Cistrome and Epicistrome Features Shape the Regulatory DNA Landscape.

The cistrome is the complete set of transcription factor (TF) binding sites (cis-elements) in an organism, while an epicistrome incorporates tissue-specific DNA chemical modifications and TF-specific chemical sensitivities into these binding profiles. Robust methods to construct comprehensive cistrome and epicistrome maps are critical for elucidating complex transcriptional networks that underlie growth, behavior, and disease. Here, we describe DNA affinity purification sequencing (DAP-seq), a high-throughput TF binding site discovery method that interrogates genomic DNA with in-vitro-expressed TFs. Using DAP-seq, we defined the Arabidopsis cistrome by resolving motifs and peaks for 529 TFs. Because genomic DNA used in DAP-seq retains 5-methylcytosines, we determined that >75% (248/327) of Arabidopsis TFs surveyed were methylation sensitive, a property that strongly impacts the epicistrome landscape. DAP-seq datasets also yielded insight into the biology and binding site architecture of numerous TFs, demonstrating the value of DAP-seq for cost-effective cistromic and epicistromic annotation in any organism.

Determination and inference of eukaryotic transcription factor sequence specificity.

Transcription factor (TF) DNA sequence preferences direct their regulatory activity, but are currently known for only ∼1% of eukaryotic TFs. Broadly sampling DNA-binding domain (DBD) types from multiple eukaryotic clades, we determined DNA sequence preferences for >1,000 TFs encompassing 54 different DBD classes from 131 diverse eukaryotes. We find that closely related DBDs almost always have very similar DNA sequence preferences, enabling inference of motifs for ∼34% of the ∼170,000 known or predicted eukaryotic TFs. Sequences matching both measured and inferred motifs are enriched in chromatin immunoprecipitation sequencing (ChIP-seq) peaks and upstream of transcription start sites in diverse eukaryotic lineages. SNPs defining expression quantitative trait loci in Arabidopsis promoters are also enriched for predicted TF binding sites. Importantly, our motif "library" can be used to identify specific TFs whose binding may be altered by human disease risk alleles. These data present a powerful resource for mapping transcriptional networks across eukaryotes.

The retrograde signaling regulator ANAC017 recruits the MKK9-MPK3/6, ethylene, and auxin signaling pathways to balance mitochondrial dysfunction with growth.

In plant cells, mitochondria are ideally positioned to sense and balance changes in energy metabolism in response to changing environmental conditions. Retrograde signaling from mitochondria to the nucleus is crucial for adjusting the required transcriptional responses. We show that ANAC017, the master regulator of mitochondrial stress, directly recruits a signaling cascade involving the plant hormones ethylene and auxin as well as the MAP KINASE KINASE (MKK) 9-MAP KINASE (MPK) 3/6 pathway in Arabidopsis thaliana. Chromatin immunoprecipitation followed by sequencing and overexpression demonstrated that ANAC017 directly regulates several genes of the ethylene and auxin pathways, including MKK9, 1-AMINO-CYCLOPROPANE-1-CARBOXYLATE SYNTHASE 2, and YUCCA 5, in addition to genes encoding transcription factors regulating plant growth and stress responses such as BASIC REGION/LEUCINE ZIPPER MOTIF (bZIP) 60, bZIP53, ANAC081/ATAF2, and RADICAL-INDUCED CELL DEATH1. A time-resolved RNA-seq experiment established that ethylene signaling precedes the stimulation of auxin signaling in the mitochondrial stress response, with a large part of the transcriptional regulation dependent on ETHYLENE-INSENSITIVE 3. These results were confirmed by mutant analyses. Our findings identify the molecular components controlled by ANAC017, which integrates the primary stress responses to mitochondrial dysfunction with whole plant growth via the activation of regulatory and partly antagonistic feedback loops.

Spatially resolved transcriptomic analysis of the germinating barley grain.

