The Arabidopsis Phospholipase D Family. Characterization of a Calcium-Independent and Phosphatidylcholine-Selective PLD 1 with Distinct Regulatory Domains

Four types of phospholipase D (PLD), PLD , , , and , have been characterized in Arabidopsis, and they display different requirements for Ca , phosphatidylinositol 4,5-bisphosphate (PIP2), substrate vesicle composition, and/or free fatty acids. However, all previously cloned plant PLDs contain a Ca -dependent phospholipid-binding C2 domain and require Ca for activity. This study documents a new type of PLD, PLD 1, which is distinctively different from previously characterized PLDs. It contains at the N terminus a Phox homology domain and a pleckstrin homology domain, but not the C2 domain. A full-length cDNA for Arabidopsis PLD 1 has been identified and used to express catalytically active PLD in Escherichia coli. PLD 1 does not require Ca or any other divalent cation for activity. In addition, it selectively hydrolyzes phosphatidylcholine, whereas the other Arabidopsis PLDs use several phospholipids as substrates. PLD 1 requires PIP2 for activity, but unlike the PIP2-requiring PLD or , phosphatidylethanolamine is not needed in substrate vesicles. These differences are described, together with a genomic analysis of 12 putative Arabidopsis PLD genes that are grouped into , , , , and based on their gene architectures, sequence similarities, domain structures, and biochemical properties.