Fitness Cost Associated With Enhanced EPSPS Gene Copy Number and Glyphosate Resistance in an Amaranthus tuberculatus Population

The evolution of resistance to pesticides in agricultural systems provides an opportunity to study the fitness costs and benefits of novel adaptive traits. Here, we studied a population of Amaranthus tuberculatus (common waterhemp), which has evolved resistance to glyphosate. The growth and fitness of seed families with contrasting levels of glyphosate resistance was assessed in the absence of glyphosate to determine their ability to compete for resources under intra- and interspecific competition. We identified a positive correlation between the level of glyphosate resistance and gene copy number for the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) glyphosate target, thus identifying gene amplification as the mechanism of resistance within the population. Resistant A. tuberculatus plants were found to have a lower competitive response when compared to the susceptible phenotypes with 2.76 glyphosate resistant plants being required to have an equal competitive effect as a single susceptible plant. A growth trade-off was associated with the gene amplification mechanism under intra-phenotypic competition where 20 extra gene copies were associated with a 26.5 % reduction in dry biomass. Interestingly, this growth trade-off was mitigated when assessed under interspecific competition from maize.

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No Impact of Increased EPSPS Gene Copy Number on Growth and Fecundity of Glyphosate-Resistant Kochia (Bassia scoparia)

Weed Science ◽

10.1017/wsc.2018.82 ◽

2018 ◽

Vol 67 (1) ◽

pp. 22-28 ◽

Cited By ~ 1

Author(s):

O. Adewale Osipitan ◽

J. Anita Dille

Keyword(s):

Weed Management ◽

Copy Number ◽

Plant Biomass ◽

Management Strategies ◽

Gene Copy Number ◽

Glyphosate Resistance ◽

Gene Copy ◽

Competitive Response ◽

Epsps Gene

AbstractThe level of glyphosate resistance in kochia [Bassia scoparia(L.) A. J. Scott] was reported to be due to an increase in 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene copy number. A field study was conducted near Manhattan, KS, in 2014 and 2015 to evaluate the relationship between EPSPS gene copy number and growth and fecundity variables ofB. scopariaindividuals within suspected glyphosate-resistant (GR) populations from western Kansas. Initial assays of EPSPS gene copy and in vivo shikimate accumulation showed thatB. scopariapopulations from Finney (FN-R), Scott (SC-R), and Thomas (TH-R) counties were segregating for glyphosate resistance, with some individuals still being glyphosate susceptible (GS). A target-neighborhood competition approach was used to evaluate the competitive response of individual target plants with relatively low (classified as GS) and high (classified as GR) EPSPS gene copy number within the populations. There was no relationship observed between EPSPS gene copy number and vegetative or fecundity variables. There was no differential competitive response of target plant biomass to increasing neighbor density between individuals with low and high EPSPS gene copy number within each population. Lack of associated vegetative growth and fecundity cost to the increased EPSPS gene copy in the GRB. scopariaplants suggests that the plants are likely to persist in field populations, except when effective weed management strategies are adopted that would prevent their growth and seed production.

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A Multistate Study of the Association Between Glyphosate Resistance and EPSPS Gene Amplification in Waterhemp (Amaranthus tuberculatus)

Weed Science ◽

10.1614/ws-d-14-00149.1 ◽

2015 ◽

Vol 63 (3) ◽

pp. 569-577 ◽

Cited By ~ 31

Author(s):

Laura A. Chatham ◽

Kevin W. Bradley ◽

Greg R. Kruger ◽

James R. Martin ◽

Micheal D. K. Owen ◽

...