Seeds are a vital source of calories for humans and a unique stage in the life cycle of flowering plants. During seed germination, the embryo undergoes major developmental transitions to become a seedling. Studying gene expression in individual seed cell types has been challenging due to the lack of spatial information or low throughput of existing methods. To overcome these limitations, a spatial transcriptomics workflow was developed for germinating barley grain. This approach enabled high-throughput analysis of spatial gene expression, revealing specific spatial expression patterns of various functional gene categories at a sub-tissue level. This study revealed over 14 000 genes differentially regulated during the first 24 h after imbibition. Individual genes, such as the aquaporin gene family, starch degradation, cell wall modification, transport processes, ribosomal proteins and transcription factors, were found to have specific spatial expression patterns over time. Using spatial autocorrelation algorithms, we identified auxin transport genes that had increasingly focused expression within subdomains of the embryo over time, suggesting their role in establishing the embryo axis. Overall, our study provides an unprecedented spatially resolved cellular map for barley germination and identifies specific functional genomics targets to better understand cellular restricted processes during germination. The data can be viewed at https://spatial.latrobe.edu.au/.

Spatiotemporal deposition of cell wall polysaccharides in oat endosperm during grain development.

Oat (Avena sativa) is a cereal crop whose grains are rich in (1,3;1,4)-ß-D-glucan (mixed-linkage glucan or MLG), a soluble dietary fiber. In our study, we analyzed oat endosperm development in 2 Canadian varieties with differing MLG content and nutritional value. We confirmed that oat undergoes a nuclear type of endosperm development but with a shorter cellularization phase than barley (Hordeum vulgare). Callose and cellulose were the first polysaccharides to be detected in the early anticlinal cell walls at 11 days postemergence (DPE) of the panicle. Other polysaccharides such as heteromannan and homogalacturonan were deposited early in cellularization around 12 DPE after the first periclinal walls are laid down. In contrast to barley, heteroxylan deposition coincided with completion of cellularization and was detected from 14 DPE but was only detectable after demasking. Notably, MLG was the last polysaccharide to be laid down at 18 DPE within the differentiation phase, rather than during cellularization. In addition, differences in the spatiotemporal patterning of MLG were also observed between the 2 varieties. The lower MLG-containing cultivar AC Morgan (3.5% w/w groats) was marked by the presence of a discontinuous pattern of MLG labeling, while labeling in the same walls in CDC Morrison (5.6% w/w groats) was mostly even and continuous. RNA-sequencing analysis revealed higher transcript levels of multiple MLG biosynthetic cellulose synthase-like F (CSLF) and CSLH genes during grain development in CDC Morrison compared with AC Morgan that likely contributes to the increased abundance of MLG at maturity in CDC Morrison. CDC Morrison was also observed to have smaller endosperm cells with thicker walls than AC Morgan from cellularization onwards, suggesting the processes controlling cell size and shape are established early in development. This study has highlighted that the molecular processes influencing MLG content and deposition are more complex than previously imagined.

Genomic and Cell-Specific Regulation of Benzylisoquinoline Alkaloid Biosynthesis in Opium Poppy.

Opium poppy is a crop of great commercial value as a source of several opium alkaloids for the pharmaceutical industries including morphine, codeine, thebaine, noscapine and papaverine. Most enzymes involved in benzylisoquinoline alkaloids (BIAs) biosynthesis in opium poppy have been functionally characterized, and opium poppy currently serves as a model system to study BIA metabolism in plants. BIA biosynthesis in opium poppy involves two biosynthetic gene clusters associated respectively with the morphine and noscapine branches. Recent reports have shown that genes in the same cluster are co-expressed, suggesting they might also be co-regulated. However, the transcriptional regulation of opium poppy BIA biosynthesis is not well studied. Opium poppy BIA biosynthesis involves three cell types associated with the phloem system: companion cells, sieve elements and laticifers. The transcripts and enzymes associated with BIA biosynthesis are distributed across cell types, requiring the translocation of key enzymes and pathway intermediates between cell types. Together, these suggest that the regulation of BIA biosynthesis in opium poppy is multilayered and complex, involving biochemical, genomic, and physiological mechanisms. In this review, we highlight recent advances in genome sequencing and single cell and spatial transcriptomics with a focus on how these efforts can improve our understanding of the genomic and cell-specific regulation of BIA biosynthesis. Such knowledge is vital for opium poppy genetic improvement and metabolic engineering efforts targeting the modulation of alkaloid yield and composition.

