CO2 Response Screen in Grass Brachypodium Reveals Key Role of a MAP Kinase in CO2-Triggered Stomatal Closure.

Plants respond to increased CO2 concentrations through stomatal closure, which can contribute to increased water use efficiency. Grasses display faster stomatal responses than eudicots due to dumbbell-shaped guard cells flanked by subsidiary cells working in opposition. However, forward genetic screening for stomatal CO2 signal transduction mutants in grasses has yet to be reported. The grass model Brachypodium distachyon is closely related to agronomically important cereal crops, sharing largely collinear genomes. To gain insights into CO2 control mechanisms of stomatal movements in grasses, we developed an unbiased forward genetic screen with an EMS-mutagenized Brachypodium distachyon M5 generation population using infrared imaging to identify plants with altered leaf temperatures at elevated CO2. Among isolated mutants, a "chill1" mutant exhibited cooler leaf temperatures than wildtype Bd21-3 parent control plants after exposure to increased [CO2]. chill1 plants showed strongly impaired high CO2-induced stomatal closure despite retaining a robust abscisic acid-induced stomatal closing response. Through bulked segregant whole-genome-sequencing analyses followed by analyses of further backcrossed F4 generation plants and generation and characterization of sodium-azide and CRISPR-cas9 mutants, chill1 was mapped to a protein kinase, Mitogen-Activated Protein Kinase 5 (BdMPK5). The chill1 mutation impaired BdMPK5 protein-mediated CO2/HCO3- sensing together with the High Temperature 1 (HT1) Raf-like kinase in vitro. Furthermore, AlphaFold2-directed structural modeling predicted that the identified BdMPK5-D90N chill1 mutant residue is located at the interface of BdMPK5 with the BdHT1 Raf-like kinase. BdMPK5 is a key signaling component that mediates CO2-induced stomatal movements and is proposed to function as a component of the primary CO2 sensor in grasses.

The H3K4 demethylase JMJ1 is required for proper timing of flowering in Brachypodium distachyon.

Flowering is a key developmental transition in the plant life cycle. In temperate climates, flowering often occurs in response to the perception of seasonal cues such as changes in day-length and temperature. However, the mechanisms that have evolved to control the timing of flowering in temperate grasses are not fully understood. We identified a Brachypodium distachyon mutant whose flowering is delayed under inductive long-day conditions due to a mutation in the JMJ1 gene, which encodes a Jumonji domain-containing protein. JMJ1 is a histone demethylase that mainly demethylates H3K4me2 and H3K4me3 in vitro and in vivo. Analysis of the genome-wide distribution of H3K4me1, H3K4me2, and H3K4me3 in wild-type plants by chromatin immunoprecipitation and sequencing (ChIP-seq) combined with RNA sequencing (RNA-seq) revealed that H3K4m1 and H3K4me3 are positively associated with gene transcript levels, whereas H3K4me2 is negatively correlated with transcript levels. Furthermore, JMJ1 directly binds to the chromatin of the flowering regulator genes VRN1 and ID1 and affects their transcription by modifying their H3K4me2 and H3K4me3 levels. Genetic analyses indicated that JMJ1 promotes flowering by activating VRN1 expression. Our study reveals a role for JMJ1-mediated chromatin modification in the proper timing of flowering in B. distachyon.

Polygenic architecture of flowering time and its relationship with local environments in the grass Brachypodium distachyon.

Synchronizing the timing of reproduction with the environment is crucial in the wild. Among the multiple mechanisms, annual plants evolved to sense their environment, the requirement of cold-mediated vernalization is a major process that prevents individuals from flowering during winter. In many annual plants including crops, both a long and short vernalization requirement can be observed within species, resulting in so-called early-(spring) and late-(winter) flowering genotypes. Here, using the grass model Brachypodium distachyon, we explored the link between flowering-time-related traits (vernalization requirement and flowering time), environmental variation, and diversity at flowering-time genes by combining measurements under greenhouse and outdoor conditions. These experiments confirmed that B. distachyon natural accessions display large differences regarding vernalization requirements and ultimately flowering time. We underline significant, albeit quantitative effects of current environmental conditions on flowering-time-related traits. While disentangling the confounding effects of population structure on flowering-time-related traits remains challenging, population genomics analyses indicate that well-characterized flowering-time genes may contribute significantly to flowering-time variation and display signs of polygenic selection. Flowering-time genes, however, do not colocalize with genome-wide association peaks obtained with outdoor measurements, suggesting that additional genetic factors contribute to flowering-time variation in the wild. Altogether, our study fosters our understanding of the polygenic architecture of flowering time in a natural grass system and opens new avenues of research to investigate the gene-by-environment interaction at play for this trait.