Keyword(s):

Gene Amplification ◽

Copy Number ◽

Quantitative Relationship ◽

Glyphosate Resistance ◽

Epsps Gene ◽

Number Magnitude ◽

Site Of Action ◽

Amaranthus Tuberculatus ◽

Polymerase Chain ◽

The Relationship

Waterhemp is an increasingly problematic weed in the U.S. Midwest, having now evolved resistances to herbicides from six different site-of-action groups. Glyphosate-resistant waterhemp in the Midwest is especially concerning given the economic importance of glyphosate in corn and soybean production. Amplification of the target-site gene, 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) was found to be the mechanism of glyphosate resistance in Palmer amaranth, a species closely related to waterhemp. Here, the relationship between glyphosate resistance and EPSPS gene amplification in waterhemp was investigated. Glyphosate dose response studies were performed at field sites with glyphosate-resistant waterhemp in Illinois, Kansas, Kentucky, Missouri, and Nebraska, and relative EPSPS copy number of survivors was determined via quantitative real-time polymerase chain reaction (qPCR). Waterhemp control increased with increasing glyphosate rate at all locations, but no population was completely controlled even at the highest rate (3,360 g ae ha−1). EPSPS gene amplification was present in plants from four of five locations (Illinois, Kansas, Missouri, and Nebraska) and the proportion of plants with elevated copy number was generally higher in survivors from glyphosate-treated plots than in plants from the untreated control plots. Copy number magnitude varied by site, but an overall trend of increasing copy number with increasing rate was observed in populations with gene amplification, suggesting that waterhemp plants with more EPSPS copies are more resistant. Survivors from the Kentucky population did not have elevated EPSPS copy number. Instead, resistance in this population was attributed to the EPSPS Pro106Ser mutation. Results herein show a quantitative relationship between glyphosate resistance and EPSPS gene amplification in some waterhemp populations, while highlighting that other mechanisms also confer glyphosate resistance in waterhemp.

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Prediction of glyphosate resistance level based on EPSPS gene copy number in Kochia scoparia

10.1101/047878 ◽

2016 ◽

Author(s):

Todd A. Gaines ◽

Abigail L. Barker ◽

Eric L. Patterson ◽

Philip Westra ◽

Eric P. Westra ◽

...

Keyword(s):

Gene Duplication ◽

Empirical Model ◽

Copy Number ◽

Gene Copy Number ◽

Glyphosate Resistance ◽

Gene Copy ◽

Kochia Scoparia ◽

Resistance Level ◽

Epsps Gene ◽

Fallow Systems

AbstractGlyphosate-resistant (GR) Kochia scoparia has evolved in dryland chemical fallow systems throughout North America and the mechanism involves 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene duplication. Sugarbeet fields in four states were surveyed for K. scoparia in 2013 and tested for glyphosate-resistance level and EPSPS gene copy number. Glyphosate resistance was confirmed in K. scoparia populations collected from sugarbeet fields in Colorado, Wyoming, and Nebraska. The GR samples all had increased EPSPS gene copy number, with median population values up to 11. An empirical model was developed to estimate the level of glyphosate-resistance in K. scoparia based on EPSPS gene copy number. The results suggested that glyphosate susceptibility can be accurately diagnosed using EPSPS gene copy number, and further increases in EPSPS gene copy number could increase resistance levels up to 8-fold relative to susceptible K. scoparia. These trends suggest that continued glyphosate selection pressure is selecting for higher EPSPS copy number and higher resistance levels in K. scoparia. By including multiple K. scoparia samples lacking EPSPS gene duplication, our empirical model provides a more realistic estimate of fold-resistance due to EPSPS gene copy number compared to methods that do not account for normal variation of herbicide response in susceptible biotypes.

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EPSPS Gene Copy Number and Whole-Plant Glyphosate Resistance Level in Kochia scoparia

PLoS ONE ◽

10.1371/journal.pone.0168295 ◽

2016 ◽

Vol 11 (12) ◽

pp. e0168295 ◽

Cited By ~ 18

Author(s):

Todd A. Gaines ◽

Abigail L. Barker ◽

Eric L. Patterson ◽

Philip Westra ◽

Eric P. Westra ◽

...

