Plant and Cell Physiology Advance Access published online on April 22, 2007
Plant and Cell Physiology, doi:10.1093/pcp/pcm046
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Potential role of annexin AnnAt1 from Arabidopsis thaliana in pH-mediated cellular response to environmental stimuli
1Department of Cellular Biochemistry, Nencki Institute of Experimental Biology, Polish Academy of Sciences, 3 Pasteur Street, PL-02093 Warsaw, Poland
2Université Lyon 1, UMR CNRS 5013, UFR de Chimie-Biochimie, 43 Boulevard du 11 Novembre 1918, F-69622 Villeurbanne, France
Corresponding author: Dr. Slawomir Pikula. Department of Cellular Biochemistry, Nencki Institute of Experimental Biology, Polish Academy of Sciences, 3 Pasteur Street, 02-093 Warsaw, Poland. phone: + 4822-589-2347, FAX: + 4822-822-5342, E-mail: s.pikula{at}nencki.gov.pl
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Abstract |
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Plant annexins, Ca2+- and membrane-binding proteins, are likely implicated in 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 A. 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 pS 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 pH from 7.2 to 5.2. These changes were characterized by an increase in ß-sheet content at the expense of 
-helix structures, and were accompanied by reversible formation of AnnAt1 oligomers as probed by ultracentrifugation in 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 by 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 – pH-induced ion channels - ion homeostasis - environmental response - Arabidopsis thaliana
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