Dynamic Phytomeric Growth Contributes to Local Adaptation in Barley.

Vascular plants have segmented body axes with iterative nodes and internodes. Appropriate node initiation and internode elongation are fundamental to plant fitness and crop yield; however, how these events are spatiotemporally coordinated remains elusive. We show that in barley (Hordeum vulgare L.), selections during domestication have extended the apical meristematic phase to promote node initiation, but constrained subsequent internode elongation. In both vegetative and reproductive phases, internode elongation displays a dynamic proximal-distal gradient, and among subpopulations of domesticated barleys worldwide, node initiation and proximal internode elongation are associated with latitudinal and longitudinal gradients, respectively. Genetic and functional analyses suggest that, in addition to their converging roles in node initiation, flowering-time genes have been repurposed to specify the timing and duration of internode elongation. Our study provides an integrated view of barley node initiation and internode elongation and suggests that plant architecture should be recognized as a collection of dynamic phytomeric units in the context of crop adaptive evolution.

Combining CRISPR-Cas9 and brain imaging to study the link from genes to molecules to networks.

Receptors, transporters, and ion channels are important targets for therapy development in neurological diseases, but their mechanistic role in pathogenesis is often poorly understood. Gene editing and in vivo imaging approaches will help to identify the molecular and functional role of these targets and the consequence of their regional dysfunction on the whole-brain level. We combine CRISPR-Cas9 gene editing with in vivo positron emission tomography (PET) and functional MRI (fMRI) to investigate the direct link between genes, molecules, and the brain connectome. The extensive knowledge of the Slc18a2 gene encoding the vesicular monoamine transporter (VMAT2), involved in the storage and release of dopamine, makes it an excellent target for studying the gene network relationships while structurally preserving neuronal integrity and function. We edited the Slc18a2 in the substantia nigra pars compacta of adult rats and used in vivo molecular imaging besides behavioral, histological, and biochemical assessments to characterize the CRISPR-Cas9-mediated VMAT2 knockdown. Simultaneous PET/fMRI was performed to investigate molecular and functional brain alterations. We found that stage-specific adaptations of brain functional connectivity follow the selective impairment of presynaptic dopamine storage and release. Our study reveals that recruiting different brain networks is an early response to the dopaminergic dysfunction preceding neuronal cell loss. Our combinatorial approach is a tool to investigate the impact of specific genes on brain molecular and functional dynamics, which will help to develop tailored therapies for normalizing brain function.

Western diet increases brain metabolism and adaptive immune responses in a mouse model of amyloidosis.

Diet-induced increase in body weight is a growing health concern worldwide. Often accompanied by a low-grade metabolic inflammation that changes systemic functions, diet-induced alterations may contribute to neurodegenerative disorder progression as well. This study aims to non-invasively investigate diet-induced metabolic and inflammatory effects in the brain of an APPPS1 mouse model of Alzheimer's disease. [18F]FDG, [18F]FTHA, and [18F]GE-180 were used for in vivo PET imaging in wild-type and APPPS1 mice. Ex vivo flow cytometry and histology in brains complemented the in vivo findings. 1H- magnetic resonance spectroscopy in the liver, plasma metabolomics and flow cytometry of the white adipose tissue were used to confirm metaflammatory condition in the periphery. We found disrupted glucose and fatty acid metabolism after Western diet consumption, with only small regional changes in glial-dependent neuroinflammation in the brains of APPPS1 mice. Further ex vivo investigations revealed cytotoxic T cell involvement in the brains of Western diet-fed mice and a disrupted plasma metabolome. 1H-magentic resonance spectroscopy and immunological results revealed diet-dependent inflammatory-like misbalance in livers and fatty tissue. Our multimodal imaging study highlights the role of the brain-liver-fat axis and the adaptive immune system in the disruption of brain homeostasis in amyloid models of Alzheimer's disease.

[18F]pFBC, a Covalent CLIP-Tag Radiotracer for Detection of Viral Reporter Gene Transfer in the Murine Brain.