Keyword(s):

Copy Number ◽

Gene Copy Number ◽

Glyphosate Resistance ◽

Gene Copy ◽

Kochia Scoparia ◽

Resistance Level ◽

Epsps Gene ◽

Whole Plant

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Effect of EPSPS Gene Copy Number and Glyphosate Selection on Fitness of Glyphosate-Resistant Bassia Scoparia in The Field

10.21203/rs.3.rs-155310/v1 ◽

2021 ◽

Author(s):

Charlemagne Ajoc Lim ◽

Prashant Jha ◽

Vipan Kumar ◽

Alan T. Dyer

Keyword(s):

Sugar Beet ◽

Seed Production ◽

Great Plains ◽

Copy Number ◽

Gene Copy Number ◽

Reproductive Traits ◽

Gene Copy ◽

Epsps Gene ◽

Gene Copies

Abstract The widespread evolution of glyphosate-resistant (GR) Bassia scoparia in the U.S. Great Plains poses a serious threat to the long-term sustainability of GR sugar beet. Glyphosate resistance in B. scoparia is due to an increase in the EPSPS (5-enolpyruvyl-shikimate-3-phosphate) gene copy number. The variation in EPSPS gene copies among individuals from within a single GR B. scoparia population indicated a differential response to glyphosate selection. We tested the hypothesis of reduced GR B. scoparia fitness (reproductive traits) to increasing glyphosate rates (applied as single or sequential applications) potentially experienced within a GR sugar beet field. The variation in EPSPS gene copy number and total glyphosate rate (single or sequential applications) did not influence any of the reproductive traits of GR B. scoparia, except seed production. Sequential applications of glyphosate with a total rate of 2,214 g ae ha− 1 or higher prevented seed production in B. scoparia plants with 2–4 (low levels of resistance) and 5–6 (moderate levels of resistance) EPSPS gene copies. Timely sequential applications of glyphosate (full recommended rates) can potentially slow down the evolution of GR B. scoparia with low to moderate levels of resistance (2–6 EPSPS gene copies), but any survivors (highly-resistant individuals with ≥ 8 EPSPS gene copies) need to be mechanically removed before flowering from GR sugar beet fields. This research warrants the need to adopt ecologically based, multi-tactic strategies to reduce exposure of B. scoparia to glyphosate in GR sugar beet.

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MET Gene High Copy Number (Amplification/Polysomy) Identified in Melanoma for Potential Targeted Therapy

American Journal of Clinical Pathology ◽

10.1093/ajcp/aqab171 ◽

2021 ◽

Author(s):

Nisha S Ramani ◽

Ajaykumar C Morani ◽

Shengle Zhang

Keyword(s):

Targeted Therapy ◽

Gene Amplification ◽

Copy Number ◽

Kinase Inhibitors ◽

Gene Copy Number ◽

Tissue Microarrays ◽

High Copy Number ◽

Gene Copy ◽

Aberrant Expression ◽

Mesenchymal Epithelial Transition Factor

Abstract Objectives Aberrant expression of the mesenchymal epithelial transition factor (MET) gene has been observed in several malignancies, and drugs targeting the MET gene have been implicated in clinical trials with promising results. Hence, MET is a potentially targetable oncogenic driver. We explored the frequency of MET gene high copy number in melanomas and carcinomas. Methods The study group included 135 patients. Tissue microarrays were constructed with 19 melanomas and 116 carcinomas diagnosed from 2010 to 2012. We screened MET gene copy number by fluorescence in situ hybridization analysis using probes for MET gene and CEP7 as control. Results We found MET gene amplification in 2 (11%) of 19 melanoma cases, whereas 5 (26%) of 19 melanoma cases showed polysomy. For carcinomas, there was no MET gene amplification identified. However, 8 (7%) of 116 cases showed polysomy. Conclusions In our study, MET gene amplification was identified in 11% of melanomas and is relatively concordant with few reported studies. However, about 26% of the additional melanoma cases showed MET gene polysomy, which has not been reported as per our knowledge. If these results are validated with further orthogonal studies, more of the melanoma cases could potentially benefit from targeted therapy with MET tyrosine kinase inhibitors.

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Resistance to methotrexate due to gene amplification in a patient with acute leukemia.

Journal of Clinical Oncology ◽

10.1200/jco.1984.2.1.16 ◽

1984 ◽

Vol 2 (1) ◽

pp. 16-20 ◽

Cited By ~ 92

Author(s):

M D Carman ◽

J H Schornagel ◽

R S Rivest ◽

S Srimatkandada ◽

C S Portlock ◽

...

