Herbicide point mutation

In goose grass (E. indica), two alleles of a-tubulin 1 (each is the result of a single unique point mutation) have been described, which confer either an intermediate or high level of tolerance to a number of antimicrotubule herbicides, for example, dinitroanilines and phosphoroamidates (Anthony and Hussey, 1999 Yamamoto et al, 1998 Yamamoto and Bird, 1999 Zeng and Baird, 1997). A positive transformation system may be developed using these herbicides. 

This point mutation is the result of a change of one amino-acid residue and the subsequent loss of the protein-herbicide affinity. Resistant chloroplasts are 1,000-fold more resistant than susceptible chloroplasts, when assayed in vitro. Whole plants also show at least a 200-fold increased resistance, which confers a clear, selective advantage in triazine treated fields, orchards and vineyards (13). Evidence that a specific gene is responsible for resistance was proven by the production of tolerant transgenic tobacco (12). Now, nearly 50 species are known to have at least one triazine resistant population and several million hectares in more than 15 countries are infected with triazine resistant weed populations (1). 

Considerable effort has been undertaken in recent years to elucidate the mode of action of herbicides on ACC at the molecular level. Point mutations in the... 

The reaction center of photosystem II (PSII) consists of three proteins The 32-kDa protein (32K, also referred to as D,), Dj, and cytochrome bjs, (1,2). It has several features in common with the reaction center from purple bacteria (3,4), including amino acid sequence homology in functional regions (3,5), arrangement of the transmembrane helices (6,7,8), and conservation of the binding sites for chlorophylls, pheophytins, quinones and a non-heme iron (3,4,6,7,9). Furthermore, 32K and the L-subunit of the bacterial reaction center are the site of triazine herbicide action (10-12), and point mutations at conserved residues in these proteins can confer herbicide resistance (3,13-15). 

Tabk I shows the rq)lacement of amino add residues by nonsynonymous nucleotide substitutions in the S conserved amino add sequences on the ALS enzyme. The alteration of tyrosine (Y) to histidine (H) in the first amino acid sequence (1) of R on A was found in 1 resistant bio pe. But it seems tiiat this replacement does not confer resistance, because the other resistant biotypes did not have such a replacement All S resistant biotypes had various point mutations (A, S, L) in the codon for a proline residue (P) in the second amino acid sequence (2) of Region A, whereas all 7 susceptible biotypes had the proline residue (P). Because this proline codon (P) is common with oth susceptible plants sequenced so far, it is possible tiiat its replacement confers resistance to SU herbicides in L, micrantha. Finalty, all of the resistant plants had an alteration from proline (P) to s me (A), alanine (S), or lysine (L) compared with the amino acid sequence of susceptible plants reported. 

The continuous use of a particular class of hobicide leads to a marked decrease in genetic diversity within populations. At the stone time, it selects plants possessing specific genetic traits conforing herbicide resistance. In addition, substitutions of amino acid residues confer resistance to SU herbicides. In this case, various point mutations occurred in the prolme codon and encoded several possible arttino acid substitutions. When we reviewed the data on genetic relationships and amino acid substitutions of the 69 plants, the emerged picture showed that resistant biotypes wo-e independent selected fiom these areas because of at least five different amino acid substitutions (i.e., 1 KTMr+KTNr 2 YCGr 3 YSSr 4 AKTr 5 ASTr). 

The first amino acid shows the sequences between position 361 and the C-terminal position 644 in the mutated ALS, and second amino acid sequence does that in the wild-type ALS. The mutations involved the residues of tryptophan 548 to leucine and serine 627 to isoleucine. This double mutation on rice is a new combination of spontaneous mutations with the novel substitution at the serine position (7). One-point mutated ALS genes were then prepared to compare the sensitivities of their recombinant ALS s to the ALS-inhibiting herbicides with that of the two-point mutant (Fig. 10). 

On the conttary, the ALS expressed from the two-point mutated ALS gene showed quite different sensitivities to the herbicides. This ALS showed a... 

The one-point mutated ALS except for the mutation of serine at position 627 to isoleucine exhibited a high resistance to the SU herbicide, chlorsulfuron. On the contrary, the resistance level of these mutated ALS s to the IM herbicide, imazaquin, was lower than that of chlorsulfuron. The mutation of proline at position 171 to alanine and histidine did not confer resistance to imazaquin. On the odier hand, the resistance level of die proline mutated ALS s to the PC herbicides was moderate between diose of chlorsulfuron and imazaquin. These results were correlated to the cross-resistance pattern of the proline-mutated ALS of K. scoparia as already described. From these results, it is considered that rice mutated recombinant ALS s, especially the proline mutants, are useful as resistant enzyme models for the herbicide resistance management at newly developed or developing ALS-inhibiting herbicides. We are now preparing other kinds of proline mutants to confirm this idea. 

Fig. 2.1.6. AHAS mutations conferring herbicide resistance. Arrows point to positions in the sequences of AHAS from plant Arabidopsis thaliana), yeast Saccharomyces cerevisiae), and bacterial (Escherichia coli, isozyme II) sources where spontaneous or induced mutations result in an herbicide-insensitive enzyme. Colors designate substitutions occurring in more than one species.

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