Three kinds of discs were taken from tobacco leaves whose lower epidermis had been peeled off, half-peeled or unpeeled. The role of the epidermis and its relation to the kinetin effect on chlorophyll degradation during senescence were studied. Our results follow. Chlorophyll degradation due to kinetin was retarded only when the lower epidermis was present. The decrease in chlorophyll content in leaf discs on water during senescence was nearly proportional to the size of the lower epidermis attached to the discs; i.e., unpeeled discs>half-peeled discs>peeled discs. Cellular fractions possessing activity which induces chlorophyll degradation were extracted from the isolated lower epidermis (i, ii) and its acetone powder (iii): (i) L-2 fraction (1.14≤d≤1.16) was separated by stepwise sucrose density-gradient centrifugation from the 10,000×g pellet of the cell homogenate. (ii) The A-fraction (M.W.≥5,000) was precipitated with 0–80% saturation of ammonium sulfate from 105,000 × g supernatant of cell homogenate and eluted in the void volume by Sephadex G-25 column chromatography. (iii) The fraction precipitated with 0–30% saturation of ammonium sulfate from the 105,000×g supernatant, contained a large amount of DNA and its activity remained even if DNA was removed. Activity was not retained when the fractions were obtained from isolated lower epidermis pretreated with 2×10−5M kinetin for 2 hr in darkness at 25°C.
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Kunihiro Kasamo (1976) studied this question.