Keyword(s):

Gene Amplification ◽

Copy Number ◽

Cdna Probe ◽

Leukemia Cell ◽

Gene Copy Number ◽

Leukemia Cell Line ◽

Gene Copy ◽

Acute Myelocytic Leukemia ◽

Human Leukemia ◽

Dot Blot

A patient is described with acute myelocytic leukemia refractory to conventional therapy, who also became highly resistant to methotrexate (MTX) after repeated courses of this drug. Leukemia cells from this patient were found to contain an elevated level of dihydrofolate reductase (DHFR) activity, with no change in the affinity of the enzyme for MTX. A sensitive "dot blot" assay revealed a fourfold increase in the gene copy number of DHFR. Southern blot analysis with a human DHFR cDNA probe confirmed this increase in the gene copy number, and demonstrated a similar restriction pattern with Eco R1, Hind III, and Pst 1 as seen with a highly amplified human leukemia cell line, K562. Additional DHFR fragments were detected, not seen in the K562 blot, suggesting the presence of pseudogenes, or a result of gene rearrangements occurring as part of the amplification process. Resistance to MTX in this patient was therefore ascribed to gene amplification and overproduction of DHFR.

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Gene copy number of epidermal growth factor receptor (EGFR) by fluorescent in situ hybridization (FISH) in head and neck squamous cell carcinomas (HNSCC) is closely correlated with EGFR protein levels assessed by automated quantitative protein analysis (AQUA)

Journal of Clinical Oncology ◽

10.1200/jco.2007.25.18_suppl.6023 ◽

2007 ◽

Vol 25 (18_suppl) ◽

pp. 6023-6023

Author(s):

P. Weinberger ◽

A. Psyrri ◽

P. Kountourakis ◽

T. Rampias ◽

C. Sasaki ◽

...

Keyword(s):

In Situ Hybridization ◽

Protein Expression ◽

Protein Content ◽

Gene Amplification ◽

Copy Number ◽

Gene Copy Number ◽

Gene Copy ◽

Egfr Gene ◽

Protein Levels

6023 Background: EGFR overexpression correlates with recurrence and with treatment resistance in HNSCC. The mechanisms of EGFR protein overexpression are poorly understood. Nonetheless, previous investigators have not demonstrated a correlation between EGFR gene copy number and protein content, using conventional immunohistochemistry (IHC). The aim of this study was to evaluate the relationship of EGFR gene copy number and protein expression utilizing fluorescence in situ hybridization (FISH) and AQUA, a novel, immunohistochemical method of automated quantitative in situ proteomic analysis which permits subcellular localization. Methods: A tissue microarray composed of 137 HNSCC treated with (chemo)radiation was constructed and analyzed for EGFR copy number by FISH (Vysis/Abbot) and EGFR protein expression (DAKO antibody) using AQUA analysis of EGFR staining scored on a scale of 0–255 and by conventional IHC. Agreement was assessed using kappa. Results: Sixteen (15%) of one-hundred six tumors with FISH results demonstrated EGFR high polysomy and/or gene amplification (FISH+). AQUA demonstrated a range of 3.6–102.2; protein levels assessed by AQUA in the FISH amplified cases were significantly higher (p =0.008) than in the FISH non- amplified ones. Using the EGFR 75th percentile as a cut-off, AQUA and FISH showed significant agreement (percentage of overall agreement 82%, kappa=0.458, p=0.003). To the contrary there was no concordance between FISH and conventional IHC results in this series. Conclusions: The discrepancy between EGFR gene amplification rate and protein expression by IHC reported previously may be due to the limitations and nonquantitative nature of conventional IHC. EGFR protein content correlates with gene copy number if protein content is quantitated and automatically analyzed, as with AQUA. No significant financial relationships to disclose.