INDETERMINATE1-mediated expression of FT family genes is required for proper timing of flowering in Brachypodium distachyon.

The transition to flowering is a major developmental switch in plants. In many temperate grasses, perception of indicators of seasonal change, such as changing day-length and temperature, leads to expression of FLOWERING LOCUS T1 (FT1) and FT-Like (FTL) genes that are essential for promoting the transition to flowering. However, little is known about the upstream regulators of FT1 and FTL genes in temperate grasses. Here, we characterize the monocot-specific gene INDETERMINATE1 (BdID1) in Brachypodium distachyon and demonstrate that BdID1 is a regulator of FT family genes. Mutations in ID1 impact the ability of the short-day (SD) vernalization, cold vernalization, and long-day (LD) photoperiod pathways to induce certain FTL genes. BdID1 is required for upregulation of FTL9 (FT-LIKE9) expression by the SD vernalization pathway, and overexpression of FTL9 in an id1 background can partially restore the delayed flowering phenotype of id1. We show that BdID1 binds in vitro to the promoter region of FTL genes suggesting that ID1 directly activates FTL expression. Transcriptome analysis shows that BdID1 is required for FT1, FT2, FTL12, and FTL13 expression under inductive LD photoperiods, indicating that BdID1 is a regulator of the FT gene family. Moreover, overexpression of FT1 in the id1 background results in rapid flowering similar to overexpressing FT1 in the wild type, demonstrating that BdID1 is upstream of FT family genes. Interestingly, ID1 negatively regulates a previously uncharacterized FTL gene, FTL4, and we show that FTL4 is a repressor of flowering. Thus, BdID1 is critical for proper timing of flowering in temperate grasses.

Wheat bZIPC1 interacts with FT2 and contributes to the regulation of spikelet number per spike.

KEY MESSAGE: The wheat transcription factor bZIPC1 interacts with FT2 and affects spikelet and grain number per spike. We identified a natural allele with positive effects on these two economically important traits. Loss-of-function mutations and natural variation in the gene FLOWERING LOCUS T2 (FT2) in wheat have previously been shown to affect spikelet number per spike (SNS). However, while other FT-like wheat proteins interact with bZIP-containing transcription factors from the A-group, FT2 does not interact with any of them. In this study, we used a yeast-two-hybrid screen with FT2 as bait and identified a grass-specific bZIP-containing transcription factor from the C-group, designated here as bZIPC1. Within the C-group, we identified four clades including wheat proteins that show Y2H interactions with different sets of FT-like and CEN-like encoded proteins. bZIPC1 and FT2 expression partially overlap in the developing spike, including the inflorescence meristem. Combined loss-of-function mutations in bZIPC-A1 and bZIPC-B1 (bzipc1) in tetraploid wheat resulted in a drastic reduction in SNS with a limited effect on heading date. Analysis of natural variation in the bZIPC-B1 (TraesCS5B02G444100) region revealed three major haplotypes (H1-H3), with the H1 haplotype showing significantly higher SNS, grain number per spike and grain weight per spike than both the H2 and H3 haplotypes. The favorable effect of the H1 haplotype was also supported by its increased frequency from the ancestral cultivated tetraploids to the modern tetraploid and hexaploid wheat varieties. We developed markers for the two non-synonymous SNPs that differentiate the bZIPC-B1b allele in the H1 haplotype from the ancestral bZIPC-B1a allele present in all other haplotypes. These diagnostic markers are useful tools to accelerate the deployment of the favorable bZIPC-B1b allele in pasta and bread wheat breeding programs.

EARLY FLOWERING 3 interactions with PHYTOCHROME B and PHOTOPERIOD1 are critical for the photoperiodic regulation of wheat heading time.