Sink strength, nutrient allocation, cannabinoid yield and associated transcript profiles vary in two drug-type Cannabis chemovars.

Cannabis sativa L. is one of the oldest domesticated crops. Hemp-type cultivars, which predominantly produce non-intoxicating cannabidiol (CBD), have been selected for their fast growth, seed, and fibre production, while drug-type chemovars were bred for high accumulation of tetrahydrocannabinol (THC). We investigated how the generation of CBD-dominant chemovars by introgression of hemp- into drug-type Cannabis impacted plant performance. The THC-dominant chemovar showed superior sink strength, higher flower biomass and demand-driven control of nutrient uptake. By contrast, the CBD-dominant chemovar hyperaccumulated phosphate in sink organs leading to reduced carbon and nitrogen assimilation in leaves, which limited flower biomass and cannabinoid yield. RNA-seq analyses determined organ- and chemovar-specific differences in expression of genes associated with nitrate and phosphate homeostasis as well as growth-regulating transcription factors that were correlated with measured traits. Among these were genes positively selected for during Cannabis domestication encoding an inhibitor of the phosphate starvation response SPX DOMAIN GENE3, nitrate reductase and two nitrate transporters. Altered nutrient sensing, acquisition or distribution are likely a consequence of adaption to growth on marginal, low-nutrient input lands in hemp. Our data provide evidence that such ancestral traits may become detrimental for female flower development and consequently overall CBD yield in protected cropping environments.

Management of esophageal anastomotic leaks, a systematic review and network meta-analysis.

There is currently no consensus as to how to manage esophageal anastomotic leaks. Intervention with endoscopic vacuum-assisted closure (EVAC), stenting, reoperation, and conservative management have all been mooted as potential options. To conduct a systematic review and network meta-analysis (NMA) to evaluate the optimal management strategy for esophageal anastomotic leaks. A systematic review was performed as per the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines with extension for NMA. NMA was performed using R packages and Shiny. In total, 12 retrospective studies were included, which included 511 patients. Of the 449 patients for whom data regarding sex was available, 371 (82.6%) were male, 78 (17.4%) were female. The average age of patients was 62.6 years (standard deviation 10.2). The stenting cohort included 245 (47.9%) patients. The EVAC cohort included 123 (24.1%) patients. The conservative cohort included 87 (17.0%) patients. The reoperation cohort included 56 (10.9%) patients. EVAC had a significantly decreased complication rate compared to stenting (odds ratio 0.23 95%, confidence interval [CI] 0.09;0.58). EVAC had a significantly lower mortality rate than stenting (odds ratio 0.43, 95% CI 0.21; 0.87). Reoperation was used in significantly larger leaks than stenting (mean difference 14.66, 95% CI 4.61;24.70). The growing use of EVAC as a first-line intervention in esophageal anastomotic leaks should continue given its proven effectiveness and significant reduction in both complication and mortality rates. Surgical management is often necessary for significantly larger leaks and will likely remain an effective option in uncontained leaks with systemic features.

Exercise breaks prevent attenuation in cerebrovascular function following an acute bout of uninterrupted sitting in healthy children.

