Differential energization of the tonoplast in suspension cells and seedlings from Picea abies |
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Authors: | Wolfgang Pfeiffer |
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Institution: | Institut für Pflanzenphysiologie, Universit?t Salzburg, Hellbrunner Strasse 34, A-5020 Salzburg, Austria e-mail: Wolfgang.Pfeiffer@sbg.ac.at; Fax: 00430662 8044 619, AT
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Abstract: | Vacuolar ATPase (EC 3.6.1.3) and PPase (EC 3.6.1.1) were studied in suspension cells and seedlings from spruce Picea abies (L.) Karst. Proton transport activity and uncoupler (1 μM nigericin) stimulated substrate hydrolysis were measured in tonoplast
enriched membrane vesicles. In suspension cells the vacuolar PPase exhibited 1.8-fold activity of the ATPase. In roots and
needles from 12-week-old spruce seedlings the vacuolar PPase was inactive, whereas the ATPase was active. Therefore, we investigated
whether the preparation of spruce tonoplast vesicles from roots and needles inactivates the vacuolar PPase but not the ATPase.
For this purpose, maize (Zea mays L.) tonoplast membranes exhibiting vacuolar PPase as well as ATPase activity were used as a probe and added to the homogenization
medium prior to the preparation of spruce vesicles. The preparation of spruce vesicles was more inhibitory to the vacuolar
ATPase than to the PPase. The comparison of vacuolar PPases from spruce suspension cells and maize roots revealed similar
enzymatic properties. After isopycnic centrifugation on continuous sucrose gradients the vacuolar PPase from spruce suspension
cells co-purified with the vacuolar ATPase. Together, these data show: (1) vacuolar PPases from spruce suspension cells and
maize roots are similar, (2) the preparation of tonoplast vesicles from spruce roots and needles does not inactivate the vacuolar
PPase, (3) tonoplasts of suspension cultured cells and seedlings from spruce are differentially energized by the vacuolar
pyrophosphatase that may indicate a difference in pyrophosphate metabolism between embryogenic and differentiated spruce cells,
and (4) tonoplast vesicles from spruce seedlings may allow investigations of the effect of pyrophosphate on the vacuolar ATPase
in the absence of vacuolar PPase activity.
Received: 2 July 1998 / Accepted: 14 September 1998 |
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Keywords: | Cell suspension culture Picea abies Vacuolar ATPase Vacuolar PPase |
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