Preclinical models of neurological diseases and gene therapy are essential for neurobiological research. However, the evaluation of such models lacks reliable reporter systems for use with noninvasive imaging methods. Here, we report the development of a reporter system based on the CLIP-tag enzyme and [18F]pFBC, an 18F-labeled covalent CLIP-tag-ligand synthesized via a DoE-optimized and fully automated process. We demonstrated its specificity using a subcutaneous xenograft model and a model of viral vector-mediated brain gene transfer by engineering HEK293 cells and striatal neurons to express membrane-tethered CLIP-tag protein. After in vitro characterization of the reporter, mice carrying either CLIP-tag expressing or control subcutaneous xenografts underwent dynamic [18F]pFBC PET imaging. The CLIP-tag expressing xenografts showed a significantly higher uptake than control xenografts (tumor-to-muscle ratio 5.0 vs 1.7, p = 0.0379). In vivo, metabolite analysis by radio-HPLC from plasma and brain homogenates showed only one radio-metabolite in plasma and none in the brain. In addition, [18F]pFBC showed fast uptake and rapid clearance from the brain in animals injected with adeno-associated virus (AAV)-CLIP in the right striatum but no right-to-left (R-L) uptake difference in the striata in the acquired PET data. In contrast, autoradiography showed a clear accumulation of radioactivity in the AAV-CLIP-injected right striatum compared to the sham-injected left striatum control. CLIP-tag expression and brain integrity were verified by immunofluorescence and light sheet microscopy. In conclusion, we established a novel reporter gene system for PET imaging of gene expression in the brain and periphery and demonstrated its potential for a wide range of applications, particularly for neurobiological research and gene therapy with viral vectors.

Investigating the genetic control of plant development in spring barley under speed breeding conditions.

KEY MESSAGE: This study found that the genes, PPD-H1 and ELF3, control the acceleration of plant development under speed breeding, with important implications for optimizing the delivery of climate-resilient crops. Speed breeding is a tool to accelerate breeding and research programmes. Despite its success and growing popularity with breeders, the genetic basis of plant development under speed breeding remains unknown. This study explored the developmental advancements of barley genotypes under different photoperiod regimes. A subset of the HEB-25 Nested Association Mapping population was evaluated for days to heading and maturity under two contrasting photoperiod conditions: (1) Speed breeding (SB) consisting of 22 h of light and 2 h of darkness, and (2) normal breeding (NB) consisting of 16 h of light and 8 h of darkness. GWAS revealed that developmental responses under both conditions were largely controlled by two loci: PPDH-1 and ELF3. Allelic variants at these genes determine whether plants display early flowering and maturity under both conditions. At key QTL regions, domesticated alleles were associated with late flowering and maturity in NB and early flowering and maturity in SB, whereas wild alleles were associated with early flowering under both conditions. We hypothesize that this is related to the dark-dependent repression of PPD-H1 by ELF3 which might be more prominent in NB conditions. Furthermore, by comparing development under two photoperiod regimes, we derived an estimate of plasticity for the two traits. Interestingly, plasticity in development was largely attributed to allelic variation at ELF3. Our results have important implications for our understanding and optimization of speed breeding protocols particularly for introgression breeding and the design of breeding programmes to support the delivery of climate-resilient crops.

Open-source flow setup for rapid and efficient [18 F]fluoride drying for automation of PET tracer syntheses.

One of the key strategies for radiochemical research facilities is the automation of synthesis processes. Unnecessary manual operations increase the radiation exposure of personnel, while simultaneously threatening the reliability of syntheses. We have previously reported an affordable open-source system comprising 3D-printed continuous flow reactors, a custom syringe pump, and a pressure regulator that can be used to perform radiofluorinations. In this paper, we address additional essential processes that are needed for radiotracer development and synthesis, with the aim of making laboratory work safer and research more efficient. We have designed and evaluated a fully automated system for rapidly and effectively processing and drying aqueous [18 F]fluoride that can be directly connected to the cyclotron. This process relies on triflyl fluoride gas generation and allows nucleophilic [18 F]fluoride to be prepared safely in a hotcell within 10 min and an activity recovery of 91.7 ± 1.6% (n = 5). Owing to the need for convenient radiofluorinated prosthetic ligands, we have adapted our continuous flow system to produce [18 F]fluoroethyl tosylate (FEOTs) and [18 F]fluoroethyl triflate (FEOTf), prosthetic groups that are widely used for late-stage fluoroethylation of PET tracers. The processes as well as the radiolabeling of different groups are compared and comprehensively discussed. Having a method providing [18 F]fluoroethyl tosylate (FEOTs) as well as [18 F]fluoroethyl triflate (FEOTf) quickly and highly efficiently is beneficial for radiochemical research.

