Journal Article

Potential Role of Annexin AnnAt1 from <i>Arabidopsis thaliana</i> in pH-Mediated Cellular Response to Environmental Stimuli

Karolina M. Gorecka, Cyril Thouverey, Rene Buchet and Slawomir Pikula

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 48, issue 6, pages 792-803
Published in print June 2007 | ISSN: 0032-0781
Published online June 2007 | e-ISSN: 1471-9053 | DOI: http://dx.doi.org/10.1093/pcp/pcm046
Potential Role of Annexin AnnAt1 from Arabidopsis thaliana in pH-Mediated Cellular Response to Environmental Stimuli

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Plant annexins, Ca2+- and membrane-binding proteins, are probably implicated in the cellular response to stress resulting from acidification of cytosol. To understand how annexins can contribute to cellular ion homeostasis, we investigated the pH-induced changes in the structure and function of recombinant annexin AnnAt1 from Arabidopsis thaliana. The decrease of pH from 7.0 to 5.8 reduced the time of the formation of ion channels by AnnAt1 in artificial lipid membranes from 3.5 h to 15–20 min and increased their unitary conductance from 32 to 63 pS. These changes were accompanied by an increase in AnnAt1 hydrophobicity as revealed by hydrophobicity predictions, by an increase in fluorescence of 2-(p-toluidino)naphthalene-6-sulfonic acid (TNS) bound to AnnAt1 and fluorescence resonance energy transfer from AnnAt1 tryptophan residues to TNS. Concomitant lipid partition of AnnAt1 at acidic pH resulted in its partial protection from proteolytic digestion. Secondary structures of AnnAt1 determined by circular dichroism and infrared spectroscopy were also affected by lowering the pH from 7.2 to 5.2. These changes were characterized by an increase in β-sheet content at the expense of α-helical structures, and were accompanied by reversible formation of AnnAt1 oligomers as probed by ultracentrifugation in a sucrose gradient. A further decrease of pH from 5.2 to 4.5 or lower led to the formation of irreversible aggregates and loss of AnnAt1 ionic conductance. Our findings suggest that AnnAt1 can sense changes of the pH milieu over the pH range from 7 to 5 and respond by changes in ion channel conductance, hydrophobicity, secondary structure of the protein and formation of oligomers. Further acidification irreversibly inactivated AnnAt1. We suggest that the pH-sensitive ion channel activity of AnnAt1 may play a role in intracellular ion homeostasis.

Keywords: Annexins; Arabidopsis thaliana; Environmental response; Ion homeostasis; pH-induced ion channels

Journal Article.  6746 words.  Illustrated.

Subjects: Biochemistry ; Molecular and Cell Biology ; Plant Sciences and Forestry

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