ALOMYbase, a resource to investigate non-target-site-based resistance to herbicides inhibiting acetolactate-synthase (ALS) in the major grass weed Alopecurus myosuroides (black-grass)

Abstract

BackgroundHerbicide resistance in agrestal weeds is a global problem threatening food security. Non-target-site resistance (NTSR) endowed by mechanisms neutralising the herbicide or compensating for its action is considered the most agronomically noxious type of resistance. Contrary to target-site resistance, NTSR mechanisms are far from being fully elucidated. A part of weed response to herbicide stress, NTSR is considered to be largely driven by gene regulation. Our purpose was to establish a transcriptome resource allowing investigation of the transcriptomic bases of NTSR in the major grass weed Alopecurus myosuroides L. (Poaceae) for which almost no genomic or transcriptomic data was available.ResultsRNA-Seq was performed from plants in one F2 population that were sensitive or expressing NTSR to herbicides inhibiting acetolactate-synthase. Cloned plants were sampled over seven time-points ranging from before until 73 h after herbicide application. Assembly of over 159M high-quality Illumina reads generated a transcriptomic resource (ALOMYbase) containing 65,558 potentially active contigs (N50 = 1240 nucleotides) predicted to encode 32,138 peptides with 74 % GO annotation, of which 2017 were assigned to protein families presumably involved in NTSR. Comparison with the fully sequenced grass genomes indicated good coverage and correct representation of A. myosuroides transcriptome in ALOMYbase. The part of the herbicide transcriptomic response common to the resistant and the sensitive plants was consistent with the expected effects of acetolactate-synthase inhibition, with striking similarities observed with published Arabidopsis thaliana data. A. myosuroides plants with NTSR were first affected by herbicide action like sensitive plants, but ultimately overcame it. Analysis of differences in transcriptomic herbicide response between resistant and sensitive plants did not allow identification of processes directly explaining NTSR. Five contigs associated to NTSR in the F2 population studied were tentatively identified. They were predicted to encode three cytochromes P450 (CYP71A, CYP71B and CYP81D), one peroxidase and one disease resistance protein.ConclusionsOur data confirmed that gene regulation is at the root of herbicide response and of NTSR. ALOMYbase proved to be a relevant resource to support NTSR transcriptomic studies, and constitutes a valuable tool for future research aiming at elucidating gene regulations involved in NTSR in A. myosuroides.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1804-x) contains supplementary material, which is available to authorized users.

Optimizing RNA‐seq studies to investigate herbicide resistance

“…A general review of the literature (Tables and S1) reveals a wide array of RNA‐seq experimental set‐ups employed by weed researchers over the last few years . These initial set‐up decisions can drastically influence the number of transcripts that appear as significantly differentially expressed by the end of the analysis.…”

“…All three studies that attempted to control for the genetic background produced shorter DET lists, even when the number of replicates was low. In fact, one of these studies compared only one pooled HR sample with one pooled HS sample and produced 1310 DETs . Other studies with a single replicate returned much longer DET lists, with an average of 44 000 DETs returned in those papers .…”

“…Other studies with a single replicate returned much longer DET lists, with an average of 44 000 DETs returned in those papers . Controlling for the genetic background was done through either recurrent selection of a single population or by studying a segregating F 2 population …”

“…This problem is evidenced in many of the RNA‐seq papers from Table . In one study, there is overall upregulation of many more transcripts in the HS‐treated condition than the HR‐treated condition (7.7× more HS than HR transcripts overexpressed on average across the time points). As may be expected, the identities of many of these HS‐specific transcripts are related to plant stress and death pathways, including protein ubiquitination, cell‐cycle arrest, RNA processing/metabolism, response to wounding, and response to oxidative stress.…”

High conservation of the transcriptional response to acetolactate‐synthase‐inhibiting herbicides across plant species

“…Recently, the transcriptional response to ALS inhibitors of A. myosuroides (black‐grass) was found consistent with the physiological effects of these herbicides observed in broad‐leaved species (Gardin et al ., ). This suggested that pathways involved in ALS‐inhibitor response may be conserved in plants.…”