Acidosis-mediated increase in IFN-γ-induced PD-L1 expression on cancer cells as an immune escape mechanism in solid tumors.

Immune checkpoint inhibitors have revolutionized cancer therapy, yet the efficacy of these treatments is often limited by the heterogeneous and hypoxic tumor microenvironment (TME) of solid tumors. In the TME, programmed death-ligand 1 (PD-L1) expression on cancer cells is mainly regulated by Interferon-gamma (IFN-γ), which induces T cell exhaustion and enables tumor immune evasion. In this study, we demonstrate that acidosis, a common characteristic of solid tumors, significantly increases IFN-γ-induced PD-L1 expression on aggressive cancer cells, thus promoting immune escape. Using preclinical models, we found that acidosis enhances the genomic expression and phosphorylation of signal transducer and activator of transcription 1 (STAT1), and the translation of STAT1 mRNA by eukaryotic initiation factor 4F (elF4F), resulting in an increased PD-L1 expression. We observed this effect in murine and human anti-PD-L1-responsive tumor cell lines, but not in anti-PD-L1-nonresponsive tumor cell lines. In vivo studies fully validated our in vitro findings and revealed that neutralizing the acidic extracellular tumor pH by sodium bicarbonate treatment suppresses IFN-γ-induced PD-L1 expression and promotes immune cell infiltration in responsive tumors and thus reduces tumor growth. However, this effect was not observed in anti-PD-L1-nonresponsive tumors. In vivo experiments in tumor-bearing IFN-γ-/- mice validated the dependency on immune cell-derived IFN-γ for acidosis-mediated cancer cell PD-L1 induction and tumor immune escape. Thus, acidosis and IFN-γ-induced elevation of PD-L1 expression on cancer cells represent a previously unknown immune escape mechanism that may serve as a novel biomarker for anti-PD-L1/PD-1 treatment response. These findings have important implications for the development of new strategies to enhance the efficacy of immunotherapy in cancer patients.

Recombination landscape divergence between populations is marked by larger low-recombining regions in domesticated rye.

The genomic landscape of recombination plays an essential role in evolution. Patterns of recombination are highly variable along chromosomes, between sexes, individuals, populations, and species. In many eukaryotes, recombination rates are elevated in sub-telomeric regions and drastically reduced near centromeres, resulting in large low-recombining (LR) regions. The processes of recombination are influenced by genetic factors, such as different alleles of genes involved in meiosis and chromatin structure, as well as external environmental stimuli like temperature and overall stress. In this work, we focused on the genomic landscapes of recombination in a collection of 916 rye (Secale cereale) individuals. By analysing population structure among individuals of different domestication status and geographic origin, we detected high levels of admixture, reflecting the reproductive biology of a self-incompatible, wind-pollinating grass species. We then analysed patterns of recombination in overlapping subpopulations, which revealed substantial variation in the physical size of LR regions, with a tendency for larger LR regions in domesticated subpopulations. Genome-wide association scans (GWAS) for LR region size revealed a major quantitative-trait-locus (QTL) at which, among 18 annotated genes, an ortholog of histone H4 acetyltransferase ESA1 was located. Rye individuals belonging to domesticated subpopulations showed increased synaptonemal complex length, but no difference in crossover frequency, indicating that only the recombination landscape is different. Furthermore, the genomic region harbouring rye ScESA1 showed moderate patterns of selection in domesticated subpopulations, suggesting that larger LR regions were indirectly selected for during domestication to achieve more homogeneous populations for agricultural use.

An exotic allele of barley EARLY FLOWERING 3 contributes to developmental plasticity at elevated temperatures.

