To determine if chlorosis caused by tentoxin, a toxin produced by Alternaria tenuis Nees., is due to interference with chlorophyll synthesis directly or to disruption of normal chloroplast development, the effects of the toxin on these processes in cucumber (Cucumis sativus L.) and cabbage (Brassica oleracea L., var. capitata) were studied. Cucumber cotyledons are highly sensitive to the toxin but exhibited no interference with the conversion of protochlorophyll(ide) to chlorophyll(ide) or with the general time course pattern of chlorophyll synthesis, although there was a 90% reduction in chlorophyll concentration. In cabbage, which shows no chlorosis in the presence of the toxin, there was a slight stimulation of chlorophyll synthesis in the presence of the toxin. Electron microscopy revealed that in cucumber, toxin treatment interferes with development of prolamellar bodies and lamellae, and results in deformed plastids. No such effects were noted in toxin-treated cabbage tissues. Plastids in toxin-treated cotyledons of both cucumber and cabbage contained more starch than plastids in nontreated tissues. It was concluded that tentoxin acts through disruption of normal plastid development, rather than through direct interference with chlorophyll synthesis.A chlorosis-inducing toxin (trivial name: tentoxin) produced by the fungus Alternaria tenuis Nees. has been characterized (8) as cycloleucyl-N-methyl alanylglycyl-N-methyl dehydrophenylalanyl. The toxin causes a sharply delimited, variegated, yellow chlorosis in germinating seedlings of many dicotyledonous species, but it has no apparent effect on tomato and members of the Cruciferae and Gramineae (1, 7). Templeton et al. (7) observed that cucumber seedlings were more sensitive to tentoxin in darkness than under continuous light, and that those exposed 48 to 50 hr after the initiation of germination were insensitive to the toxin. They suggested that activity of the toxin is associated with some step late in the development of chloroplasts. Saad et al. (4) (b) interfere with plastid development, thereby affecting chlorophyll synthesis indirectly. The present investigation was undertaken to determine if either of these hypotheses is correct and to determine if cellular organelles other than plastids are affected by the toxin. A preliminary report of this study has been published (2).
MATERIALS AND METHODSProcedures for the production, purification, and quantitation of tentoxin have been described (4). Seeds of cucumber (Cucumis sativus L., c.v. SMR-18) and cabbage (Brassica oleracea L., var. capitata, c.v. Jersey Queen) were germinated in distilled water or tentoxin (30 ,ug/ml), in darkness, at 24 C, for 96 hr, unless otherwise noted. Seedlings were either harvested under a green safelight or exposed to 200 ft-c from General Electric Cool-white fluorescent lamps at 24 C.Protochlorophyll and chlorophyll were extracted by homogenizing 1 g (fresh wt) of cotyledons in 50 volumes (w/v) of acetone in a Sorvall Omni-mixer. The acetone solution was centrifuged a...
