Response Of Phloem Sap MRNA And Proteins To Brassinosteroids In Cucumis Sativus

Brassinosteroids(BRs), a new kind of steroidal hormones play important roles not only in plant growth and development but also in respons to biotic and abiotic stresses. The plant long-distance translocation systems played a critical role in the development of an effective whole-plant circulatory and communication network. Studying the response of phloem sap mRNAs and proteins to BRs may be helpful to reveal the roles of phloem information network in cooperating plant growth, development and stress tolerance between aerial and under-ground organs. The study is also of significant importance for developmenting methods to improve systemically the whole-plant stress resistance.Cucumber (Cucumis sativus L.) was used to study the BRs-regulated mRNAs and proteins in phloem sap. Through the establishment of suppression subtractive library interrogated by bioinformatic tools, those transcripts specifically expressed in response to EBR in the metabolic network of cucumber were indentified. The reliability of the suppression subtractive library was also verified by using RT-PCR method. Meanwhile, changes in proteome of phloem sap after EBR treatment were studied by 2D-PAGE. The findings are as follows:1) A cDNA library of genes which were preferentially expressed in the EBR-treated phloem sap was constructed by suppression subtractive hybridization (SSH) to study the effects of BRs on the transcripts in cucumber phloem sap, 680 clones were obtained,and 260 clones were randomly selected for sequencing, resulting in 28 unique transcripts and fall them into seven categories after analyzed by BLASTX and Go: energy generation and photosynthesis, transportation channels, transport and synthesis of protein, cytoskeleton, control of cell division, hormone response and regulation of stress response. The high occurrence of stress related genes in the SSH cDNA library points to an important roles of BRs in stress response. The appearance of auxin and gibberellin response factor genes in the library suggests that the interactions between BRs and other hormones may exist.2) RT-PCR verified that EBR foliar application mRNA levels increased significantly for those genes: C60,C453,C568,C681,C579,C594,C564,C623,C637,C420, this prove that EBR promote this genes expression. 3) The two-dimensional polyacrylamide gel electrophoresis of cucumber phloem protein revealed that 6 h after 0.1 mg L^-1 EBR foliar application the abundance of a 28 KDa protein was upregulated and three other protein (60KDa,75 KDa,95 KDa) were downregulated by EBR. This suggests that EBR induce synthesis of some related proteins in cucumber leaves and transport to other organs through the phloem translocation stream.In conclusion, EBR influences the expression of RNA and protein in cucumber phloem sap. We hypothesize that BRs regulate systemically the development and stress resistance of distant organs by mRNA and protein trafficking through the phloem.