Cloning And Functional Analysis Of Genes About Phythphthora Boehmeriae Inducing Non-Host Resistance In Non-Heading Chinese Cabbage

We clonged and functional analysed genes about Phythphthora Boehmeriae inducing non-host resistance in non-heading Chinese cabbage. From these genes, we studied BCLCB₂ which was up-regulated in hypersensitive cell death.We described a suppression subtractive library-based approach to isolate the plant's ESTs upregulated in the early induction/execution of the HR induced by elicitor PB90 from Phytophthora boehmeriae. We analyzed 200 clones, according to their putative identification in BLAST searches against the three Genome Databases, 70 up-regulated genes were classified in 9 parts: some aspect of primary 'metabolism' or 'energy' production; 'protein synthesis' or 'protein fate'; cellular communication/signal transduction mechanism; cell fate, transcription; cellular transport and hypothetical proteins or to proteins for which the function has yet to be determined. And we used RT-PCR to analyse Beclin, thioredoxin, HLA-B, MAP3K, SPT1, SPT2 and AGO1. These genes were up-regulated induced by PB90, suggesting that the genes may play an important role in PB90-triggered HR.We isolated BCLCB₂ from non-heading Chinese cabbage (Brassica campestris ssp. chinensis) encoding the LCB2 subunit of serine palmitoyltransferase (SPT) that is up-regulated during hypersensitive response (HR) induced by elicitor PB90 from Phytophthora boehmeriae. The transcript level of the gene that shares 90% homology with Arabidopsis thaliana is higher in flower and stem than in leaf and root. Overexpression of BCLCB₂ in Nicotiana leaves by Agrobacterium-mediated transient expression suppressed HR initiated by various elicitors such as PB90, elicitin, MgNep1 (from Magnaporthe grisea) and hrf_1xoo (from Xanthomonas oxyzae pv. oryzae). BcLCB₂ also suppressed Bax-mediated HR-like in Nicotiana. Overexpression of BcLCB₂ inhibited H₂O₂ accumulation in Nicotiana triggered by PB90 via DAB staining. Virus inducing NbLCB₂ silencing in N. benthamiana resulted in enhanced hypersensitive cell death triggered by elicitors. In compatible host-pathogen interactions, symptoms in Nicotiana tabacum leaves inoculated of Ralstonia solanacearum were suppressed by overexpression of BcLCB₂. BcLCB₂ also inhibited Bax- and oxidant stress-induced cell death in yeast. Reactive oxygen species (ROS) production triggered by Bax in yeast was inhibited by expression of BcLCB₂. The results demonstrated that BcLCB₂ is a negative regulator in HCD triggered by elicitors in Nicotiana leaves, in cell death during compatible host-pathogen interactions or in Bax-induced cell death in yeast via inhibiting ROS accumulation. The results demonstrated that BcLCB₂ function as cell death attenuator across kingdoms, suggesting signal transduction pathways associated with plant HCD and stress-induced cell death share a common molecular switch.