Leaf senescence represents a genetically programmed sequence of events highly coordinated at the cell and tissue levels that happens at the final stage of development. The main symptom of leaf senescence is the yellowing of the blade, due to the loss in the chloroplasts of chlorophylls and membrane structure. It is a common observation that greening and senescence occur at opposite ends of the life span of plant tissues. Greening generally implies differentiation of etioplasts or proplastids into chloroplasts in the presence of light, characterized by the formation of thylakoids. In contrast, during chloroplast senescence, the structure changes markedly: the chloroplast polyribosome population decreases, the organized granal stacks are lost, the plastoglobuli appear larger and more abundant, and finally the plastid envelope ruptures. The present work gives evidence for re-differentiation of senescent chloroplasts (gerontoplasts) in cytokinin-induced regreening of senescent Nicotiana leaves. During regreening the chlorophyll content, chloroplast structure and photosynthetic activity recovered. Ultrathin sections of leaf tissue were examined at several times during senescence and regreening. It was apparent that granal stacks were rebuilding in the senescent chloroplasts, which were still recognizable by the presence of some plastoglobuli. No ultrastructural evidence of proplastids or chloroplast division was observed at any of the developmental stages examined. Trends in plastid numbers per cell also supported the gerontoplast-re-differentiation route. Plastid numbers did not increase during regreening, and when many plastids were lost during advanced senescence, regreening capacity deteriorated. Re-differentiation of gerontoplasts was associated with increased levels of light-harvesting chlorophyll a/b-binding protein (LHCP-2), cytochrome f , small and large subunits of Rubisco, and the appearance of NADP-protochlorophyllide oxidoreductase (POR) detected by Western blotting. Differentiation, de-differentiation, and re-differentiation concepts are discussed.

Key words: chloroplasts, cytokinins., Nicotiana, re-differentiation, senescence