Previously, it was reported that plant protoplasts isolated from Betula platyphylla (white birch) callus secreted bundles of hollow callose fibrils in acidic culture medium containing a high concentration of calcium ions (Ca2+). Here, the callose synthase was characterized from in situ and in vitro perspectives. Localization of callose synthases at the secreting site of callose fiber was indicated from in situ immunostaining observation of protoplasts. For in vitro analyses, membrane proteins were extracted from membrane fraction of protoplasts with a 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonate (CHAPS) treatment. The CHAPS extract aggregated in the presence of a high concentration of Ca2+, suggesting that Ca2+ may promote the arrangement of callose synthases in the plasma membrane. The callose synthase activity was dependent on pH and Ca2+, similar to the callose synthase of Arabidopsis thaliana. However, the synthesized fibril products were longer than those produced by callose synthases of herbaceous plants. This is the first insight into the specific properties of callose synthases of woody plants that secrete of callose hollow fibers.
We thank Prof. R. Malcolm Brown Jr. for providing the β-1,3-glucan synthase antibody. We are grateful to the Research Support Center, Research Center for Human Disease Modeling, Kyushu University Graduate School of Medical Sciences, for assistance with analyses using the multimode plate reader, and the Center for Advanced Instrumental and Educational Supports, Faculty of Agriculture, Kyushu University, for assistance with confocal laser scanning microscopy.
Author contributions: All the authors have accepted responsibility for the entire content of this submitted manuscript and approved submission.
Research funding: None declared.
Employment or leadership: None declared.
Honorarium: None declared.
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