In Vivo 31P-Nuclear Magnetic Resonance Studies of Glyphosate Uptake, Vacuolar Sequestration, and Tonoplast Pump Activity in Glyphosate-Resistant Horseweed  

Abstract

Horseweed (Conyza canadensis) is considered a significant glyphosate-resistant (GR) weed in agriculture, spreading to 21 states in the United States and now found globally on five continents. This laboratory previously reported rapid vacuolar sequestration of glyphosate as the mechanism of resistance in GR horseweed. The observation of vacuole sequestration is consistent with the existence of a tonoplast-bound transporter. 31 P-Nuclear magnetic resonance experiments performed in vivo with GR horseweed leaf tissue show that glyphosate entry into the plant cell (cytosolic compartment) is (1) first order in extracellular glyphosate concentration, independent of pH and dependent upon ATP; (2) competitively inhibited by alternative substrates (aminomethyl phosphonate [AMPA] and N-methyl glyphosate [NMG]), which themselves enter the plant cell; and (3) blocked by vanadate, a known inhibitor/blocker of ATP-dependent transporters. Vacuole sequestration of glyphosate is (1) first order in cytosolic glyphosate concentration and dependent upon ATP; (2) competitively inhibited by alternative substrates (AMPA and NMG), which themselves enter the plant vacuole; and (3) saturable.31 P-Nuclear magnetic resonance findings with GR horseweed are consistent with the active transport of glyphosate and alternative substrates (AMPA and NMG) across the plasma membrane and tonoplast in a manner characteristic of ATP-binding cassette transporters, similar to those that have been identified in mammalian cells.

Glyphosate resistance in Ambrosia trifida: Part 2. Rapid response physiology and non‐target‐site resistance

“…GR RR and GR NRR phenotypes also show a higher CPi/VPi ratio than the GS phenotype. The lower VPi levels would suggest that GR RR and NRR cells are less capable of buffering a pH challenge . At 16 h onwards, the GR RR phenotype shows a progressive loss in ATP signal and a decline in CPi and VPi, which indicates cellular death as a result of the RR phenotype (Fig.…”

“…Glyphosate mobility in the plant follows sucrose transport movement, and glyphosate mobility is essential for its herbicidal activity . Reduced translocation of glyphosate has been associated with sequestration of glyphosate into the vacuoles in several weed species . The involvement of reduced translocation of glyphosate in GR A. trifida remains unclear, with some reports of reduced translocation in the GR phenotype, while other studies reported no differences in translocation …”

De Novo Genome Assembly of the Economically Important Weed Horseweed Using Integrated Data from Multiple Sequencing Platforms

“…With few genomic resources for weeds and little expertise to utilize the available resources among the weed science community (Stewart, 2009), discovering the molecular mechanisms underlying nontarget resistance has been extremely difficult. However, now that the genomics era has found its way to weed science, we can begin to answer fundamental questions about what makes weeds so weedy and capable of adapting to control measures and, thereby, design approaches that reduce the further evolution of herbicide resistance in weeds (Basu et al, 2004;Stewart et al, 2009) and , EPSPS gene amplification (Gaines et al, 2010(Gaines et al, , 2013Jugulam et al, 2014), and vacuolar sequestration of glyphosate (Ge et al, 2010(Ge et al, , 2012(Ge et al, , 2014; however, in the last case, the genes conferring resistance have not been identified.…”

Procedures for Detection of Resistant Weeds Using 14C- Herbicide Absorption, Translocation, and Metabolism

“…In order to study glyphosate resistance, 31 P nuclear magnetic resonance (NMR) spectroscopy studies were employed to track glyphosate movement and metabolism in resistant and susceptible biotypes of horseweed (Conyza canadensis), and the results showed that the rate of vacuole accumulation of this herbicide is faster and occurs to a greater extent in the resistant biotype rather than in the susceptible [15].…”