An isoleucine residue within the carboxyl-transferase domain of multidomain acetyl-coenzyme A carboxylase is a major determinant of sensitivity to aryloxyphenoxypropionate but not to cyclohexanedione inhibitors

Abstract

A 3,300-bp DNA fragment encoding the carboxyl-transferase domain of the multidomain, chloroplastic acetyl-coenzyme A carboxylase (ACCase) was sequenced in aryloxyphenoxypropionate (APP)-resistant and -sensitive Alopecurus myosuroides (Huds.). No resistant plant contained an Ile-1,781-Leu substitution, previously shown to confer resistance to APPs and cyclohexanediones (CHDs). Instead, an Ile-2,041-Asn substitution was found in resistant plants. Phylogenetic analysis of the sequences revealed that Asn-2,041 ACCase alleles derived from several distinct origins. Allele-specific polymerase chain reaction associated the presence of Asn-2,041 with seedling resistance to APPs but not to CHDs. ACCase enzyme assays confirmed that Asn-2,041 ACCase activity was moderately resistant to CHDs but highly resistant to APPs. Thus, the Ile-2,041-Asn substitution, which is located outside a domain previously shown to control sensitivity to APPs and CHDs in wheat (Triticum aestivum), is a direct cause of resistance to APPs only. In known multidomain ACCases, the position corresponding to the Ile/Asn-2,041 residue in A. myosuroides is occupied by an Ile or a Val residue. In Lolium rigidum (Gaud.), we found Ile-Asn and Ile-Val substitutions. The Ile-Val change did not confer resistance to the APP clodinafop, whereas the Ile-Asn change did. The position and the particular substitution at this position are of importance for sensitivity to APPs.

Figure 1.

Inhibition of ACCase activity in sensitive (00-017, ▪) and resistant (02-F1, ▴) A. myosuroides population by clethodim and cycloxydim (CHDs) and by fenoxaprop, diclofop, clodinafop, and haloxyfop (APPs). ACCase from sensitive plants have Ile at positions 1,781 and 2,041, whereas ACCase from resistant plants have Ile and Asn at positions 1,781 and 2,041, respectively. Averages of two independent experiments are shown with error bars. ACCase activity is expressed as a percentage of ACCase activity without inhibitor for each population.

Figure 2.

Alignment of amino acid sequences of multidomain ACCases around the site of the Ile residue critical for APP sensitivity (in bold). Higher plant ACCases are underlined. C, Chloroplastic ACCases. L, ACCase isoforms containing a Leu residue at position 1,780 in sequence AJ310767. Dots, Residues identical to those in sequence AJ310767. Dashes, Gaps. The fragment shown extends from Asp 2,002 to Ser 2,079 in sequence AJ310767.

Figure 3.

Phylogenetic tree of the 29 A. myosuroides ACCase haplotypes calculated by the maximum parsimony method. The haplotype names are the same as in EMBL accession number ALIGN_000483. Bootstrap values ≥ 50% are shown. Haplotypes containing an Ile-2,041-Asn mutation are in bold.