The photoperiodic response is critical for plants to adjust their reproductive phase to the most favorable season. Wheat heads earlier under long days (LD) than under short days (SD) and this difference is mainly regulated by the PHOTOPERIOD1 (PPD1) gene. Tetraploid wheat plants carrying the Ppd-A1a allele with a large deletion in the promoter head earlier under SD than plants carrying the wildtype Ppd-A1b allele with an intact promoter. Phytochromes PHYB and PHYC are necessary for the light activation of PPD1, and mutations in either of these genes result in the downregulation of PPD1 and very late heading time. We show here that both effects are reverted when the phyB mutant is combined with loss-of-function mutations in EARLY FLOWERING 3 (ELF3), a component of the Evening Complex (EC) in the circadian clock. We also show that the wheat ELF3 protein interacts with PHYB and PHYC, is rapidly modified by light, and binds to the PPD1 promoter in planta (likely as part of the EC). Deletion of the ELF3 binding region in the Ppd-A1a promoter results in PPD1 upregulation at dawn, similar to PPD1 alleles with intact promoters in the elf3 mutant background. The upregulation of PPD1 is correlated with the upregulation of the florigen gene FLOWERING LOCUS T1 (FT1) and early heading time. Loss-of-function mutations in PPD1 result in the downregulation of FT1 and delayed heading, even when combined with the elf3 mutation. Taken together, these results indicate that ELF3 operates downstream of PHYB as a direct transcriptional repressor of PPD1, and that this repression is relaxed both by light and by the deletion of the ELF3 binding region in the Ppd-A1a promoter. In summary, the regulation of the light mediated activation of PPD1 by ELF3 is critical for the photoperiodic regulation of wheat heading time.

PHYTOCHROME C regulation of photoperiodic flowering via PHOTOPERIOD1 is mediated by EARLY FLOWERING 3 in Brachypodium distachyon.

Daylength sensing in many plants is critical for coordinating the timing of flowering with the appropriate season. Temperate climate-adapted grasses such as Brachypodium distachyon flower during the spring when days are becoming longer. The photoreceptor PHYTOCHROME C is essential for long-day (LD) flowering in B. distachyon. PHYC is required for the LD activation of a suite of genes in the photoperiod pathway including PHOTOPERIOD1 (PPD1) that, in turn, result in the activation of FLOWERING LOCUS T (FT1)/FLORIGEN, which causes flowering. Thus, B. distachyon phyC mutants are extremely delayed in flowering. Here we show that PHYC-mediated activation of PPD1 occurs via EARLY FLOWERING 3 (ELF3), a component of the evening complex in the circadian clock. The extreme delay of flowering of the phyC mutant disappears when combined with an elf3 loss-of-function mutation. Moreover, the dampened PPD1 expression in phyC mutant plants is elevated in phyC/elf3 mutant plants consistent with the rapid flowering of the double mutant. We show that loss of PPD1 function also results in reduced FT1 expression and extremely delayed flowering consistent with results from wheat and barley. Additionally, elf3 mutant plants have elevated expression levels of PPD1, and we show that overexpression of ELF3 results in delayed flowering associated with a reduction of PPD1 and FT1 expression, indicating that ELF3 represses PPD1 transcription consistent with previous studies showing that ELF3 binds to the PPD1 promoter. Indeed, PPD1 is the main target of ELF3-mediated flowering as elf3/ppd1 double mutant plants are delayed flowering. Our results indicate that ELF3 operates downstream from PHYC and acts as a repressor of PPD1 in the photoperiod flowering pathway of B. distachyon.

MiR172-APETALA2-like genes integrate vernalization and plant age to control flowering time in wheat.

Plants possess regulatory mechanisms that allow them to flower under conditions that maximize reproductive success. Selection of natural variants affecting those mechanisms has been critical in agriculture to modulate the flowering response of crops to specific environments and to increase yield. In the temperate cereals, wheat and barley, the photoperiod and vernalization pathways explain most of the natural variation in flowering time. However, other pathways also participate in fine-tuning the flowering response. In this work, we integrate the conserved microRNA miR172 and its targets APETALA2-like (AP2L) genes into the temperate grass flowering network involving VERNALIZATION 1 (VRN1), VRN2 and FLOWERING LOCUS T 1 (FT1 = VRN3) genes. Using mutants, transgenics and different growing conditions, we show that miR172 promotes flowering in wheat, while its target genes AP2L1 (TaTOE1) and AP2L5 (Q) act as flowering repressors. Moreover, we reveal that the miR172-AP2L pathway regulates FT1 expression in the leaves, and that this regulation is independent of VRN2 and VRN1. In addition, we show that the miR172-AP2L module and flowering are both controlled by plant age through miR156 in spring cultivars. However, in winter cultivars, flowering and the regulation of AP2L1 expression are decoupled from miR156 downregulation with age, and induction of VRN1 by vernalization is required to repress AP2L1 in the leaves and promote flowering. Interestingly, the levels of miR172 and both AP2L genes modulate the flowering response to different vernalization treatments in winter cultivars. In summary, our results show that conserved and grass specific gene networks interact to modulate the flowering response, and that natural or induced mutations in AP2L genes are useful tools for fine-tuning wheat flowering time in a changing environment.

