| Rice stripe disease caused by Rice stripe virus(RSV)is one kind of important virus diseases. This disease occurs widely in our country and has caused great losses to rice yields. The use of resistant cultivars is the most economical and environmentally sound method to reduce the damage. Therefore, understanding the mechanisms underlying disease-resistance would be very important for improving disease resistance in rice. Transcription profiling of Wuyujing 3 (susceptible variety) and KT95-418 (resistant variety) conducted previously in our lab showed that some early auxin-responsive genes expressed differentially in RSV infected rice plants, implicating the involvement of the auxin signal pathway in rice-RSV interaction.Auxin exerts pleiotropic effects on plant growth and development by regulating the expression of early auxin-responsive genes including auxin/indoleacetic acid (Aux/IAAs). These genes have been studied extensively in dicots like Arabidopsis. But in rice,the role of Aux/IAA in auxin signaling and their functions in the host–pathogen interactions have not been clearly elucidated. In this study, OsIAA6, a rice gene that was shown to be differentially expressed during RSV infection in the susceptible variety by our transcriptom studies, was selected and functionally studied.The expression changes of OsIAA6 were validated by real-time RT-PCR. The expression was constantly uprgulated in rice susnsion culcure cells infected by RSV. However, only a transient up-regulation at 4 d after RSV infection in rice plants was detected and the expression pattern at 16 d after RSV infection were consistent with the result of microarray analysis. The samples investigated in microarray were collected at about 16 d after RSV infection.To investigate its function, OsIAA6 gene was cloned by RT-PCR and fused with EGFP to construct the transformation vector pEGAD-OsIAA6. On confocal laser-scanning microscopy and ordinary fluorescence microscopes, green fluorescent signals were localized in nucleus in tobacco leaf cells and onion epidermal cells upon Agrobacterium-mediated transformation and transent expression. The results suggest that OsIAA6 encodes a nuclear protein.We selected the plant expression vector pKYLX and used Agrobacterium-mediated leaf disc transformation to get transgenic tobacco K326,which overexpressed OsIAA6 gene. PCR analysis revealed that a total of 11 transgenic plants were obtained. The differentiations of 4 plants stopped after application on of rooting culture and the other 7 plants were slow-growing compared with wild type plants. Further analysis of TMV infection and replication in transgenic plants and wild-type plants revealed significant difference. TMV replication in transgenic plants was a few times more than that in the the wild-type plants. The reason was unclear now.To further disclose the function of rice OsIAA6, we also selected vector pKYLX to construct OsIAA6 sense and anti-sense expression vectors and transferred them into rice by Agrobacterium- mediated transformation. We obtained the OsIAA6 downregulated and overexpressed rice lines. The rice callus of sense OsIAA6 overexpessing plants could hardly differentiate. And only 1 transgenic plant was obtained. But it showed a severe defect in development and growth similar to transgenic tobacco. Whereas 50 plants expressing anti-sense OsIAA6 were gained and they displayed high rate of differentiation, elongation of plant height, increased panicle length and number of tillers. This implied that rice OsIAA6 plays an important regulatory role in plant growth. |
