Imaging of the Yellow Cameleon 3.6 Indicator Reveals That Elevations in Cytosolic Ca2+ Follow Oscillating Increases in Growth in Root Hairs of Arabidopsis

Abstract

AbstractIn tip-growing cells, the tip-high Ca2+ gradient is thought to regulate the activity of components of the growth machinery, including the cytoskeleton, Ca2+-dependent regulatory proteins, and the secretory apparatus. In pollen tubes, both the Ca2+ gradient and cell elongation show oscillatory behavior, reinforcing the link between the two. We report that in growing root hairs of Arabidopsis (Arabidopsis thaliana), an oscillating tip-focused Ca2+ gradient can be resolved through imaging of a cytosolically expressed Yellow Cameleon 3.6 fluorescence resonance energy transfer-based Ca2+ sensor. Both elongation of the root hairs and the associated tip-focused Ca2+ gradient show a similar dynamic character, oscillating with a frequency of 2 to 4 min−1. Cross-correlation analysis indicates that the Ca2+ oscillations lag the growth oscillations by 5.3 ± 0.3 s. However, growth never completely stops, even during the slow cycle of an oscillation, and the concomitant tip Ca2+ level is always slightly elevated compared with the resting Ca2+ concentration along the distal shaft, behind the growing tip. Artificially increasing Ca2+ using the Ca2+ ionophore A23187 leads to immediate cessation of elongation and thickening of the apical cell wall. In contrast, dissipating the Ca2+ gradient using either the Ca2+ channel blocker La3+ or the Ca2+ chelator EGTA is accompanied by an increase in the rate of cell expansion and eventual bursting of the root hair tip. These observations are consistent with a model in which the maximal oscillatory increase in cytosolic Ca2+ is triggered by cell expansion associated with tip growth and plays a role in the subsequent restriction of growth.