Integrated metabolomics and transcriptomics reveal that Lolium rigidum ABCC3.2 contributes to multiple herbicide resistance.
The widespread application of glyphosate has resulted in the evolution of glyphosate resistance in Lolium rigidum. This study investigated the resistance mechanism in an Australian population of L. rigidum, designated WALR60. Transcriptome sequencing coupled with quantitative real-time PCR (qRT-PCR) analyses identified significant upregulation of ABCC3.2, ABCB11, and ABCC13 in the WALR60 population compared to susceptible controls. Structural analysis revealed that LrABCC3.2 lacks the N-terminal TMD0 domain present in its homolog LrABCC3.1, representing a core ABC transporter. LrABCC3.2-transformed yeast cells exhibited enhanced tolerance to glyphosate, as compared to the empty vector control. Similarly, rice calli and seedlings overexpressing ABCC3.2 (ABCC3.2-OE) showed increased glyphosate resistance relative to the corresponding GFP-overexpressing (GFP-OE) controls. Additionally, the LrABCC3.2-OE lines displayed resistance to haloxyfop and pinoxaden. Integrative transcriptomic and metabolomic analyses of transgenic rice revealed that LrABCC3.2 overexpression significantly enriches the glycerophospholipid metabolism pathway, with positive correlations between the expression of GPAT3, PLDα2, PLA14 and the accumulation of phosphatidylcholine (PC) and choline. Comparative transcriptome analysis between resistant L. rigidum and LrABCC3.2-OE rice identified 10 commonly upregulated and 18 commonly downregulated genes, indicating a conserved detoxification mechanism. In conclusion, this study demonstrates that ABCC3.2 overexpression contributes to glyphosate resistance in the WALR60 population and elucidates a potential downstream metabolic pathway involved in this trait.