Expression and Physiological Relevance of Agrobacterium tumefaciens Phosphatidylcholine Biosynthesis Genes

Abstract

Phosphatidylcholine (PC), or lecithin, is the major phospholipid in eukaryotic membranes, whereas only 10% of all bacteria are predicted to synthesize PC. In Rhizobiaceae, including the phytopathogenic bacterium Agrobacterium tumefaciens, PC is essential for the establishment of a successful host-microbe interaction. A. tumefaciens produces PC via two alternative pathways, the methylation pathway and the Pcs pathway. The responsible genes, pmtA (coding for a phospholipid N-methyltransferase) and pcs (coding for a PC synthase), are located on the circular chromosome of A. tumefaciens C58. Recombinant expression of pmtA and pcs in Escherichia coli revealed that the individual proteins carry out the annotated enzyme functions. Both genes and a putative ABC transporter operon downstream of PC are constitutively expressed in A. tumefaciens. The amount of PC in A. tumefaciens membranes reaches around 23% of total membrane lipids. We show that PC is distributed in both the inner and outer membranes. Loss of PC results in reduced motility and increased biofilm formation, two processes known to be involved in virulence. Our work documents the critical importance of membrane lipid homeostasis for diverse cellular processes in A. tumefaciens.

S -Adenosylmethionine-Binding Properties of a Bacterial Phospholipid N -Methyltransferase

“…2B, lane V). As reported previously (26), MMPE, DMPE, and PC were synthesized when wt PmtA was expressed (Fig. 2B, lane WT).…”

“…An A. tumefaciens strain lacking both PC biosynthesis pathways is unable to elicit plant tumors (52). Recently, we produced recombinant PmtA by using Escherichia coli and characterized the enzyme properties in vitro (1,26). PmtA acts as a monomeric enzyme of 22.3 kDa and is inhibited by the end products PC and S-adenosylhomocysteine (SAH).…”

In Vitro Characterization of the Enzyme Properties of the Phospholipid N -Methyltransferase PmtA from Agrobacterium tumefaciens

“…It is possible that this in vitro finding is relevant in vivo, since it might be an elegant way to adjust the proper balance between PE and PC in the bacterial membrane. The PC content in A. tumefaciens membranes is around 23% (24). The relative level of PC does not exceed 40 to 50% of total membrane lipids in B. japonicum membranes (17,30) or 25% in E. coli cells overexpressing Pmt enzymes (17).…”

“…Recombinant A. tumefaciens PmtA is able to convert E. coli PE to PC in the heterologous host (24). Therefore, the activity of purified PmtA was initially tested by using isolated E. coli lipids as the substrate.…”

“…Although PC biosynthesis is restricted to a minority of bacteria, it plays an important physiological role in these microorganisms (15,26). In A. tumefaciens, PC is required for plant infection, stress management, motility, and normal biofilm formation (24,42).…”