| Rice (Oryza sativa L.) is not only the most important crop of China, butalso the model plant for the monocots. The accomplishment of rice genomesequencing project declare that the era of rice functional genomics is right onthe horizon. Proteomics, the large-scale analysis of proteins, will contributegreatly to our understanding of gene function in the post-genomic era.Through optimizing the rice protein extraction method andelectrophoresis conditions, a large-scale, high-resolution and reproducible2-DE analysis systerm was established based on the technical platform ofBio-Rad large-scale 2-DE system. At least 1000 protein spots were detected onthe gels of rice leaf. The correlation coefficient between two replicates is0.964.Bacterial blight, caused by Xanthomonas oryzae pv. oryzae (Xoo), is themost serious bacterial disease of rice worldwide. It is an ideal model systemfor the study of the interaction between plants and their bacterial pathogens.The bacterial blight resistance trait from Oryza meyeriana, a wild ricespecies, was introduced into an elite japonica rice cultivar using asymmetricsomatic hybridization. This study was carried out by using differentialproteomic analyses with the intention of understanding the molecularmechanism of incompatible interaction between Xoo and the stable somatichybrids. Proteins were extracted from leaves at 24, 48, and 72 hatter Xooinoculation and separated by high resolution 2-DE. Then, the differentially expressed proteins identified by large-scale mass spectrometry. The results aresummarized as follows:1. Forty-eight hour may be the pivotal time point for the incompatibleinteraction between rice and Xoo in protein level. Quantitative image analysisrevealed that a total of 77 protein spots changed their intensities significantly (p＜0.05) by more than 1.5-fold at least at one time point. Seventeen spots weredifferentially expressed at all three time points, 8 at both 24 h and 48 h, 20 atboth 48 h and 72 h, 31 only at 48 h and 1 only at 24 h. Among 77 protein spots,76 were differentially expressed at 48 h time point.2. Three novel proteins were found involved in the pathogen response. Thefirst novel protein, which belongs to Ser/Thr kinase family, is phytosulfokinereceptor precursor (PSKR), containing leucine rich-repeat region (LRR) andSer/Thr kinase domain (STK). It is located on the membrane, which can triggerthe signal cascade through binding phytosulfokine. The second is thephosphoinositide 3-kinase-related protein kinase (PIKK). The PIKKs comprisea family of signaling proteins that play central roles in the control of cell growth,gene expression genome surveillance and repair in eukaryotic cells. The PIKKfamily of proteins has a key role in many of the stress responses of theeukaryotic cell. The third one is auxin-regulated protein. Sequence analysisindicated that this protein contains two repeats of the N-terminal domain ofGSTs.3. Chloroplast may be the organelle mostly affected by Xoo infection. Using TargetP program, 15 of the identified proteins were predicted to belocated in the chloroplast. Up-regulation of Rubisco large subunit (RcbL)fragments indicated that RcbL degraded following Xoo attack, which wasfurther confirmed by Western Blot analysis.We first applied proteome method to detect protein exppression changes ofthe stable somatic hybrids responding to Xoo. The present study revealed thatthere existed a complex cellular network in the leaf cells of the stable somatichybrids in response to the bacterial pathogen infection. The network coveredphotosynthesis, antioxidant defense, protein turnover, signal transduction, andmetabolisms. Of the 64 identifed protein spots, 23 were predicted to havespecific subcellular localization. Fifteen and eight protein spots were suggestedto be located in chloroplast and mitochondria, respectively. Thus, a full pictureof the bacterical pathogen induced response will require subcellular proteomicanalyses so that the role of particular protein isoforms and organelles will beclarified. And further research of the three novel proteins will contribute to theunderstanding the pathogen resistance mechanism of the stable somatic hybrids.
|
PDF Full Text Request