Functional characterization of phospholipase Dzeta1 and zeta2 in root development and in response to phosphate deficiency in Arabidopsis thaliana

An electrospray ionization tandem mass spectrometry (ESI-MS/MS) based method was established to profile plant polar glycerolipids, including phospholipids, lysophospholipids, and galactolipids. This approach was applied to characterize the gene function of phospholipases in this study.; Phospholipase D (PLD) hydrolyzes phospholipids into phosphatidic acid (PA) and a head group. There are 12 PLD genes in Arabidopsis, including PLDzeta1 and PLDzeta2 . Gene expression analysis by real time PCR showed that PLDzeta1 and PLDzeta2 have higher transcriptional levels in roots and old leaves than in other tissues. This study investigated the roles of PLDzeta1 and PLDzeta2 in root-related functions and in response to the senescence-related phytohormone, ethylene.; Knockout mutant analysis revealed that permanent disruption of PLDzeta1 and PLDzeta2 did not impair root hair differentiation and root growth under normal growth conditions. Disruption of both PLDzeta1 and PLDzeta2 retarded primary root growth and promoted lateral root growth during deficiency of phosphorous, an essential element for plant growth and development. Disruption of PLDzeta1 or PLDzeta2 alone did not affect root growth. These results suggest that PLDzeta1 and PLDzeta2 function to regulate root growth during phosphate deficiency.; Without phosphate in the culture media, disruption of both PLDzeta1 and PLDzeta2 resulted in a higher level of phosphatidylcholine (PC) and a lower level of digatactosyldiacylglycerol (DGDG) in roots; this was not the case when either PLDzeta1 or PLDzeta2 was disrupted alone. These results suggest that PLDzeta1 and PLDzeta2 function in lipid turnover that produces galactolipids at the expense of phospholipids when phosphate is severely limited.; The PLDzeta1 mutant and PLDzeta2 mutant did not show any apparent phenotype upon ethylene treatment, and their responses were similar to wild type in terms of lipid levels. The PA level was increased in leaves of wild type and PLDzeta mutants in response to ethylene treatment. The interaction of PA and a putative PA binding motif of CONSTITUTIVE TRIPLE RESPONSE1, an essential component in the ethylene signaling pathway, was characterized by circular dichroism spectrometry.