Abstract
Polyamines, such as putrescine, spermidine and spermine (Spm), are low-molecular-weight polycationic molecules present in all living organisms. Despite their implication in plant cellular processes, little is known about their molecular mode of action. Here, we demonstrate that polyamines trigger a rapid increase in the regulatory membrane lipid phosphatidylinositol 4,5-bisphosphate (PIP(2) ), and that this increase is required for polyamine effects on K(+) efflux in Arabidopsis roots. Using in vivo (32) P(i) -labelling of Arabidopsis seedlings, low physiological (μm) concentrations of Spm were found to promote a rapid PIP(2) increase in roots that was time- and dose-dependent. Confocal imaging of a genetically encoded PIP(2) biosensor revealed that this increase was triggered at the plasma membrane. Differential (32) P(i) -labelling suggested that the increase in PIP(2) was generated through activation of phosphatidylinositol 4-phosphate 5-kinase (PIP5K) activity rather than inhibition of a phospholipase C or PIP(2) 5-phosphatase activity. Systematic analysis of transfer DNA insertion mutants identified PIP5K7 and PIP5K9 as the main candidates involved in the Spm-induced PIP(2) response. Using non-invasive microelectrode ion flux estimation, we discovered that the Spm-triggered K(+) efflux response was strongly reduced in pip5k7 pip5k9 seedlings. Together, our results provide biochemical and genetic evidence for a physiological role of PIP(2) in polyamine-mediated signalling controlling K(+) flux in plants.