SOLUBLE AND BOUND APOPLASTIC PROTEINS AND ISOZYMES OF PEROXIDASE, ESTERASE AND MALATE-DEHYDROGENASE IN OAT PRIMARY LEAVES

The lower epidermis was peeled from 6 day old oat (Avena sativa L.) primary leaves and leaf segments vacuum-infiltrated with pH 6.5 buffered 200 mM NaCl to elute an intercellular washing solution (IWS) containing soluble and weakly ionically bound apoplastic proteins and enzymes (Plant Physiol. 90, 185-190). Segments were then homogenized in buffer, centrifuged, and the supernatant used as a source of soluble cytosolic constituents. The pellet was washed and resuspended in 1M NaCl to solubilize proteins and enzymes strongly ionically bound to cell walls. Four IWS proteins (44, 45, 56 and 71 kD) and two strongly ionically bound proteins (34 and 35kD) were restricted to the apoplast. Wounding by peeling away the lower epidermis 24 h before sampling induced two IWS proteins (32 and 38 kD) whose induction was inhibited by cycloheximide and partially inhibited by tunicamycin. Wounding induced specific peroxidase isozymes in the apoplastic fractions. Apoplastic and symplastic malate dehydrogenase isozyme patterns were similar and not influenced by wounding. Anodic esterase isozymes were found in both apoplastic and cytosolic fractions. Cathodic esterases were restricted to the apoplast. The importance of including soluble and weakly ionically bound apoplastic fractions in studies of cell wall proteins and enzymes is emphasized. Problems encountered in working with solubilized apoplastic fractions are considered. © 1990, Gustav Fischer Verlag, Stuttgart. All rights reserved.