Mesotrione is a new herbicide being developed for the selective pre- and post-emergence control of a wide range of broad-leaved and grass weeds in maize (Zea mays). It is a member of the benzoylcyclohexane-1,3-dione family of herbicides, which are chemically derived from a natural phytotoxin obtained from the Californian bottlebrush plant, Callistemon citrinus. The compound acts by competitive inhibition of the enzyme 4-hydroxyphenylpyruvate dioxygenase (HPPD), a component of the biochemical pathway that converts tyrosine to plastoquinone and alpha-tocopherol. Mesotrione is an extremely potent inhibitor of HPPD from Arabidopsis thaliana, with a Ki value of c 6-18 pM. It is rapidly taken up by weed species following foliar application, and is distributed within the plants by both acropetal and basipetal movement. Maize is tolerant to mesotrione as a consequence of selective metabolism by the crop plant. Slower uptake of mesotrione, relative to susceptible weed species, may also contribute to its utility as a selective herbicide for use in maize.

Abstract.Temperatures of more than 25 ~ C adversely affect the activity of soluble starch synthase (SSS), an amyloplastic enzyme, in endosperm of wheat (Triticum aestivum L. cv. Mardler). Enzyme rate was found to have a temperature optimum between 20 and 25~ This effect was apparently reversible after a short period of exposure to elevated temperature. We also found that with a prolonged period of exposure to elevated temperature there was another temperature-related phenomenon which caused a loss of enzyme activity that appeared to be much slower to reverse. We have termed this effect of temperature on SSS activity "knockdown". The knockdown in SSS activity also occurred in-vivo. However, elevated temperature did not affect the activities of several other enzymes in the pathway of starch synthesis (ADP-glucose pyrophosphorylase, UDP-glucose pyrophosphorylase, sucrose synthase, phosphoglucomutase, phosphoglucose isomerase, bound starch synthase or hexokinase). Because the knockdown effect appeared to be specific to the enzyme SSS, we quantified the effect of knockdown on flux of carbon into starch and used these data to calculate the flux-control coefficient for SSS. Using data at 10-20~ the flux-control coefficient was C Starchl(~20C = 0.50, whereas at 20-30 ~ C the flux-control coefficient was C Starch2(~30C= 1.38, and between 30-40~ the flux-control coefficient was C s~"rch3w4~ =0.69. Using data at 10-30~ the flux-control coefficient was cStarch~W3~ = 1.15, and at 10M0~ the flux-control coefficient was cStarchl(~40C=0.82. In conclusion, we suggest that SSS is a major site of regulation of starch synthesis 515) 6 85 25 48 in developing wheat grain. During periods of high temperature the control point in the pathway of starch synthesis is apparently not associated exclusively with ADPglucose pyrophosphorylase. In field conditions, in which temperatures are fluctuating, there will likely be periods of control of starch synthesis being exerted predominantly by SSS. During periods at lower temperature, control of flux may be exerted by SSS, perhaps in combination with other flux-controlling enzymes in the pathway. Our data point-out a crucial new aspect of quantifying control strengths of enzymes in plants: the determination of enzyme control strengths should be done in carefully regulated temperature conditions. Thus, since temperature is a major determinant of real flux through a pathway and the individual enzymes can respond differently to changing temperature conditions, the control strengths of individual steps in a pathway may vary with changing environmental conditions. This is particularly pronounced in starch deposition, because of the temperature instability of SSS.

Several GSTs have been characterised in maize. GST I is a homodimer of 29 kDa subunits, GST II a hetrodimer of 27 kDa and 29 kDa subunits and GST IV a homodimer of 27 kDa subunits. We report the isolation and characterization of a herbicide-safener inducible cDNA clone, GST-27. Based on partial amino acid sequence, GST-27 encodes the 27 kDa subunit present in both glutathione S-transferase isoforms GST II and IV. Northern blotting was used to compare the expression patterns of GST-27 with that of GST-29. Transcripts corresponding to GST-27 were found to be constitutively expressed in RNA isolated from the root, but no expression was detected in RNA isolated from aerial parts of the plant. The application of herbicide safener caused a dramatic increase in the expression of GST-27 in all aerial plant parts tested. GST-29 was found to be constitutively expressed in RNA isolated from a number of maize tissues. The basal level of GST-29 expression showed a minimal increase upon herbicide safener treatment. Although a range of hormonal, environmental and physiological stimuli failed to elevate GST-27 levels, some increase in GST-27 mRNA was observed in the late stages of leaf senescence and after treatments resulting in phytotoxic effects.

The safener-induced maize (Zea mays L.) glutathione S-transferase, GST II (EC 2.5.1.18) and another predominant isoform, GST I, were purified from extracts of maize roots treated with the safeners R-25788 (N,N-diallyl-2-dichloroacetamide) or R-29148 (3-dichloroacetyl-2,2,5-trimethyl-1,3-oxazolidone). The isoforms GST I and GST II are respectively a homodimer of 29-kDa (GST-29) subunits and a heterodimer of 29- and 27-kDa (GST-27) subunits, while GST I is twice as active with 1-chloro-2,4-dinitrobenzene as GST II, GST II is about seven times more active against the herbicide, alachlor. Western blotting using antisera raised against GST-29 and GST-27 showed that GST-29 is present throughout the maize plant prior to safener treatment. In contrast, GST-27 is only present in roots of untreated plants but is induced in all the major aerial organs of maize after root-drenching with safener. The amino-acid sequences of proteolytic fragments of GST-27 show that it is related to GST-29 and identical to the 27-kDa subunit of GST IV.

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