Increase in ambient temperatures caused by climate change affects various morphological and developmental traits of plants, threatening crop yield stability. In the model plant Arabidopsis thaliana, EARLY FLOWERING 3 (ELF3) plays prominent roles in temperature sensing and thermomorphogenesis signal transduction. However, how crop species respond to elevated temperatures is poorly understood. Here, we show that the barley ortholog of AtELF3 interacts with high temperature to control growth and development. We used heterogeneous inbred family (HIF) pairs generated from a segregating mapping population and systematically studied the role of exotic ELF3 variants in barley temperature responses. An exotic ELF3 allele of Syrian origin promoted elongation growth in barley at elevated temperatures, whereas plant area and estimated biomass were drastically reduced, resulting in an open canopy architecture. The same allele accelerated inflorescence development at high temperature, which correlated with early transcriptional induction of MADS-box floral identity genes BM3 and BM8. Consequently, barley plants carrying the exotic ELF3 allele displayed stable total grain number at elevated temperatures. Our findings therefore demonstrate that exotic ELF3 variants can contribute to phenotypic and developmental acclimation to elevated temperatures, providing a stimulus for breeding of climate-resilient crops.

Novel exotic alleles of EARLY FLOWERING 3 determine plant development in barley.

EARLY FLOWERING 3 (ELF3) is an important regulator of various physiological and developmental processes and hence may serve to improve plant adaptation which will be essential for future plant breeding. To expand the limited knowledge on barley ELF3 in determining agronomic traits, we conducted field studies with heterogeneous inbred families (HIFs) derived from selected lines of the wild barley nested association mapping population HEB-25. During two growing seasons, phenotypes of nearly isogenic HIF sister lines, segregating for exotic and cultivated alleles at the ELF3 locus, were compared for 10 developmental and yield-related traits. We determine novel exotic ELF3 alleles and show that HIF lines, carrying the exotic ELF3 allele, accelerated plant development compared with the cultivated ELF3 allele, depending on the genetic background. Remarkably, the most extreme effects on phenology could be attributed to one exotic ELF3 allele differing from the cultivated Barke ELF3 allele in only one single nucleotide polymorphism (SNP). This SNP causes an amino acid substitution (W669G), which as predicted has an impact on the protein structure of ELF3. Consequently, it may affect phase separation behaviour and nano-compartment formation of ELF3 and, potentially, also its local cellular interactions causing significant trait differences between HIF sister lines.

The Structural Combination of SIL and MODAG Scaffolds Fails to Enhance Binding to α-Synuclein but Reveals Promising Affinity to Amyloid ß.

A technique to image α-synuclein (αSYN) fibrils in vivo is an unmet scientific and clinical need that would represent a transformative tool in the understanding, diagnosis, and treatment of various neurodegenerative diseases. Several classes of compounds have shown promising results as potential PET tracers, but no candidate has yet exhibited the affinity and selectivity required to reach clinical application. We hypothesized that the application of the rational drug design technique of molecular hybridization to two promising lead scaffolds could enhance the binding to αSYN up to the fulfillment of those requirements. By combining the structures of SIL and MODAG tracers, we developed a library of diarylpyrazoles (DAPs). In vitro evaluation through competition assays against [3H]SIL26 and [3H]MODAG-001 showed the novel hybrid scaffold to have preferential binding affinity for amyloid ß (Aß) over αSYN fibrils. A ring-opening modification on the phenothiazine building block to produce analogs with increased three-dimensional flexibility did not result in an improved αSYN binding but a complete loss of competition, as well as a significant reduction in Aß affinity. The combination of the phenothiazine and the 3,5-diphenylpyrazole scaffolds into DAP hybrids did not generate an enhanced αSYN PET tracer lead compound. Instead, these efforts identified a scaffold for promising Aß ligands that may be relevant to the treatment and monitoring of Alzheimer's disease (AD).

Two experts and a newbie: [18F]PARPi vs [18F]FTT vs [18F]FPyPARP-a comparison of PARP imaging agents.

