Functional Analysis Of E3 Ubiquitin Ligases OsUBC26IP2 And OsUBC26IP4 In Rice

Post-translational modification is central to protein stability and activity.The ubiquitin/26S proteasome system constitutes the major protein degradation pathway in the cell.It is an important mechanism for plant to regulate protein turnover for growth,development,and responses to abiotic and biotic stresses.Among the three main kinds of UPS enzymes,E3 ubiquitin ligases are highly flexible and diverse,and each has unique specificity in recognizing substrates.Many studies have indicated that E3s play pivotal roles during plant-pathogen interactions.In previous work,we found that OsUBC26,an ub-conjugating enzyme(E2),is related to the rice disease resistance and OsUBC26IP2,OsUBC26IP4 are the OsUBC26 interacting proteins by yeast two hybrid screening.Bioinformatics analysis shows that both OsUBC26IP2 and OsUBC26IP4 are RING domain containing proteins.Real time PCR assay indicates OsUBC26IP2 and OsUBC26IP4 have differential expression patterns after M.oryzae infection,implying that they may be involved in the interaction between rice and M.oryzae.One by one yeast two hybrid interaction assay indicates that OsUBC26 interacts with OsUBC26IP2 and OsUBC26IP4.Yeast two hybrid screening for OsUBC26IP2 shows that there are 25 interacting proteins of OsUBC26IP2 in rice,including OsUBC26IP2 and OsUBC26IP4.One by one yeast two hybrid assay confirms that OsUBC26IP2 can interact with OsUBC26IP2 and OsUBC26IP4.Therefore,OsUBC26IP2 may function by formation of OsUBC26IP2/2 homo-dimers and OsUBC26IP2/4 hetero-dimers.The over-expression transgenic rice seedlings of OsUBC26IP2 and OsUBC26IP4,and knock-out transgenic rice seedlings of OsUBC26IP2 are generated for further functional analysis.