Annexin 1 regulates the H2O2-induced calcium signature in Arabidopsis thaliana roots

Hydrogen peroxide is the most stable of the reactive oxygen species (ROS) and is a regulator of development, immunity and adaptation to stress. It frequently acts by elevating cytosolic free Ca2+ ([Ca2+](cyt)) as a second messenger, with activation of plasma membrane Ca2+-permeable influx channels as a fundamental part of this process. At the genetic level, to date only the Ca2+-permeable Stelar K+ Outward Rectifier (SKOR) channel has been identified as being responsive to hydrogen peroxide. We show here that the ROS-regulated Ca2+ transport protein Annexin 1 in Arabidopsis thaliana (AtANN1) is involved in regulating the root epidermal [Ca2+](cyt) response to stress levels of extracellular hydrogen peroxide. Peroxide-stimulated [Ca2+](cyt) elevation (determined using aequorin luminometry) was aberrant in roots and root epidermal protoplasts of the Atann1 knockout mutant. Similarly, peroxide-stimulated net Ca2+ influx and K+ efflux were aberrant in Atann1 root mature epidermis, determined using extracellular vibrating ion-selective microelectrodes. Peroxide induction of GSTU1 (Glutathione-S-Transferase1 Tau 1), which is known to be [Ca2+](cyt)-dependent was impaired in mutant roots, consistent with a lesion in signalling. Expression of AtANN1 in roots was suppressed by peroxide, consistent with the need to restrict further Ca2+ influx. Differential regulation of annexin expression was evident, with AtANN2 down-regulation but up-regulation of AtANN3 and AtANN4. Overall the results point to involvement of AtANN1 in shaping the root peroxide-induced [Ca2+](cyt) signature and downstream signalling.