Standing genetic variation fuels rapid evolution of herbicide resistance in blackgrass.

Repeated herbicide applications in agricultural fields exert strong selection on weeds such as blackgrass (Alopecurus myosuroides), which is a major threat for temperate climate cereal crops. This inadvertent selection pressure provides an opportunity for investigating the underlying genetic mechanisms and evolutionary processes of rapid adaptation, which can occur both through mutations in the direct targets of herbicides and through changes in other, often metabolic, pathways, known as non-target-site resistance. How much target-site resistance (TSR) relies on de novo mutations vs. standing variation is important for developing strategies to manage herbicide resistance. We first generated a chromosome-level reference genome for A. myosuroides for population genomic studies of herbicide resistance and genome-wide diversity across Europe in this species. Next, through empirical data in the form of highly accurate long-read amplicons of alleles encoding acetyl-CoA carboxylase (ACCase) and acetolactate synthase (ALS) variants, we showed that most populations with resistance due to TSR mutations-23 out of 27 and six out of nine populations for ACCase and ALS, respectively-contained at least two TSR haplotypes, indicating that soft sweeps are the norm. Finally, through forward-in-time simulations, we inferred that TSR is likely to mainly result from standing genetic variation, with only a minor role for de novo mutations.

DAP5 enables main ORF translation on mRNAs with structured and uORF-containing 5' leaders.

Half of mammalian transcripts contain short upstream open reading frames (uORFs) that potentially regulate translation of the downstream coding sequence (CDS). The molecular mechanisms governing these events remain poorly understood. Here, we find that the non-canonical initiation factor Death-associated protein 5 (DAP5 or eIF4G2) is required for translation initiation on select transcripts. Using ribosome profiling and luciferase-based reporters coupled with mutational analysis we show that DAP5-mediated translation occurs on messenger RNAs (mRNAs) with long, structure-prone 5' leader sequences and persistent uORF translation. These mRNAs preferentially code for signalling factors such as kinases and phosphatases. We also report that cap/eIF4F- and eIF4A-dependent recruitment of DAP5 to the mRNA facilitates main CDS, but not uORF, translation suggesting a role for DAP5 in translation re-initiation. Our study reveals important mechanistic insights into how a non-canonical translation initiation factor involved in stem cell fate shapes the synthesis of specific signalling factors.

Characterization of monocyte subtypes regarding their phenotype and development in the context of graft-versus-host disease.

Graft-versus-host disease (GvHD) is a major cause of morbidity and mortality after allogeneic hematopoietic cell transplantation (HCT). In this study, monocyte subtypes were characterized regarding cytokine expression pattern and development in the context of GvHD. Using inflammatory S100 proteins for monocyte stimulation, it could be demonstrated that intermediate monocytes are the main producers of inflammatory cytokines such as IL-6 and TNFα known to be involved in the development of Th17 cells pointing towards an inflammatory phenotype of this monocyte subtype. Furthermore, novel aspects regarding monocyte subtype development were found. Our data reveal that prednisolone promotes the induction of intermediate monocytes from classical monocytes which correlates with HSP70 expression levels. However, 1α,25-Dihydroxyvitamin D3 treatment results in the abrogation of the prednisolone-mediated induction of this inflammatory monocyte subset and low HSP70 expression levels. Treatment of classical monocytes with pifithrin-µ, a specific HSP70 inhibitor, also leads to an inhibited induction of intermediate monocytes in the presence of prednisolone. These data point towards a predominant role of HSP70 in the development of intermediate monocytes. Thus, HSP70 might be a promising target for GvHD therapy, especially in combination with glucocorticoids, in order to decrease intermediate monocyte subset levels.

Increase of Intermediate Monocytes in Graft-versus-Host Disease: Correlation with MDR1+Th17.1 Levels and the Effect of Prednisolone and 1α,25-Dihydroxyvitamin D3.

Graft-versus-host disease (GVHD) remains one of the major complications after allogeneic hematopoietic stem cell transplantation that is mainly treated with glucocorticoids such as prednisolone. In this study the influence of monocyte subpopulations, prednisolone, and 1α,25-dihydroxyvitamin D3 (1α,25-(OH)2D3) on the induction of a proinflammatory subset of Th17 cells (MDR+Th17.1) characterized by CCR6+CXCR3hiCCR4loCCR10-CD161+ and stable expression of the multidrug resistance protein type 1 (MDR1) was investigated. Our results demonstrate that intermediate monocytes are increased in patients with acute GVHD, promoting the induction of proinflammatory MDR1+Th17.1 cells. Furthermore, prednisolone induces the development of MDR1+Th17.1 cells, whereas 1α,25-(OH)2D3 acts as an anti-inflammatory, leading to diminished percentages of proinflammatory MDR1+Th17.1 cells in the presence of prednisolone after stimulation with the TLR4-ligand S100A8/S100A9. Moreover, 1α,25-(OH)2D3 decreased the expression level of the targets JAK2 and CD74, both associated with T cell activation, in monocytes. Thus, in steroid-resistant GVHD, 1α,25-(OH)2D3 could be an important regulator in monocyte-induced development of proinflammatory MDR1+Th17.1 cells and might therefore be a potential therapeutic agent in combination with glucocorticoids for GVHD treatment.