Cloning and expression of AtPLC6, a gene encoding a phosphatidylinositol-specific phospholipase C in Arabidopsis thaliana
Chinese Science Bulletin
A full-length cDNA clone corresponding to a putative phosphatidylinositol-specific phospholipase C (PI-PLC) was isolated from Arabidopsis thaliana by screening a cDNA library and using RT-PCR strategy. The cDNA, designated AtPLC6, encodes a putative polypeptide of 578 amino acid residues with a calculated molecular mass of 66251.84 D and a pI of 7.24. The sequence analysis indicates that the polypeptide contains X, Y, EF-hand and C2 domains. The overall structure of putative AtPLC6 protein,
... AtPLC6 protein, like other plant PI-PLCs, is most similar to that of mammalian PLC . The recombinant AtPLC6 protein expressed in E. coli was able to hydrolyze phosphatidylinositol 4,5-biophosphate (PIP2) to generate inositol 1,4,5-trisphate (IP3) and 1,2-diacylglycerol (DAG). The protein hydrolyzes PIP2 in a Ca 2+ -dependent manner and the optimum concentration of Ca 2+ is 10 mol/L. These results suggested that AtPLC6 gene encodes a genuine PI-PLC. Northern blot analysis showed that the AtPLC6 gene is expressed at low level in all examined tissues, such as roots, stems, leaves, flowers, siliques and seedlings under normal growth conditions. The gene is strongly induced under low temperature and weakly induced under various stresses, such as ABA, high-salt stress and heat. These results suggested that AtPLC6 might be involved in the signal-transduction pathways of cold responses of the plants.