Lipid kinases PIP5K7 and PIP5K9 are required for polyamine-triggered K+efflux in Arabidopsis roots

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 (PIP2), and that this increase is required for polyamine effects on K(+)efflux in Arabidopsis roots. Usingin vivo(32)P(i)-labelling of Arabidopsis seedlings, low physiological (mu m) 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 identifiedPIP5K7andPIP5K9as 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 inpip5k7 pip5k9seedlings. Together, our results provide biochemical and genetic evidence for a physiological role of PIP(2)in polyamine-mediated signalling controlling K(+)flux in plants.