The wild grass Brachypodium distachyon as a developmental model system.

The arrival of cheap and high-throughput sequencing paired with efficient gene editing technologies allows us to use non-traditional model systems and mechanistically approach biological phenomena beyond what was conceivable just a decade ago. Venturing into different model systems enables us to explore for example clade-specific environmental responses to changing climates or the genetics and development of clade-specific organs, tissues and cell types. We-both early career researchers working with the wild grass model Brachypodium distachyon-want to use this review to (1) highlight why we think B. distachyon is a fantastic grass developmental model system, (2) summarize the tools and resources that have enabled discoveries made in B. distachyon, and (3) discuss a handful of developmental biology vignettes made possible by using B. distachyon as a model system. Finally, we want to conclude by (4) relating our personal stories with this emerging model system and (5) share what we think is important to consider before starting work with an emerging model system.

WAPO-A1 is the causal gene of the 7AL QTL for spikelet number per spike in wheat.

Improving our understanding of the genes regulating grain yield can contribute to the development of more productive wheat varieties. Previously, a highly significant QTL affecting spikelet number per spike (SNS), grain number per spike (GNS) and grain yield was detected on chromosome arm 7AL in multiple genome-wide association studies. Using a high-resolution genetic map, we established that the A-genome homeolog of WHEAT ORTHOLOG OF APO1 (WAPO-A1) was a leading candidate gene for this QTL. Using mutants and transgenic plants, we demonstrate in this study that WAPO-A1 is the causal gene underpinning this QTL. Loss-of-function mutants wapo-A1 and wapo-B1 showed reduced SNS in tetraploid wheat, and the effect was exacerbated in wapo1 combining both mutations. By contrast, spikes of transgenic wheat plants carrying extra copies of WAPO-A1 driven by its native promoter had higher SNS, a more compact spike apical region and a smaller terminal spikelet than the wild type. Taken together, these results indicate that WAPO1 affects SNS by regulating the timing of terminal spikelet formation. Both transgenic and wapo1 mutant plants showed a wide range of floral abnormalities, indicating additional roles of WAPO1 on wheat floral development. Previously, we found three widespread haplotypes in the QTL region (H1, H2 and H3), each associated with particular WAPO-A1 alleles. Results from this and our previous study show that the WAPO-A1 allele in the H1 haplotype (115-bp deletion in the promoter) is expressed at significantly lower levels in the developing spikes than the alleles in the H2 and H3 haplotypes, resulting in reduced SNS. Field experiments also showed that the H2 haplotype is associated with the strongest effects in increasing SNS and GNS (H2>H3>H1). The H2 haplotype is already present in most modern common wheat varieties but is rare in durum wheat, where it might be particularly useful to improve grain yield.

EARLY FLOWERING 3 and Photoperiod Sensing in Brachypodium distachyon.

The proper timing of flowering, which is key to maximize reproductive success and yield, relies in many plant species on the coordination between environmental cues and endogenous developmental programs. The perception of changes in day length is one of the most reliable cues of seasonal change, and this involves the interplay between the sensing of light signals and the circadian clock. Here, we describe a Brachypodium distachyon mutant allele of the evening complex protein EARLY FLOWERING 3 (ELF3). We show that the elf3 mutant flowers more rapidly than wild type plants in short days as well as under longer photoperiods but, in very long (20 h) days, flowering is equally rapid in elf3 and wild type. Furthermore, flowering in the elf3 mutant is still sensitive to vernalization, but not to ambient temperature changes. Molecular analyses revealed that the expression of a short-day marker gene is suppressed in elf3 grown in short days, and the expression patterns of clock genes and flowering time regulators are altered. We also explored the mechanisms of photoperiodic perception in temperate grasses by exposing B. distachyon plants grown under a 12 h photoperiod to a daily night break consisting of a mixture of red and far-red light. We showed that 2 h breaks are sufficient to accelerate flowering in B. distachyon under non-inductive photoperiods and that this acceleration of flowering is mediated by red light. Finally, we discuss advances and perspectives for research on the perception of photoperiod in temperate grasses.

Epistatic interactions between PHOTOPERIOD1, CONSTANS1 and CONSTANS2 modulate the photoperiodic response in wheat.