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Defense by duplication: The relation between phenotypic glyphosate resistance and EPSPS gene copy number variation in Amaranthus palmeri

10.1101/2021.04.13.439524 ◽

2021 ◽

Author(s):

Sarah B Yakimowski ◽

Zachary Teitel ◽

Christina M. Caruso

Keyword(s):

Copy Number Variation ◽

Copy Number ◽

Gene Copy Number ◽

Glyphosate Resistance ◽

Gene Copy ◽

Quantitative Relation ◽

Stabilizing Selection ◽

Amaranthus Palmeri ◽

Gene Copy Number Variation ◽

Number Variation

Gene copy number variation (CNV) has been increasingly associated with organismal responses to environmental stress, but we know little about the quantitative relation between CNV and phenotypic variation. In this study we quantify variation in EPSPS (5-enolpyruvylshikimate-3-phosphate synthase) copy number using digital drop PCR and variation in phenotypic glyphosate resistance in 22 populations of Amaranthus palmeri (Palmer Amaranth), a range-expanding agricultural weed. Overall, we detected a significant positive relation between population mean copy number and mean resistance. The majority of populations exhibited high glyphosate resistance, yet maintained low-resistance individuals resulting in bimodality in many populations. We investigated linear and threshold models for the relation between copy number and resistance, and found evidence for a threshold of ~15 EPSPS copies: there was a steep increase in resistance before the threshold, followed by a much shallower slope. Moreover, as copy number increases, the range of variation in resistance decreases. This suggests a working hypothesis that as EPSPS copies and dosage increases, negative epistatic interactions may be compensated. We detected a quadratic relation between mean resistance and variation (s.d.) in resistance, consistent with the prediction that as phenotypic resistance increases in populations, stabilizing selection decreases variation in the trait. Finally, patterns of variation across the landscape are consistent with less variation among populations in mean copy number / resistance in Georgia where glyphosate resistance was first detected, and wider variation among populations in resistance and copy number in a more northern state where resistance evolution may be at a younger evolutionary state.

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Level of HER2 Gene Amplification Predicts Response and Overall Survival in HER2-Positive Advanced Gastric Cancer Treated With Trastuzumab

Journal of Clinical Oncology ◽

10.1200/jco.2013.48.9070 ◽

2013 ◽

Vol 31 (35) ◽

pp. 4445-4452 ◽

Cited By ~ 110

Author(s):

Carlos Gomez-Martin ◽

Jose Carlos Plaza ◽

Roberto Pazo-Cid ◽

Antonieta Salud ◽

Francesc Pons ◽

...

Keyword(s):

Gastric Cancer ◽

Overall Survival ◽

Advanced Gastric Cancer ◽

Gene Amplification ◽

Copy Number ◽

Gene Copy Number ◽

Gene Copy ◽

Her2 Gene ◽

Optimal Cutoff ◽

Her2 Gene Amplification

Purpose Previous studies have highlighted the importance of an appropriate human epidermal growth factor receptor 2 (HER2) evaluation for the proper identification of patients eligible for treatment with anti-HER2 targeted therapies. Today, the relationship remains unclear between the level of HER2 amplification and the outcome of HER2-positive gastric cancer treated with first-line chemotherapy with trastuzumab. The aim of this study was to determine whether the level of HER2 gene amplification determined by the HER2/CEP17 ratio and HER2 gene copy number could significantly predict some benefit in overall survival and response to therapy in advanced gastric cancer treated with trastuzumab-based chemotherapy. Patients and Methods Ninety patients with metastatic gastric cancer treated with first-line trastuzumab-based chemotherapy were studied. The optimal cutoff values for HER2/CEP17 ratio and HER2 gene copy number (GCN) for discriminating positive results in terms of response and prolonged survival were determined using receiver operating characteristic curves analyses. Results In this study, a median HER2/CEP17 ratio of 6.11 (95% CI, 2.27 to 21.90) and a median HER2 gene copy number of 11.90 (95% CI, 3.30 to 43.80) were found. A mean HER2/CEP17 ratio of 4.7 was identified as the optimal cutoff value discriminating sensitive and refractory patients (P = .005). Similarly, the optimal cutoff for predicting survival longer than 12 months was 4.45 (P = .005), and for survival longer than 16 months was 5.15 (P = .004). For HER2 GCN, the optimal cutoff values were 9.4, 10.0, and 9.5, respectively (P = .02). Conclusion The level of HER2 gene amplification significantly predicts sensitivity to therapy and overall survival in advanced gastric cancer treated with trastuzumab-based chemotherapy.

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