Monitoring single root hairs using a micro-chambered hydroponics system MiCHy reveals a new mode of action of the aminosteroid U73122

Abstract

When using chemical probes and inhibitors for cell biological studies, the uniform and rapid access of the chemical to the sample is key for the accuracy of the analysis. Until now, however, this could only be accomplished by using expensive equipment and labor-intensive and often lengthy procedures. Here, we introduce MiCHy, a micro-chambered hydroponics system, as an accessible and versatile method for chemical treatments and demonstrate its use for imaging roots of Arabidopsis thaliana, the major model system for plant cell biological research. Using lipid biosensors in combination with established chemicals, such as the phosphoinositol 4-kinase (PI4K) inhibitor phenylarsine oxide and the phosphoinositide-specific phospholipase C (PI-PLC) inhibitor U73122, we demonstrate the suitability of MiCHy for studying the relationship between growth and lipid dynamics in root hairs. We further reveal novel effects of U73122 on the plasma membrane localization of phosphatidylinositol-4-phosphate 5-kinase 3 (PIP5K3), which is the first evidence demonstrating a role for U73122 besides its role as a PI-PLC inhibitor in plants.