiquitin-conjugating enzyme catalytic domain,without N-terminal and C-terminal extension,which belong to class I ubiquitin-conjugating enzyme.Plant cis-acting elements analysis in the promoter region indicates that one hypersensitivity response cis-acting element and four pathogen resistance cis-acting elements have higher distribution in the promoter region of rice ubiquitin-conjugating enzyme genes.qRT-PCR analysis showed that some ubiquitin-conjugating enzyme gene’s expression can be induced by the inoculation of rice blast fungus both in compatible and incompatible monogenic rice.The use of BDOsUBC26 as bait to screening rice cDNA library received a total of 19 OsUBC26 candidate interacting proteins,including six putative E3 ligase protein,an U-box domain containing E3 ligase protein,a putative deubiquitination enzyme.Up to now,14 out of 19 candidate genes have been successfully cloned,among which an E3 ubiquitin ligase was constructed into AD vector and co-transform with BD-OsUBC26 into yeast which can reappear the interaction phenotype.Bimolecular fluorescence complementation experiment further confirmed the interaction between OsUBC26 and this E3 ligase.In short,this study lays a strong foundation for functional analysis of rice ubiquitin-conjugating enzyme genes,especially OsUBC26 in rice-Magnaporthe oryzae interaction. |
