Peroxidase isoenzymes as markers of cell de-differentiation in grapevines (Vitis vinifera)

The expression, and the tissue localization and the subcellular localization of peroxidase isoenzymes were studied during the development of mesocarp and hypodermal tissues of Vitis vinifera fruits cvs Airen and Monastrell until fruit softening. In addition the de-differentiation process of mesocarp tissues to form callus cultures was investigated. Both grapevine cultivars contain, as the only component of peroxidase polymorphism in the whole fruit, the peroxidase isoenzyme Bg (the sole component of the peroxidase isoenzyme group HpI BPrx in grapevines), which is both developmentally-regulated and tissue-specific. The establishment of cell cultures from the mesocarp of grapevine fruits is accompanied by the de novo expression of one acidic (APrx) and one basic (LpI BPrx) peroxidase isoenzyme group, whose particular isoenzyme composition is cultivar-specific and dependent on subcellular location (soluble or bound to cell wall). These results suggest that, during the establishment of grape vine cell cultures from mesocarp tissues of grapevine fruits, there is a differential expression of the peroxidase isoenzyme groups APrx and LpI BPrx located in both soluble and cell wall-bound fractions. This de novo gene expression probably expresses at molecular level the totipotency of grapevine somatic cells, which is linked to the de-differentiation process associated with the in vitro culture

The localization of NADPH oxidase and reactive oxygen species in in vitro-cultured Mesembryanthemum crystallinum L. hypocotyls discloses their differing roles in rhizogenesis

This work demonstrated how reactive oxygen species (ROS) are involved in the regulation of rhizogenesis from hypocotyls of Mesembryanthemum crystallinum L. cultured on a medium containing 1-naphthaleneacetic acid (NAA). The increase of NADPH oxidase activity was correlated with an increase of hydrogen peroxide (H2O2) content and induction of mitotic activity in vascular cylinder cells, leading to root formation from cultured hypocotyls. Diphenylene iodonium (DPI), an inhibitor of NADPH oxidase, inhibited H2O2 production and blocked rhizogenesis. Ultrastructural studies revealed differences in H2O2 localization between the vascular cylinder cells and cortex parenchyma cells of cultured explants. We suggest that NADPH oxidase is responsible for H2O2 level regulation in vascular cylinder cells, while peroxidase (POD) participates in H2O2 level regulation in cortex cells. Blue formazan (NBT) precipitates indicating superoxide radical (O2 •−) accumulation were localized within the vascular cylinder cells during the early stages of rhizogenesis and at the tip of root primordia, as well as in the distal and middle parts of newly formed organs. 3,3′-diaminobenzidine (DAB) staining of H2O2 was more intense in vascular bundle cells and in cortex cells. In newly formed roots, H2O2 was localized in vascular tissue. Adding DPI to the medium led to a decrease in the intensity of NBT and DAB staining in cultured explants. Accumulation of O2 •− was then limited to epidermis cells, while H2O2 was accumulated only in vascular tissue. These results indicate that O2 •− is engaged in processes of rhizogenesis induction involving division of competent cells, while H2O2 is engaged in developmental processes mainly involving cell growth