Altered acetyl-CoA carboxylase confers herbicide tolerance and may affect fatty acid biosynthesis

--Lorelei C. Marshall, Robin A. Keith, Donald L. Wyse, Burle G. Gengenbach, and John W. Gronwald

Acetyl-CoA carboxylase (ACCase) activity from five mutants exhibiting partially dominant tolerance to ACCase-inhibiting herbicides (Parker et al., PNAS 87:7175, 1990) was 16- to 350-fold less inhibited by these herbicides than ACCase activity in wildtype (susceptible) maize. Seedlings of the homozygous tolerant mutants survived 2- to > 130-fold higher rates of these herbicides than wildtype seedlings in greenhouse tests. The tolerance to representatives of two classes of ACCase-inhibiting herbicides (cyclohexanediones, e.g. sethoxydim, and aryloxyphenoxypropionates, e.g. haloxyfop) cosegregated and increased in parallel for plant and ACCase responses in the mutants. Based on allelism tests, the mutations appeared to be located in a single gene or in closely linked genes. These results suggested that we have identified at least four alleles of the ACCase structural gene--one wildtype susceptible allele (acc) and at least three alleles representing distinguishable mutant phenotypes. All mutants conferred moderate tolerance to haloxyfop, but each conferred different levels of tolerance to sethoxydim. The Acc-H1 allele had no sethoxydim tolerance, Acc-H2 had intermediate tolerance and Acc-S1, Acc-S2, Acc-S3 exhibited tolerance to high rates of sethoxydim. Differences in cross-tolerance to the two herbicides suggested that sites on ACCase that interact with the different herbicide chemistries do not completely overlap.

The single gene inheritance, high levels of herbicide tolerance, and retention of ACCase catalytic activity in the mutants suggested that the tolerance alleles may be of agronomic interest. Supporting this, the herbicide-tolerance alleles were expressed in diverse genetic backgrounds including F1 and backcross combinations with the inbreds A188, A619, A641, A665, A682, B73 and W153R. The tolerance trait has been genetically stable over at least four generations. Herbicide treatments at rates recommended for control of annual grass weeds did not adversely affect grain yield or quality of tolerant lines. Incorporation of the tolerance trait was not associated with deleterious effects on grain yield or quality in the absence of herbicide. These results warrant efforts to transfer the tolerance trait into elite maize lines.

The mutants will be useful in studying maize ACCase. ACCase is the first committed step in fatty acid biosynthesis and may regulate lipid biosynthesis and thus kernel oil deposition. Homozygous Acc-H1 and Acc-H2 mutants had ACCase activity in the absence of herbicide that was about 25% lower than wildtype, suggesting alterations in ACCase that affected some catalytic function. Homozygous Acc-S1, S2, and S3 mutants had wildtype levels of ACCase activity and a slight tendency for increased kernel oil concentration in the absence of herbicide treatments (overall mean of 9% more than wildtype). These results suggested that alteration(s) conferring reduced herbicide inhibition of ACCase activity may alter the regulatory properties of ACCase in controlling fatty acid biosynthesis and thereby affect lipid biosynthesis.

To further investigate the role of ACCase in kernel oil deposition, we are currently comparing the herbicide inhibition of mutant and wildtype ACCase extracted from seedling leaves and developing embryo and endosperm. Our preliminary results indicate that the herbicide-tolerant form of ACCase is expressed in all three tissues. Furthermore, the herbicide-tolerant ACCase is expressed throughout kernel development in both embryo and endosperm. These results lead us to speculate that a single ACCase gene is expressed throughout the plant and that it produces precursor for synthesis of fatty acids for both the housekeeping function of membrane maintenance and for oil deposition in the developing kernel. Based on this one gene model it is possible to predict that an acc-null allele will be lethal provided that other ACCase isozymes do not exist. A recessive mutant exhibiting a substantial reduction in ACCase activity should have low kernel oil and be an albino lethal; bleached leaves are observed when wildtype seedlings are treated with sublethal rates of the ACCase inhibiting herbicides presumably because of a reduction in ACCase activity. This predicted phenotype is similar to the description of loc1 (PlewaMJ, MNL 53:93, 1973). Such a mutant would be of great interest to us and we would appreciate any guidance to a source of loc1 or similar phenotype.

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