The Correlation Study Between SYP71, ROP2 And Auxin Rapidly Responsive Regulation Of Vesicular Transport

The auxin as a kind of phytohormones plays a crucial role in plant development and morphogenesis. Vesicular transport is a process by which specific cargos are transported to the target through vesicle trafficking to maintain homeostasis in cells and promote cell development. Higher plant cells relay on it's characteristic vesicle trafficking mechanism to accomplish material communication and signal transduction. Actually the formation of auxin polar distribution and auxin signal transduction depend on vesicular transport regulating auxin transport proteins and their receptors distribution in the membrane of the cell. The SYP71, a Qc-SNARE (Soluble N-Ethylmaleimide Sensitive Factor Attachment Protein Receptor) only found in plant, play an essential role in the process of secretions regulation. And the ROP2 (Rho-related GTPases from plant), a member of the small G protein Rho family, are also only found in plant and involved in cell signal transduction as well.In order to investigate the auxin signal response in regulating the cell vesicle transport, we cloned the cDNA of SYP71 from Arabidopsis and constructed the over expression (OX-SYP71) and antisense expression (anti-SYP71) vectors, and then transformed into membrane labeled BY-2 cell lines in which the expression auxin binding protein 1 (ABP1) is inducible. SYP71 over expression activated secretions in BY2 cells. During the time treated with IAA for 0-3min, many secretion vesicle distribute formed, and we deduced that ABP1 was transformed from the endoplasmic reticulum to the plasma membrane to act as auxin receptor with the budding when cell stimulated by exogenous auxin signal. However, the expression of anti-SYP71 inhibited the vesicular transport, and fluorescence mainly observed in endoplasmic reticulum surrounding the nucleus, which is in the initial stage of vesicular transport. The regulation of vesicular transport by SYP71 was in conformity to ABP1, vesicular exocytosis were obliviously enhanced in ABP1 over expression BY2 cells while suppressed in ABP1 inhibited BY2 cells, and the filose transport network in cytoplasm obviously decreased when ABP1 inhibited.Meanwhile, the retransformed ROP2 over expression (OX-ROP2), constitutively active expression (CA-rop2) and the dominant negative (DN-rop2) BY2 cells in which the expression auxin binding protein 1 (ABP1) was inducible and membrane labeled were treated and observed. With the treatment of IAA that the over expression of ROP2 and the expression of constitutively active CA-rop2 the cell vesicular exocytosis were obliviously enhanced in the cells. The vesicular exocytosis became even more active when ABP1 was induced to over expression previously. However, the expression of dominant-negative DN-rop2 inhibited the vesicular transport, even if the ABP1 was induced to over expression previously. The promotion of vesicular transport by OX-ROP2 and CA-rop2 was suppressed in ABP1 inhibited BY2 cells. Time-lapse observation suggested that vesicular transport induced by auxin occurred within 2 minutes only. When the cells were treated with a grade IAA, the vesicular transport from nucleus membrane to cytoplasm systems and then to plasma membrane appeared active and moved to auxin higher concentration side in CA-rop2 and ABP1 over expressed BY2 cells. These results indicate that the rapid response to auxin signal in plant is directly related to vesicular transport regulation. Synthetically, ROP2 is involved in the ABP1 mediated vesicular transport response in plant cell and accelerate vesicular transported to auxin higher concentration side, which lead to cell polar formation. Thus we deduced that the plasma membrane localised ABP1, acting as a auxin receptor, transduce the auxin signal to some kinase and activate ROP2 when combined with IAA, then ROP2 activates the G protein directly related to vesicular transport as well as SNARE (SYP71), thereby enliven the vesicular transport.