Identification of a Novel 2,4-D Metabolic Detoxification Pathway in 2,4-D-Resistant Waterhemp (Amaranthus tuberculatus).

A 2,4-dichlorophenoxyactic acid (2,4-D)-resistant population of Amaranthus tuberculatus (common waterhemp) from Nebraska, USA, was previously found to have rapid metabolic detoxification of the synthetic auxin herbicide 2,4-D. We purified the main 2,4-D metabolites from resistant and susceptible plants, solved their structures by nuclear magnetic resonance (NMR) and high-resolution mass spectrometry (HRMS), and synthesized the metabolites to determine their in planta toxicity. Susceptible plants conjugated 2,4-D to aspartate to form 2,4-D-aspartic acid (2,4-D-Asp), while resistant plants had a unique metabolic profile where 2,4-D was hydroxylated into 5-OH-2,4-D, followed by conjugation into a sugar metabolite (2,4-D-5-O-d-glucopyranoside) and subsequent malonylation into 2,4-D-(6'-O-malonyl)-5-O-d-glucopyranoside. Toxicological studies on waterhemp and Arabidopsis thaliana confirmed that the hydroxylated metabolite lost its auxinic action and toxicity. In contrast, the 2,4-D-Asp metabolite found in susceptible plants retained some auxinic action and toxicity. These results demonstrate that 2,4-D-resistant A. tuberculatus evolved novel detoxification reactions not present in susceptible plants to rapidly metabolize 2,4-D, potentially mediated by cytochrome P450 enzymes that perform the initial 5-hydroxylation reaction. This novel mechanism is more efficient to detoxify 2,4-D and produces metabolites with lower toxicity compared to the aspartic acid conjugation found in susceptible waterhemp.

An in-frame deletion mutation in the degron tail of auxin coreceptor IAA2 confers resistance to the herbicide 2,4-D in Sisymbrium orientale.

The natural auxin indole-3-acetic acid (IAA) is a key regulator of many aspects of plant growth and development. Synthetic auxin herbicides such as 2,4-D mimic the effects of IAA by inducing strong auxinic-signaling responses in plants. To determine the mechanism of 2,4-D resistance in a Sisymbrium orientale (Indian hedge mustard) weed population, we performed a transcriptome analysis of 2,4-D-resistant (R) and -susceptible (S) genotypes that revealed an in-frame 27-nucleotide deletion removing nine amino acids in the degron tail (DT) of the auxin coreceptor Aux/IAA2 (SoIAA2). The deletion allele cosegregated with 2,4-D resistance in recombinant inbred lines. Further, this deletion was also detected in several 2,4-D-resistant field populations of this species. Arabidopsis transgenic lines expressing the SoIAA2 mutant allele were resistant to 2,4-D and dicamba. The IAA2-DT deletion reduced binding to TIR1 in vitro with both natural and synthetic auxins, causing reduced association and increased dissociation rates. This mechanism of synthetic auxin herbicide resistance assigns an in planta function to the DT region of this Aux/IAA coreceptor for its role in synthetic auxin binding kinetics and reveals a potential biotechnological approach to produce synthetic auxin-resistant crops using gene-editing.

Synthetic auxin herbicides: finding the lock and key to weed resistance.

Synthetic auxin herbicides are designed to mimic indole-3-acetic acid (IAA), an integral plant hormone affecting cell growth, development, and tropism. In this review, we explore target site genes in the auxin signaling pathway including SCFTIR1/AFB, Aux/IAA, and ARFs that are confirmed or proposed mechanisms for weed resistance to synthetic auxin herbicides. Resistance to auxin herbicides by metabolism, either by enhanced cytochrome P450 detoxification or by loss of pro-herbicide activation, is a major non-target-site resistance pathway. We speculate about potential fitness costs of resistance due to effects of resistance-conferring mutations, provide insight into the role of polyploidy in synthetic auxin resistance evolution, and address the genetic resources available for weeds. This knowledge will be the key to unlock the long-standing questions as to which components of the auxin signaling pathway are most likely to have a role in resistance evolution. We propose that an ambitious research effort into synthetic auxin herbicide/target site interactions is needed to 1) explain why some synthetic auxin chemical families have activity on certain dicot plant families but not others and 2) fully elucidate target-site cross-resistance patterns among synthetic auxin chemical families to guide best practices for resistance management.