The purpose of this study was to examine the effect of an acute bout of prolonged sitting with and without exercise breaks on cerebrovascular function in 7- to 13-year-old children. Forty-two children and adolescents were recruited to a crossover trial, with 15 girls (mean age 10.1 ± 2.5 years) and 16 boys (mean age 10.5 ± 1.3 years) completing the two trial conditions: SIT, uninterrupted sitting for 3 h and CYCLE, 3 h of sitting interrupted hourly with a 10-min moderate intensity exercise break. Cerebrovascular function was measured Pre and Post SIT and CYCLE from blood flow ( Q ̇ ${\dot{Q}}$ ), diameter, and shear rate of the internal carotid artery (ICA) at rest and in response to CO2 . Blood velocity in the middle (MCA) and posterior (PCA) cerebral arteries was assessed at rest, during a neurovascular coupling task (NVC) and in response to CO2 . We demonstrate that SIT but not CYCLE reduced ICA cerebrovascular reactivity to CO2 (%Δ ICA Q ̇ ${\dot{Q}}$ /Δ end-tidal CO2 : SIT: Pre 5.0 ± 2.4%/mmHg to Post 3.3 ± 2.8%/mmHg vs. CYCLE: Pre 4.4 ± 2.3%/mmHg to Post 5.3 ± 3.4%/mmHg, P = 0.05) and slowed the MCA blood velocity onset response time to hypercapnia (SIT: Pre 57.2 ± 32.6 s to Post 76.6 ± 55.2 s, vs. CYCLE: Pre 64.1 ± 40.4 s to Post 52.3 ± 28.8 s, P = 0.05). There were no changes in NVC. Importantly, breaking up prolonged sitting with hourly exercise breaks prevented the reductions in cerebrovascular reactivity to CO2 and the slowed intracranial blood velocity onset response time to hypercapnia apparent with uninterrupted sitting in children. NEW FINDINGS: What is the central question of this study? What are the effects of interrupting prolonged sitting on cerebrovascular function in children? What is the main finding and its importance? Prolonged sitting results in declines in cerebrovascular reactivity, a valuable index of cerebrovascular health. Breaking up prolonged sitting with hourly 10 min exercise breaks prevented these changes. These initial findings suggest excessive sedentary behaviour does impact cerebrovascular function in childhood, but taking exercise breaks prevents declines.

Protecting the stars of tomorrow: do international cardiovascular preparticipation screening policies account for the paediatric athlete? A systematic review and quality appraisal.

OBJECTIVE: (1) Identify and review current policies for the cardiovascular screening of athletes to assess their applicability to the paediatric population and (2) evaluate the quality of these policy documents using the Appraisal of Guidelines for Research & Evaluation II (AGREE II) tool. DESIGN: Systematic review and quality appraisal of policy documents. DATA SOURCES: A systematic search of PubMed, MEDLINE, Scopus, Web of Science, SportDiscus and CINAHL. ELIGIBILITY CRITERIA FOR SELECTING STUDIES: An article was included if it was a policy/position statement/guideline/consensus or recommendation paper relating to athletes and cardiovascular preparticipation screening. RESULTS AND SUMMARY: Of the 1630 articles screened, 13 met the inclusion criteria. Relevance to paediatric athletes was found to be high in 3 (23%), moderate in 6 (46%) and low in 4 (31%), and only 2 provide tailored guidance for the athlete aged 12-18 years. A median 5 related citations per policy investigated solely paediatric athletes, with study designs most commonly being retrospective (72%). AGREEII overall quality scores ranged from 25% to 92%, with a median of 75%. The lowest scoring domains were rigour of development; (median 32%) stakeholder involvement (median 47%) and Applicability (median 52%). CONCLUSION: Cardiac screening policies for athletes predominantly focus on adults, with few providing specific recommendations for paediatric athletes. The overall quality of the policies was moderate, with more recent documents scoring higher. Future research is needed in paediatric athletes to inform and develop cardiac screening guidelines, to improve the cardiac care of youth athletes.

London International Consensus and Delphi study on hamstring injuries part 3: rehabilitation, running and return to sport.

