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Zea mays Annexins Modulate Cytosolic Free Ca²⁺ and Generate a Ca²⁺-Permeable Conductance^[W]

^a Department of Plant Sciences, University of Cambridge, Cambridge CB2 3EA, United Kingdom
^b Marine Biological Association, Plymouth PL1 2PB, United Kingdom
^c Structural Chemistry Program, Eskitis Institute for Cell and Molecular Therapies, Griffith University, Nathan QLD 4111, Australia
^d BioMaDe, 9747 AG, Groningen, The Netherlands
^e School of Biological Sciences, University of Reading, Whiteknights, Reading RG6 6AS, United Kingdom

⁵ Address correspondence to jmd32{at}cam.ac.uk

Regulation of reactive oxygen species and cytosolic free calcium ([Ca²⁺]_cyt) is central to plant function. Annexins are small proteins capable of Ca²⁺-dependent membrane binding or membrane insertion. They possess structural motifs that could support both peroxidase activity and calcium transport. Here, a Zea mays annexin preparation caused increases in [Ca²⁺]_cyt when added to protoplasts of Arabidopsis thaliana roots expressing aequorin. The pharmacological profile was consistent with annexin activation (at the extracellular plasma membrane face) of Arabidopsis Ca²⁺-permeable nonselective cation channels. Secreted annexins could therefore modulate Ca²⁺ influx. As maize annexins occur in the cytosol and plasma membrane, they were incorporated at the intracellular face of lipid bilayers designed to mimic the plasma membrane. Here, they generated an instantaneously activating Ca²⁺-permeable conductance at mildly acidic pH that was sensitive to verapamil and Gd³⁺ and had a Ca²⁺-to-K⁺ permeability ratio of 0.36. These results suggest that cytosolic annexins create a Ca²⁺ influx pathway directly, particularly during stress responses involving acidosis. A maize annexin preparation also demonstrated in vitro peroxidase activity that appeared independent of heme association. In conclusion, this study has demonstrated that plant annexins create Ca²⁺-permeable transport pathways, regulate [Ca²⁺]_cyt, and may function as peroxidases in vitro.

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