PURPOSE: Imaging of PARP expression has emerged as valuable strategy for prediction of tumor malignancy. While [18F]PARPi and [18F]FTT are already in clinical translation, both suffer from mainly hepatobiliary clearance hampering their use for detection of abdominal lesions, e.g., liver metastases. Our novel radiotracer [18F]FPyPARP aims to bridge this gap with a higher renal clearance and an easily translatable synthesis route for potential clinical application. METHODS: We developed a less lipophilic variant of [18F]PARPi by exchange of the fluorobenzoyl residue with a fluoronicotinoyl group and automated the radiosyntheses of the three radiotracers. We then conducted a comparative side-by-side study of [18F]PARPi, [18F]FPyPARP, and [18F]FTT in NOD.CB17-Prkdcscid/J mice bearing HCC1937 xenografts to assess xenograft uptake and pharmacokinetics focusing on excretion pathways. RESULTS: Together with decent uptake of all three radiotracers in the xenografts (tumor-to-blood ratios 3.41 ± 0.83, 3.99 ± 0.99, and 2.46 ± 0.35, respectively, for [18F]PARPi, [18F]FPyPARP, and [18F]FTT), a partial shift from hepatobiliary to renal clearance of [18F]FPyPARP was observed, whereas [18F]PARPi and [18F]FTT show almost exclusive hepatobiliary clearance. CONCLUSION: These findings imply that [18F]FPyPARP is an alternative to [18F]PARPi and [18F]FTT for PET imaging of PARP enzymes.

Genome-Wide Association Study of Salt Tolerance-Related Traits during Germination and Seedling Development in an Intermedium-Spike Barley Collection.

Increased salinity is one of the major consequences of climatic change affecting global crop production. The early stages in the barley (Hordeum vulgare L.) life cycle are considered the most critical phases due to their contributions to final crop yield. Particularly, the germination and seedling development are sensitive to numerous environmental stresses, especially soil salinity. In this study, we aimed to identify SNP markers linked with germination and seedling development at 150 mM NaCl as a salinity treatment. We performed a genome-wide association study (GWAS) using a panel of 208 intermedium-spike barley (H. vulgare convar. intermedium (Körn.) Mansf.) accessions and their genotype data (i.e., 10,323 SNPs) using the genome reference sequence of "Morex". The phenotypic results showed that the 150 mM NaCl salinity treatment significantly reduced all recorded germination and seedling-related traits compared to the control treatment. Furthermore, six accessions (HOR 11747, HOR 11718, HOR 11640, HOR 11256, HOR 11275 and HOR 11291) were identified as the most salinity tolerant from the intermedium-spike barley collection. GWAS analysis indicated that a total of 38 highly significantly associated SNP markers under control and/or salinity traits were identified. Of these, two SNP markers on chromosome (chr) 1H, two on chr 3H, and one on chr 4H were significantly linked to seedling fresh and dry weight under salinity stress treatment. In addition, two SNP markers on chr 7H were also significantly associated with seedling fresh and dry weight but under control condition. Under salinity stress, one SNP marker on chr 1H, 5H and 7H were detected for more than one phenotypic trait. We found that in most of the accessions exhibiting the highest salinity tolerance, most of the salinity-related QTLs were presented. These results form the basis for detailed studies, leading to improved salt tolerance breeding programs in barley.

[Tyrolean stroke pathway]. / Schlaganfallpfad Tirol.

Background: Recent advances in acute stroke therapy have been shown to improve clinical outcome and therefore reduce long-term disability. Acute therapy procedures in stroke have a limited time frame after onset of symptoms; thus, rapid emergency management is critical. The Tyrolean Stroke Pathway was developed to optimize the entire treatment pathway from stroke onset to outpatient rehabilitation. Objectives: The description of the Tyrolean Stroke Pathway as an example to optimize the entire treatment pathway of stroke patients in a central European alpine region with a focus on prehospital management. Materials and methods: In four subprojects from the prehospital phase up to outpatient rehabilitation, the project optimized intersections between different phases of care and evaluated the care processes using quality assurance measures. Results: After implementation, thrombolysis rates doubled and functional outcome of stroke patients after 3 months improved and complications, e.g., pneumonia, declined. Access to rehabilitation improved, particularly because of the establishment of quality assured and funded outpatient rehabilitation. Conclusions: The project showed that a holistic standardized stroke pathway from acute management up to outpatient rehabilitation can be successfully implemented in clinical routine. The project improved the entire health care pathway of stroke patients. The Stroke Action Plan for Europe 2018-2030 recommends to have national plans for stroke that encompass the entire chain of care.

