Regulation of plant phospholipid biosynthesis

Phospholipids are essential for the construction of eukaryotic cell membranes, which play a fundamental role in compartmentalising the biochemistry of life. The quantity and composition of phospholipids are tightly regulated during growth and development, and in response to environmental change, so that membranes always maintain their structure and function. Research in mammals and yeast (Saccharomyces cerevisiae) has uncovered elegant metabolite signaling mechanisms, both transcriptional and post-translational, that allow the cell to sense changes in key lipid intermediates and adjust phospholipid synthesis (and turnover) accordingly. Analogous mechanisms are also likely to exist in plants but surprisingly they have not been elucidated. Because phospholipids are essential, genetic analysis of their regulation through loss-of-function is problematic. However, we have recently isolated an Arabidopsis thaliana double mutant in two phosphatidate phosphatases (pah1 pah2) that produces approximately twice as much phospholipid in its leaves as wild type plants. To our knowledge this is the first plant mutant to over-produce phospholipids and the gain-of-function phenotype provides a unique tool. The objective of this grant proposal is to use the pah1 pah2 mutant (and corresponding genes) to discover how phospholipid biosynthesis is regulated in Arabidopsis and to investigate how it is coordinated with cell cycle progression, which requires membrane biogenesis. This discovery will be of fundamental scientific interest, particularly as there is already evidence to show that many key elements of the regulatory mechanism(s) in plants must differ from those described in mammals or yeast.

In yeast (Saccharomyces cerevisiae) the Mg2+-dependent phosphatidate phosphatase PAH1 acts as a repressor of phospholipid biosynthetic gene expression by controlling the level of the signalling molecule phosphatidic acid (PA). Our data show that in Arabidopsis thaliana two PAH proteins (PAH1&2) also govern the global rate of phospholipid synthesis at the ER. PAH is therefore a conserved 'node' in the phospholipid regulatory networks of Arabidopsis and yeast. However, despite this commonality the signal transduction pathways that couple PAH to phospholipid synthesis must be very different for two fundamental reasons. (i) Arabidopsis lacks homologues of the phospholipid regulatory genes OPI1, INO2 and INO4, which allow yeast to sense and respond to PA. (ii) Arabidopsis and yeast synthesise the bulk of their phospholipids by different metabolic pathways. The objective of this grant proposal is therefore to discover how the PAH-dependent signal transduction pathway works in Arabidopsis and also whether the cyclin-dependent protein kinase CDKA;1 is involved in coordinating phospholipid synthesis with the cell cycle by phosphorylating PAH1&2.