Hamstring injuries (HSIs) are the most common athletic injury in running and pivoting sports, but despite large amounts of research, injury rates have not declined in the last 2 decades. HSI often recur and many areas are lacking evidence and guidance for optimal rehabilitation. This study aimed to develop an international expert consensus for the management of HSI. A modified Delphi methodology and consensus process was used with an international expert panel, involving two rounds of online questionnaires and an intermediate round involving a consensus meeting. The initial information gathering round questionnaire was sent to 46 international experts, which comprised open-ended questions covering decision-making domains in HSI. Thematic analysis of responses outlined key domains, which were evaluated by a smaller international subgroup (n=15), comprising clinical academic sports medicine physicians, physiotherapists and orthopaedic surgeons in a consensus meeting. After group discussion around each domain, a series of consensus statements were prepared, debated and refined. A round 2 questionnaire was sent to 112 international hamstring experts to vote on these statements and determine level of agreement. Consensus threshold was set a priori at 70%. Expert response rates were 35/46 (76%) (first round), 15/35 (attendees/invitees to meeting day) and 99/112 (88.2%) for final survey round. Statements on rehabilitation reaching consensus centred around: exercise selection and dosage (78.8%-96.3% agreement), impact of the kinetic chain (95%), criteria to progress exercise (73%-92.7%), running and sprinting (83%-100%) in rehabilitation and criteria for return to sport (RTS) (78.3%-98.3%). Benchmarks for flexibility (40%) and strength (66.1%) and adjuncts to rehabilitation (68.9%) did not reach agreement. This consensus panel recommends individualised rehabilitation based on the athlete, sporting demands, involved muscle(s) and injury type and severity (89.8%). Early-stage rehab should avoid high strain loads and rates. Loading is important but with less consensus on optimum progression and dosage. This panel recommends rehabilitation progress based on capacity and symptoms, with pain thresholds dependent on activity, except pain-free criteria supported for sprinting (85.5%). Experts focus on the demands and capacity required for match play when deciding the rehabilitation end goal and timing of RTS (89.8%). The expert panellists in this study followed evidence on aspects of rehabilitation after HSI, suggesting rehabilitation prescription should be individualised, but clarified areas where evidence was lacking. Additional research is required to determine the optimal load dose, timing and criteria for HSI rehabilitation and the monitoring and testing metrics to determine safe rapid progression in rehabilitation and safe RTS. Further research would benefit optimising: prescription of running and sprinting, the application of adjuncts in rehabilitation and treatment of kinetic chain HSI factors.

London International Consensus and Delphi study on hamstring injuries part 1: classification.

Muscle injury classification systems for hamstring injuries have evolved to use anatomy and imaging information to aid management and prognosis. However, classification systems lack reliability and validity data and are not specific to individual hamstring muscles, potentially missing parameters vital for sport-specific and activity-specific decision making. A narrative evidence review was conducted followed by a modified Delphi study to build an international consensus on best-practice decision-making for the classification of hamstring injuries. This comprised a digital information gathering survey to a cohort of 46 international hamstring experts (sports medicine physicians, physiotherapists, surgeons, trainers and sports scientists) who were also invited to a face-to-face consensus group meeting in London . Fifteen of these expert clinicians attended to synthesise and refine statements around the management of hamstring injury. A second digital survey was sent to a wider group of 112 international experts. Acceptance was set at 70% agreement. Rounds 1 and 2 survey response rates were 35/46 (76%) and 99/112 (88.4%) of experts responding. Most commonly, experts used the British Athletics Muscle Injury Classification (BAMIC) (58%), Munich (12%) and Barcelona (6%) classification systems for hamstring injury. Issues identified to advance imaging classifications systems include: detailing individual hamstring muscles, establishing optimal use of imaging in diagnosis and classification, and testing the validity and reliability of classification systems. The most used hamstring injury classification system is the BAMIC. This consensus panel recommends hamstring injury classification systems evolve to integrate imaging and clinical parameters around: individual muscles, injury mechanism, sporting demand, functional criteria and patient-reported outcome measures. More research is needed on surgical referral and effectiveness criteria, and validity and reliability of classification systems to guide management.

Recognition of a novel variant of phosphoglycerate kinase 1 deficiency PGK1 Galveston (c.472G > C) in a child with hemolytic anemia, neurologic dysfunction and myopathy.

The enzyme phosphoglycerate kinase 1 (PGK1) catalyzes the first ATP producing reaction in the glycolysis pathway. Certain mutations to the coding gene of PGK1 present clinically with varying manifestations including hemolytic anemia, central nervous system (CNS) dysfunction and myopathy. Various PGK1 mutations have been described in the literature at the clinical and molecular level. Herein, we describe a novel case PGK1 mutation (PGK1 Galveston) in a 4-year-old boy who presented with all three manifestations. We discuss the characteristic hematopathology findings from this patient as well as provide a comparison with previously described neuroimaging findings. The variable clinical presentation of this condition along with its inherent uniqueness provide a diagnostic challenge for physicians. This presentation will add to the current body of knowledge for this condition and help guide future investigation and management.

RNA-seq analysis of laser microdissected Arabidopsis thaliana leaf epidermis, mesophyll and vasculature defines tissue-specific transcriptional responses to multiple stress treatments.

