Ion selectivity in the Chlamydomonas reinhardtii flagellar regeneration system

Extracellular Ca2+ mediates the cellular and molecular responses to cell stimulation of Chlamydomonas reinhardtii. Extracellular Ca2+ concentrations ([Ca2+]e) must exceed certain threshold values to support flagellar excision by acid shock and to stimulate flagellar outgrowth following mechanical shear of the flagella. Also, the magnitude and duration of flagellar RNA accumulations following acid shock or mechanical shear increase with increasing [Ca2+]e. To better understand the role of Ca2+ in flagellar excision, RNA induction, and outgrowth, we have performed a survey of the ion selectivity of each of these responses to acid shock. We found that flagellar excision in vivo following acid shock was supported by Sr2+ and Ca2+, but no other ion tested. LaCl3 and neomycin prevented flagellar excision following acid shock of cells in Ca2+- or Sr2+-containing buffer. Sr2+ addition to detergent-permeabilized cell models, however, failed to elicit flagellar excision in vitro. Cells failed to regrow flagella following flagellar excision in Sr2+-containing buffer unless exogenous Ca2+ was added. Flagellar RNA accumulations of lower magnitude and shorter duration were measured in cells acid-shocked in Sr2+-containing buffer than in Ca2+-containing buffer. These results demonstrate that a Sr2+ influx can evoke flagellar excision following acid shock, but cannot directly activate the machinery for flagellar excision, suggesting that a Sr2+ influx induces excision by stimulating an intracellular Ca2+ release. Furthermore, they suggest that flagellar outgrowth and normal flagellar RNA induction have a strict requirement for Ca2+, which is not satisfied by the proposed intracellular Ca2+ release.