Loss of the vacuolar cation channel, AtTPC1, does not impair Ca2+ signals induced by abiotic and biotic stresses

The putative two-pore Ca2+ channel TPC1 has been suggested to be involved in responses to abiotic and biotic stresses. We show that AtTPC1 co-localizes with the K+-selective channel AtTPK1 in the vacuolar membrane. Loss of AtTPC1 abolished Ca2+-activated slow vacuolar (SV) currents, which were increased in AtTPC1-over-expressing Arabidopsis compared to the wild-type. A Ca2+-insensitive vacuolar cation channel, as yet uncharacterized, could be resolved in tpc1-2 knockout plants. The kinetics of ABA- and CO2-induced stomatal closure were similar in wild-type and tpc1-2 knockout plants, excluding a role of SV channels in guard-cell signalling in response to these physiological stimuli. ABA-, K+-, and Ca2+-dependent root growth phenotypes were not changed in tpc1-2 compared to wild-type plants. Given the permeability of SV channels to mono- and divalent cations, the question arises as to whether TPC1 in vivo represents a pathway for Ca2+ entry into the cytosol. Ca2+ responses as measured in aequorin-expressing wild-type, tpc1-2 knockout and TPC1-over-expressing plants disprove a contribution of TPC1 to any of the stimulus-induced Ca2+ signals tested, including abiotic stresses (cold, hyperosmotic, salt and oxidative), elevation in extracellular Ca2+ concentration and biotic factors (elf18, flg22). In good agreement, stimulus- and Ca2+-dependent gene activation was not affected by alterations in TPC1 expression. Together with our finding that the loss of TPC1 did not change the activity of hyperpolarization-activated Ca2+-permeable channels in the plasma membrane, we conclude that TPC1, under physiological conditions, functions as a vacuolar cation channel without a major impact on cytosolic Ca2+ homeostasis.