An interconnection between tip-focused Ca²⁺ and anion homeostasis controls pollen tube growth

Plant reproduction is the basis for economically relevant food production. It relies on pollen tube (PTs) growth into the female flower organs for successful fertilization. The high cytosolic Ca²⁺ concentration ([Ca²⁺]_cyt) at the PT tip is sensed by Ca²⁺-dependent protein kinases (CPKs) that in turn activate R- and S-type anion channels to control polar growth. Lanthanum, a blocker for plant Ca²⁺-permeable channels was used here to demonstrate a strict dependency for anion channel activation through high PT tip [Ca²⁺]_cyt. We visualized this relationship by live-cell anion imaging and concurrent triggering of Ca²⁺-elevations with the two-electrode voltage-clamp (TEVC) technique. The anion efflux provoked by a TEVC-triggered [Ca²⁺]_cyt increase was abolished by Lanthanum and was followed by an overall rise in the cytosolic anion concentration. An interrelation between Ca²⁺ and anion homeostasis occurred also on the transcript level of CPKs and anion channels. qRT-PCR analysis demonstrated a co-regulation of anion channels and CPKs in media with different Cl^- and NO₃^- compositions. Our data provides strong evidence for the importance of a Ca²⁺-dependent anion channel regulation and point to a synchronized adjustment of CPK and anion channel transcript levels to fine-tune anion efflux at the PT tip.