In Arabidopsis, CONSTANS (CO) integrates light and circadian clock signals to promote flowering under long days (LD). In the grasses, a duplication generated two paralogs designated as CONSTANS1 (CO1) and CONSTANS2 (CO2). Here we show that in tetraploid wheat plants grown under LD, combined loss-of-function mutations in the A and B-genome homeologs of CO1 and CO2 (co1 co2) result in a small (3 d) but significant (P<0.0001) acceleration of heading time both in PHOTOPERIOD1 (PPD1) sensitive (Ppd-A1b, functional ancestral allele) and insensitive (Ppd-A1a, functional dominant allele) backgrounds. Under short days (SD), co1 co2 mutants headed 13 d earlier than the wild type (P<0.0001) in the presence of Ppd-A1a. However, in the presence of Ppd-A1b, spikes from both genotypes failed to emerge by 180 d. These results indicate that CO1 and CO2 operate mainly as weak heading time repressors in both LD and SD. By contrast, in ppd1 mutants with loss-of-function mutations in both PPD1 homeologs, the wild type Co1 allele accelerated heading time >60 d relative to the co1 mutant allele under LD. We detected significant genetic interactions among CO1, CO2 and PPD1 genes on heading time, which were reflected in complex interactions at the transcriptional and protein levels. Loss-of-function mutations in PPD1 delayed heading more than combined co1 co2 mutations and, more importantly, PPD1 was able to perceive and respond to differences in photoperiod in the absence of functional CO1 and CO2 genes. Similarly, CO1 was able to accelerate heading time in response to LD in the absence of a functional PPD1. Taken together, these results indicate that PPD1 and CO1 are able to respond to photoperiod in the absence of each other, and that interactions between these two photoperiod pathways at the transcriptional and protein levels are important to fine-tune the flowering response in wheat.

Mutations in the predicted DNA polymerase subunit POLD3 result in more rapid flowering of Brachypodium distachyon.

The timing of reproduction is a critical developmental decision in the life cycle of many plant species. Fine mapping of a rapid-flowering mutant was done using whole-genome sequence data from bulked DNA from a segregating F2 mapping populations. The causative mutation maps to a gene orthologous with the third subunit of DNA polymerase δ (POLD3), a previously uncharacterized gene in plants. Expression analyses of POLD3 were conducted via real time qPCR to determine when and in what tissues the gene is expressed. To better understand the molecular basis of the rapid-flowering phenotype, transcriptomic analyses were conducted in the mutant vs wild-type. Consistent with the rapid-flowering mutant phenotype, a range of genes involved in floral induction and flower development are upregulated in the mutant. Our results provide the first characterization of the developmental and gene expression phenotypes that result from a lesion in POLD3 in plants.

An ortholog of CURLY LEAF/ENHANCER OF ZESTE like-1 is required for proper flowering in Brachypodium distachyon.

Many plants require prolonged exposure to cold to acquire the competence to flower. The process by which cold exposure results in competence is known as vernalization. In Arabidopsis thaliana, vernalization leads to the stable repression of the floral repressor FLOWERING LOCUS C via chromatin modification, including an increase of trimethylation on lysine 27 of histone H3 (H3K27me3) by Polycomb Repressive Complex 2 (PRC2). Vernalization in pooids is associated with the stable induction of a floral promoter, VERNALIZATION 1 (VRN1). From a screen for mutants with a reduced vernalization requirement in the model grass Brachypodium distachyon, we identified two recessive alleles of ENHANCER OF ZESTE-LIKE 1 (EZL1). EZL1 is orthologous to A. thaliana CURLY LEAF 1, a gene that encodes the catalytic subunit of PRC2. B. distachyon ezl1 mutants flower rapidly without vernalization in long-day (LD) photoperiods; thus, EZL1 is required for the proper maintenance of the vegetative state prior to vernalization. Transcriptomic studies in ezl1 revealed mis-regulation of thousands of genes, including ectopic expression of several floral homeotic genes in leaves. Loss of EZL1 results in the global reduction of H3K27me3 and H3K27me2, consistent with this gene making a major contribution to PRC2 activity in B. distachyon. Furthermore, in ezl1 mutants, the flowering genes VRN1 and AGAMOUS (AG) are ectopically expressed and have reduced H3K27me3. Artificial microRNA knock-down of either VRN1 or AG in ezl1-1 mutants partially restores wild-type flowering behavior in non-vernalized plants, suggesting that ectopic expression in ezl1 mutants may contribute to the rapid-flowering phenotype.