Striatal and prefrontal D2R and SERT distributions contrastingly correlate with default-mode connectivity.

Although brain research has taken important strides in recent decades, the interaction and coupling of its different physiological levels is still not elucidated. Specifically, the molecular substrates of resting-state functional connectivity (rs-FC) remain poorly understood. The aim of this study was elucidating interactions between dopamine D2 receptors (D2R) and serotonin transporter (SERT) availabilities in the striatum (CPu) and medial prefrontal cortex (mPFC), two of the main dopaminergic and serotonergic projection areas, and the default-mode network. Additionally, we delineated its interaction with two other prominent resting-state networks (RSNs), the salience network (SN) and the sensorimotor network (SMN). To this extent, we performed simultaneous PET/fMRI scans in a total of 59 healthy rats using [11C]raclopride and [11C]DASB, two tracers used to image quantify D2R and SERT respectively. Edge, node and network-level rs-FC metrics were calculated for each subject and potential correlations with binding potentials (BPND) in the CPu and mPFC were evaluated. We found widespread negative associations between CPu D2R availability and all the RSNs investigated, consistent with the postulated role of the indirect basal ganglia pathway. Correlations between D2Rs in the mPFC were weaker and largely restricted to DMN connectivity. Strikingly, medial prefrontal SERT correlated both positively with anterior DMN rs-FC and negatively with rs-FC between and within the SN, SMN and the posterior DMN, underlining the complex role of serotonergic neurotransmission in this region. Here we show direct relationships between rs-FC and molecular properties of the brain as assessed by simultaneous PET/fMRI in healthy rodents. The findings in the present study contribute to the basic understanding of rs-FC by revealing associations between inter-subject variances of rs-FC and receptor and transporter availabilities. Additionally, since current therapeutic strategies typically target neurotransmitter systems with the aim of normalizing brain function, delineating associations between molecular and network-level brain properties is essential and may enhance the understanding of neuropathologies and support future drug development.

[11C]MODAG-001-towards a PET tracer targeting α-synuclein aggregates.

PURPOSE: Deposition of misfolded alpha-synuclein (αSYN) aggregates in the human brain is one of the major hallmarks of synucleinopathies. However, a target-specific tracer to detect pathological aggregates of αSYN remains lacking. Here, we report the development of a positron emission tomography (PET) tracer based on anle138b, a compound shown to have therapeutic activity in animal models of neurodegenerative diseases. METHODS: Specificity and selectivity of [3H]MODAG-001 were tested in in vitro binding assays using recombinant fibrils. After carbon-11 radiolabeling, the pharmacokinetic and metabolic profile was determined in mice. Specific binding was quantified in rats, inoculated with αSYN fibrils and using in vitro autoradiography in human brain sections of Lewy body dementia (LBD) cases provided by the Neurobiobank Munich (NBM). RESULTS: [3H]MODAG-001 revealed a very high affinity towards pure αSYN fibrils (Kd = 0.6 ± 0.1 nM) and only a moderate affinity to hTau46 fibrils (Kd = 19 ± 6.4 nM) as well as amyloid-ß1-42 fibrils (Kd = 20 ± 10 nM). [11C]MODAG-001 showed an excellent ability to penetrate the mouse brain. Metabolic degradation was present, but the stability of the parent compound improved after selective deuteration of the precursor. (d3)-[11C]MODAG-001 binding was confirmed in fibril-inoculated rat striata using in vivo PET imaging. In vitro autoradiography showed no detectable binding to aggregated αSYN in human brain sections of LBD cases, most likely, because of the low abundance of aggregated αSYN against background protein. CONCLUSION: MODAG-001 provides a promising lead structure for future compound development as it combines a high affinity and good selectivity in fibril-binding assays with suitable pharmacokinetics and biodistribution properties.

Scalable 18F processing conditions for copper-mediated radiofluorination chemistry facilitate DoE optimization studies and afford an improved synthesis of [18F]olaparib.

