Increase of polyphenol oxidase and decrease of polyarnines correlate with tissue browning in Virginia pine (Pinus virginiana Mill.)

Tissue browning seriously decreases in vitro regeneration from callus cultures in some of woody plants, especially plant regeneration via organogenesis in pine species. Although most of callus cultures turn to brown, a few of them do not become brown during the process of in vitro regeneration in Virginia pine (Pinus virginiana Mill.). We have recently identified a browning callus line and a non-browning callus line in Virginia pine. For understanding the molecular mechanism of tissue browning in pine, lipid peroxidation, polyphenol oxidase, antioxidant enzymes, and polyamines in the browning callus and the non-browning callus were quantitatively analyzed. An increment in lipid peroxidation and polyphenol oxidase and a decrease in antioxidant enzymes ascorbate peroxidase (APOX), glutathione reductase (GR) and superoxide dismutase (SOD) activities were observed in the browning calluses, compared to the non-browning calluses. Antioxidant enzyme activity decreased rapidly soon after the start of the culture period, especially between the third and fourth weeks of culture. Concentrations of the fraction of soluble amines were different. Putrescine (Put) was reduced 63.8-71.5% in the browning tissue, spermidine (Spd) was decreased to 47-65.6%, and spermine (Spm) content was decreased to 62.3-74.5%, at the third and fifth week of culture, respectively. Our results demonstrated that tissue browning is associated with both accumulation of polyphenol oxidase and decrease of putrescine, spermidine, and spermine in browning calluses, which inhibited callus growth, shoot differentiation, and rooting of shoots derived from browning tissues in Virginia pine. However. more research is necessary to elucidate their precise role in tissue browning in different species. (C) 2004 Elsevier Ireland Ltd. All rights reserved.