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. 2006 Mar 27;172(7):991–998. doi: 10.1083/jcb.200508116

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Copyright © 2006, The Rockefeller University Press

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Figure 5. — The Ca²⁺ sensor AtCBL1 interacts with NH₂-terminal domains of PI-4Kβ1. (A) Yeast two-hybrid interaction between AtCBL1 and the NH₂-terminal PI-4Kβ1 fragment (ΔC567-1121; Fig. 1 B) was observed on high-stringency media (−HisTrpLeu [HTL] + 3-AT). (B–D) Disruption of tip-focused Ca²⁺ gradient in root hairs abolished growth and tip-localized EYFP-RabA4b. (B) EYFP-RabA4b fluorescence was visualized in root hairs using time-lapse fluorescence microscopy. Upon treatment with the Ca²⁺ ionophore A23187 (20-min time point), root hair elongation was rapidly inhibited. This correlated with loss of EYFP-RabA4b tip localization and observation of EYFP-RabA4b along the entire root hair (24–30-min time points). When A23187 was washed out, all EYFP-RabA4b fluorescence was lost from the root hair. Neither EYFP-RabA4b tip localization nor root hair tip growth occurred after A23187 washout. The length of the root hair (C) and the percentage of tip fluorescence (D) were measured over time.