A convenient and scalable base-free method for processing [18F]fluoride as [18F]TBAF is reported and applied to copper-mediated radiofluorination radiosyntheses. A central feature of this method is that a single production of [18F]TBAF can be divided into small aliquots that can be used to perform multiple small-scale reactions in DoE optimization studies. The results of these studies can then be reliably translated to full batch tracer productions using automated synthesizers. The processing method was applied to the DoE optimization of [18F]olaparib, affording the tracer in high radiochemical yields via both manual (%RCY = 78 ± 6%, n = 4 (CMRF step only)) and automated (up to 80% (%RCY); 41% activity yield (%AY)) radiosynthesis procedures.

Identification of QTLs conferring resistance to scald (Rhynchosporium commune) in the barley nested association mapping population HEB-25.

BACKGROUND: Barley scald, caused by the fungus Rhynchosporium commune, is distributed worldwide to all barley growing areas especially in cool and humid climates. Scald is an economically important leaf disease resulting in yield losses of up to 40%. To breed resistant cultivars the identification of quantitative trait loci (QTLs) conferring resistance to scald is necessary. Introgressing promising resistance alleles of wild barley is a way to broaden the genetic basis of scald resistance in cultivated barley. Here, we apply nested association mapping (NAM) to map resistance QTLs in the barley NAM population HEB-25, comprising 1420 lines in BC1S3 generation, derived from crosses of 25 wild barley accessions with cv. Barke. RESULTS: In scald infection trials in the greenhouse variability of resistance across and within HEB-25 families was found. NAM based on 33,005 informative SNPs resulted in the identification of eight reliable QTLs for resistance against scald with most wild alleles increasing resistance as compared to cv. Barke. Three of them are located in the region of known resistance genes and two in the regions of QTLs, respectively. The most promising wild allele was found at Rrs17 in one specific wild donor. Also, novel QTLs with beneficial wild allele effects on scald resistance were detected. CONCLUSIONS: To sum up, wild barley represents a rich resource for scald resistance. As the QTLs were linked to the physical map the identified candidate genes will facilitate cloning of the scald resistance genes. The closely linked flanking molecular markers can be used for marker-assisted selection of the respective resistance genes to integrate them in elite cultivars.

Grassystatin-derived peptides selectively inhibit cathepsin E and have low affinity to cathepsin D.

Aspartic proteases are important biomarkers of human disease and interesting targets for modulation of immune response via MHC class II antigen processing inhibition. The lack of inhibitors with sufficient selectivity hampers precise analysis of the role of cathepsin E and napsin A in samples containing the ubiquitous and highly abundant homolog cathepsin D. Grassystatins from marine cyanobacteria show promising selectivity for cathepsin E but contain several ester bonds that make their synthesis cumbersome and thus limit availability of the inhibitors. Herewith, we present grassystatin-derived cathepsin E inhibitors with greatly facilitated synthesis but retained selectivity profile. We demonstrate their affinity and selectivity with both enzyme kinetic assays and streptavidin-based pull-down from cells and mouse organs. Our findings suggest that grassystatin-like inhibitors are useful tools for targeted inhibition of cathepsin E and thus provide a novel approach for cancer and immunology research.

Natural variation in meiotic recombination rate shapes introgression patterns in intraspecific hybrids between wild and domesticated barley.

Meiotic recombination rates vary considerably between species, populations and individuals. The genetic exchange between homologous chromosomes plays a major role in evolution by breaking linkage between advantageous and deleterious alleles in the case of introgressions. Identifying recombination rate modifiers is thus of both fundamental and practical interest to understand and utilize variation in meiotic recombination rates. We investigated recombination rate variation in a large intraspecific hybrid population (named HEB-25) derived from a cross between domesticated barley and 25 wild barley accessions. We observed quantitative variation in total crossover number with a maximum of a 1.4-fold difference between subpopulations and increased recombination rates across pericentromeric regions. The meiosis-specific α-kleisin cohesin subunit REC8 was identified as a candidate gene influencing crossover number and patterning. Furthermore, we quantified wild barley introgression patterns and revealed how local and genome-wide recombination rate variation shapes patterns of introgression. The identification of allelic variation in REC8 in combination with the observed changes in crossover patterning suggest a difference in how chromatin loops are tethered to the chromosome axis, resulting in reduced crossover suppression across pericentromeric regions. Local and genome-wide recombination rate variation is shaping patterns of introgressions and thereby directly influences the consequences of linkage drag.