Acclimation of plants to adverse conditions requires the coordination of gene expression and signalling pathways between tissues and cell types. As the energy and carbon capturing organs, leaves are significantly affected by abiotic and biotic stresses. However, tissue- or cell type-specific analyses of stress responses have focussed on the Arabidopsis root. Here, we comparatively explore the transcriptomes of three leaf tissues (epidermis, mesophyll, vasculature) after induction of diverse stress pathways by chemical stimuli (antimycin A, 3-amino-1,2,4-triazole, methyl viologen, salicylic acid) and ultraviolet light in Arabidopsis using laser capture microdissection followed by RNA sequencing. Stimulation of stress pathways caused an overall reduction in the number of genes expressed in a tissue-specific manner, though a small subset gained or changed their tissue specificity. We find no evidence of a common stress response, with only a few genes consistently responsive to two or more treatments in the analysed tissues. However, differentially expressed genes overlap between tissues for individual treatments. A focussed analysis provided evidence for an interaction of auxin and ethylene that mediates retrograde signalling during mitochondrial dysfunction specifically in the epidermis, and a gene regulatory network defined the hierarchy of interactions. Taken together, we have generated an extensive reference dataset that will be valuable for future experiments analysing transcriptional responses on a tissue or single-cell level. Our results will enable the tailoring of the tissue-specific engineering of stress-tolerant plants.

A MYC2/MYC3/MYC4-dependent transcription factor network regulates water spray-responsive gene expression and jasmonate levels.

Mechanical stimuli, such as wind, rain, and touch affect plant development, growth, pest resistance, and ultimately reproductive success. Using water spray to simulate rain, we demonstrate that jasmonic acid (JA) signaling plays a key role in early gene-expression changes, well before it leads to developmental changes in flowering and plant architecture. The JA-activated transcription factors MYC2/MYC3/MYC4 modulate transiently induced expression of 266 genes, most of which peak within 30 min, and control 52% of genes induced >100-fold. Chromatin immunoprecipitation-sequencing analysis indicates that MYC2 dynamically binds >1,300 promoters and trans-activation assays show that MYC2 activates these promoters. By mining our multiomic datasets, we identified a core MYC2/MYC3/MYC4-dependent "regulon" of 82 genes containing many previously unknown MYC2 targets, including transcription factors bHLH19 and ERF109 bHLH19 can in turn directly activate the ORA47 promoter, indicating that MYC2/MYC3/MYC4 initiate a hierarchical network of downstream transcription factors. Finally, we also reveal that rapid water spray-induced accumulation of JA and JA-isoleucine is directly controlled by MYC2/MYC3/MYC4 through a positive amplification loop that regulates JA-biosynthesis genes.

Poor isometric neck extension strength as a risk factor for concussion in male professional Rugby Union players.

BACKGROUND: Concussion is one of the highest burden injuries within professional Rugby Union ('rugby') and comes with a high health and financial cost to players and teams. Limited evidence exists as to the existence of modifiable intrinsic risk factors for concussion, leaving athletes and clinicians with few options when developing prevention strategies. OBJECTIVE: To investigate whether neck strength is significantly associated with concussion incidence in professional male rugby players. METHODS: 225 rugby players were assessed for neck strength at three time points throughout the 2018/2019 season using a method of isometric contraction. Associations with clinically diagnosed concussion injuries are presented as incidence rate ratios (IRRs) with 95% CIs. RESULTS: Thirty concussions occurred in 29 players during the study period; a rate of 13.7 concussions per 1000 hours played. Greater neck strength was observed at mid and end of season time points versus preseason across the study population. There was a significant association between extension strength and concussion; a 10% increase for extension strength was associated with a 13% reduction in concussion rate (adjusted IRR (95% CI) 0.87 (0.78 to 0.98). No other significant associations were observed between concussion incidence and any other unique neck strength range or composite score. CONCLUSION: Higher neck extension strength is associated with lower concussion rates in male rugby players. Neck strength is a modifiable intrinsic risk factor for concussion and may be an important component of a strength and conditioning regime.

Association between thermal responses, medical events, performance, heat acclimation and health status in male and female elite athletes during the 2019 Doha World Athletics Championships.

