| Phosphatidylinositol-specific phospholipase C (PI-PLC) is a critical enzyme in various signal-transduction pathways in eukaryotic cells. PI-PLC hydrolyzes phosphatidyl-inositol 4,5-biophosphate (PIP2) into two second messengers, inositol 1,4,5-trisphosphate (IP3) and 1,2-diacyl-glycerol (DAG). PI-PLC-mediated phosphoinositide signaling pathway has been shown to perform important roles in the responses of plant cells to extracellular stimuli, such as osmotic stress, auxin, ABA, light and gravitational force changes, pathogen attack, and pollination. Ca2+ iron is required for plant PI-PLC activities.The Arabidopsis genome contains nine putitived PI-PLC genes. Only the genes corresponding to AtPLC1S, AtPLC1F, AtPLC2 and AtPLC6 have been reported. Further analysis shows that AtPLC1S, AtPLC6 and one gene named as AtPLC5 are found tandem located in a 13 kb fragment on Arabidopsis chromosome V.108 amino acid residues from the N-terminal region of AtPLC5 was expressed as a recombinant protein in E.coli and used in generating polyclonal antibody of AtPLC5. Six other PI-PLC genes including At5g58670 (AtPLC1S), At4g38530 (AtPLC1F), At3g08510 (AtPLC2), At5g58690 (AtPLC6), At3g47220 and At3g47290 had been cloned and expressed as recombinant proteins. Western blot result shows that the antibody only can recognize AtPLC5. The cytosolic fractions and purified plasma membranes were analyzed by western blots using the anti-AtPLC5 antibody. A single band with the similar molecular mass with AtPLC5 was revealed in both cytosolic and plasma membrane fractions.Putative promoter regions of AtPLC5 fused with GUS reporter gene were transferred into Arabidopsis plant. The GUS-staining result shows that the AtPLCS is expressed in the petiole of leaves, anthers and the stigma surface of silique in the transgenic plants. And the expression of AtPLCS is induced to pressed in the vascular tissue of leaves and roots under ABA, NaCl treatments, pathogen infection and cold stress condition.
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