Abstract
There is evidence that Ca2+ can regulate vesicle-mediated secretion in plant cells, but the mechanism for this is not known. One possibility is that Ca2+ -dependent phospholipid-binding proteins (annexins) couple the Ca2+ stimulus to the exocytotic response. Using a protocol developed for the isolation of animal annexins we have identified proteins in maize (Zea mays L.) coleoptiles that have similar characteristics to annexins. The predominant polypeptide species run as a doublet of relative molecular mass (Mr) 33000–35000 on sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE); another less-abundant protein of Mr 23000 is also present. In the presence of Ca2+ these proteins bind to liposomes composed of acidic phospholipids. Calcium-sensitivity of binding differs for each protein and is also influenced by the pH of the buffer used for the liposome-binding assay. Antiserum raised to the 33 to 35-kDa doublet purified on SDS-PAGE recognises the doublet in crude extracts from maize and proteins of similar Mr in Tradescantia virginiana and tobacco Nicotiana tabacum L. The antiserum also recognises p68 (Annexin VI) from chicken gizzard extracts, indicating homology between animal annexins and the maize proteins. For the maize proteins to be involved in the regulation of exocytosis, binding to phospholipids would be expected to occur at physiological levels of Ca2+. The characteristics of the maize annexin-like proteins are described and attention drawn to the marked effect of pH in lowering the requirement for Ca2+ for phospholipid binding.
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Abbreviations
- DEAE:
-
diethylaminoethyl
- EGTA:
-
ethylene glycol-bis (β-aminoethyiether)-N,N,N′,N′-tetraacetic acid
- kDa:
-
kilodalton(s)
- Mr :
-
relative molecular mass
- SDS-PAGE:
-
sodium dodecyl sulphate-polyacrylamide gel electrophoresis
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This work was funded by the Agricultural and Food Research Council. Our thanks also to Professor P. Lowry and Dr R. Woods, Department of Biochemistry, University of Reading for facilities and advice for antiserum production, and C. Boustead, Department of Biochemistry, University of Leeds for advice on immunoblotting and phospholipid-binding assays.
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Blackbourn, H.D., Walker, J.H. & Battey, N.H. Calcium-dependent phospholipid-binding proteins in plants. Planta 184, 67–73 (1991). https://doi.org/10.1007/BF00208238
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DOI: https://doi.org/10.1007/BF00208238