PURPOSE: To determine associations between thermal responses, medical events, performance, heat acclimation and health status during a World Athletics Championships in hot-humid conditions. METHODS: From 305 marathon and race-walk starters, 83 completed a preparticipation questionnaire on health and acclimation. Core (Tcore; ingestible pill) and skin (Tskin; thermal camera) temperatures were measured in-competition in 56 and 107 athletes, respectively. 70 in-race medical events were analysed retrospectively. Performance (% personal best) and did not finish (DNF) were extracted from official results. RESULTS: Peak Tcore during competition reached 39.6°C±0.6°C (maximum 41.1°C). Tskin decreased from 32.2°C±1.3°C to 31.0°C±1.4°C during the races (p<0.001). Tcore was not related to DNF (25% of starters) or medical events (p≥0.150), whereas Tskin, Tskin rate of decrease and Tcore-to-Tskin gradient were (p≤0.029). A third of the athletes reported symptoms in the 10 days preceding the event, mainly insomnia, diarrhoea and stomach pain, with diarrhoea (9% of athletes) increasing the risk of in-race medical events (71% vs 17%, p<0.001). Athletes (63%) who performed 5-30 days heat acclimation before the competition: ranked better (18±13 vs 28±13, p=0.009), displayed a lower peak Tcore (39.4°C±0.4°C vs 39.8°C±0.7°C, p=0.044) and larger in-race decrease in Tskin (-1.4°C±1.0°C vs -0.9°C±1.2°C, p=0.060), than non-acclimated athletes. Although not significant, they also showed lower DNF (19% vs 30%, p=0.273) and medical events (19% vs 32%, p=0.179). CONCLUSION: Tskin, Tskin rate of decrease and Tcore-to-Tskin gradient were important indicators of heat tolerance. While heat-acclimated athletes ranked better, recent diarrhoea represented a significant risk factor for DNF and in-race medical events.

Temporal tissue-specific regulation of transcriptomes during barley (Hordeum vulgare) seed germination.

The distinct functions of individual cell types require cells to express specific sets of genes. The germinating seed is an excellent model to study genome regulation between cell types since the majority of the transcriptome is differentially expressed in a short period, beginning from a uniform, metabolically inactive state. In this study, we applied laser-capture microdissection RNA-sequencing to small numbers of cells from the plumule, radicle tip and scutellum of germinating barley seeds every 8 h, over a 48 h time course. Tissue-specific gene expression was notably common; 25% (910) of differentially expressed transcripts in plumule, 34% (1876) in radicle tip and 41% (2562) in scutellum were exclusive to that organ. We also determined that tissue-specific storage of transcripts occurs during seed development and maturation. Co-expression of genes had strong spatiotemporal structure, with most co-expression occurring within one organ and at a subset of specific time points during germination. Overlapping and distinct enrichment of functional categories were observed in the tissue-specific profiles. We identified candidate transcription factors amongst these that may be regulators of spatiotemporal gene expression programs. Our findings contribute to the broader goal of generating an integrative model that describes the structure and function of individual cells within seeds during germination.

The JA-pathway MYC transcription factors regulate photomorphogenic responses by targeting HY5 gene expression.

Jasmonates are key regulators of the balance between defence and growth in plants. However, the molecular mechanisms by which activation of defence reduces growth are not yet fully understood. Here, we analyze the role of MYC transcription factors (TFs) and jasmonic acid (JA) in photomorphogenic growth. We found that multiple myc mutants share light-associated phenotypes with mutants of the phytochrome B photoreceptor, such as delayed seed germination in the dark and long hypocotyl growth. Overexpression of MYC2 in a phyB background partially suppressed its long hypocotyl phenotype. Transcriptomic analysis of multiple myc mutants confirmed that MYCs are required for full expression of red (R) light-regulated genes, including the master regulator HY5. ChIP-seq analyses revealed that MYC2 and MYC3 bind directly to the promoter of HY5 and that HY5 gene expression and protein levels are compromised in multiple myc mutants. Altogether, our results pinpoint MYCs as photomorphogenic TFs that control phytochrome responses by activating HY5 expression. This has important implications in understanding the trade-off between growth and defence as the same TFs that activate defence responses are photomorphogenic growth regulators.