Primary Study Of Alternaria Alternata Crofton-Weed Toxin On Chlamydomonas Reinhardtii

Alternaria alternate Crofton-weed toxin (AAC-toxin) is a phytotoxin produced by A. alternata (Fr.)Keissler which made Crofton-weed pathogenetic. The action site and mode of AAC-toxin wereinvestigated in wild and mutants of Chlamydomonas reinhardtii. The results revealed thatAAC-toxin inhibited the synthesis of ATP, which could be due to uncoupling ATPsynthesis from the electron transport; blocking the electron transport and blocking thephosphorylation reaction itself. So, we analyzed the cause of inhibition of ATP synthesis byClark-type electrode. The study showed that both uncoupled and basal electron flow wereinhibited as concentrations of toxin increased (P＜0.01). However, phosphorylating electrontransport was not affected until the concentrations of toxin reach 40μg/mL (P＞0.05). Inaddition, the inhibitory effect on uncoupled electron transport was greater than the basaland phosphorylating electron transports. So, the inhibition of ATP synthesis was mainlyinduced by the block of electron transport. Subsequently, we measured the effect ofAAC-toxin on electron transport. The results showed that the toxin inhibited the uncoupledPSⅡelectron transport, but uncoupled PSⅠelectron transport was not affected nearly.Therefore, the primary action site of AAC-toxin was located in PSⅡ. The half time ofchlorophyll fluorescence decay and half time of chlorophyll fluorescence rise approvedfurther that toxin inhibited the acceptor side of PSⅡ, not the donor side.Fast Chl a fluorescence transient showed that the J step and F_o increased, and no K peakwas observed after the toxin treatment. The increase of the J level accounted for the lowerrate of electron flow between Q_A and Q_B, It can be conferred that toxin affected the electrontransport between Q_A and Q_B. F_o was caused by transfering the electron from Q_B to Q_A afterthe displacement of Q_B. K peak was attributed to inactivation of the OEC, no K peak in theinduction curves indicated that toxin didn't inhibited the donor side of PSⅡ. Further, Theresults of JIP test demonstrated that there were multiple effects involved in the inhibition oftoxin: (1) blocking electron transport between Q_A and Q_B at the acceptor side of PSⅡ; (2)inactivating PSⅡreaction centers (RCs); (3) decreasing the yield for primary photochemistry; (4) damaging the antennae of PSⅡcentre in part; (5) decreasing energeticconnectivity between the antennae of active and inactive centers.At last, the sensitivity profile of AAC-toxin on herbicide-resistant mutants confirmedthat toxin surely inhibited the electron transport between Q_A, and Q_B by displacing the Q_B,and was a natural PSⅡinhibitor. PSⅡinhibitors can be grouped two families, i.e. theclassical triazine-urea herbicides and phenol-type group; The principle difference betweenthem rests on the fact that the ser264ala mutant is tolerant to classical herbicides in contrastto the latter. Because ser264ala mutant was not tolerant, the mode of action of AAC-toxinwas similar to phenol type herbicides. And the binding of toxin involved in the amino acids275, 219, 256 of D1 protein, the amino acids 275 is more possible site.The aim for this study is to elucidate, and modify the active site of the chemical structureof AAC-toxin so as to develop the new bioherbicide and pave for its futureindustrialization.