Extensive gene content variation in the Brachypodium distachyon pan-genome correlates with population structure.

While prokaryotic pan-genomes have been shown to contain many more genes than any individual organism, the prevalence and functional significance of differentially present genes in eukaryotes remains poorly understood. Whole-genome de novo assembly and annotation of 54 lines of the grass Brachypodium distachyon yield a pan-genome containing nearly twice the number of genes found in any individual genome. Genes present in all lines are enriched for essential biological functions, while genes present in only some lines are enriched for conditionally beneficial functions (e.g., defense and development), display faster evolutionary rates, lie closer to transposable elements and are less likely to be syntenic with orthologous genes in other grasses. Our data suggest that differentially present genes contribute substantially to phenotypic variation within a eukaryote species, these genes have a major influence in population genetics, and transposable elements play a key role in pan-genome evolution.

Establishment of a vernalization requirement in Brachypodium distachyon requires REPRESSOR OF VERNALIZATION1.

A requirement for vernalization, the process by which prolonged cold exposure provides competence to flower, is an important adaptation to temperate climates that ensures flowering does not occur before the onset of winter. In temperate grasses, vernalization results in the up-regulation of VERNALIZATION1 (VRN1) to establish competence to flower; however, little is known about the mechanism underlying repression of VRN1 in the fall season, which is necessary to establish a vernalization requirement. Here, we report that a plant-specific gene containing a bromo-adjacent homology and transcriptional elongation factor S-II domain, which we named REPRESSOR OF VERNALIZATION1 (RVR1), represses VRN1 before vernalization in Brachypodium distachyon That RVR1 is upstream of VRN1 is supported by the observations that VRN1 is precociously elevated in an rvr1 mutant, resulting in rapid flowering without cold exposure, and the rapid-flowering rvr1 phenotype is dependent on VRN1 The precocious VRN1 expression in rvr1 is associated with reduced levels of the repressive chromatin modification H3K27me3 at VRN1, which is similar to the reduced VRN1 H3K27me3 in vernalized plants. Furthermore, the transcriptome of vernalized wild-type plants overlaps with that of nonvernalized rvr1 plants, indicating loss of rvr1 is similar to the vernalized state at a molecular level. However, loss of rvr1 results in more differentially expressed genes than does vernalization, indicating that RVR1 may be involved in processes other than vernalization despite a lack of any obvious pleiotropy in the rvr1 mutant. This study provides an example of a role for this class of plant-specific genes.

Genetic Architecture of Flowering-Time Variation in Brachypodium distachyon.

The transition to reproductive development is a crucial step in the plant life cycle, and the timing of this transition is an important factor in crop yields. Here, we report new insights into the genetic control of natural variation in flowering time in Brachypodium distachyon, a nondomesticated pooid grass closely related to cereals such as wheat (Triticum spp.) and barley (Hordeum vulgare L.). A recombinant inbred line population derived from a cross between the rapid-flowering accession Bd21 and the delayed-flowering accession Bd1-1 were grown in a variety of environmental conditions to enable exploration of the genetic architecture of flowering time. A genotyping-by-sequencing approach was used to develop SNP markers for genetic map construction, and quantitative trait loci (QTLs) that control differences in flowering time were identified. Many of the flowering-time QTLs are detected across a range of photoperiod and vernalization conditions, suggesting that the genetic control of flowering within this population is robust. The two major QTLs identified in undomesticated B. distachyon colocalize with VERNALIZATION1/PHYTOCHROME C and VERNALIZATION2, loci identified as flowering regulators in the domesticated crops wheat and barley. This suggests that variation in flowering time is controlled in part by a set of genes broadly conserved within pooid grasses.

Anterior Cruciate Ligament Injuries in National Football League Athletes From 2010 to 2013: A Descriptive Epidemiology Study.

BACKGROUND: There is a high incidence of anterior cruciate ligament (ACL) injuries among National Football League (NFL) athletes; however, the incidence of reinjury in this population is unknown. PURPOSE: This retrospective epidemiological study analyzed all publicly disclosed ACL tears occurring in NFL players between 2010 and 2013 to characterize injury trends and determine the incidence of reinjury. STUDY DESIGN: Descriptive epidemiological study. METHODS: A comprehensive online search identified any NFL player who had suffered an ACL injury from 2010 to 2013. Position, playing surface, activity, and date were recorded. Each player was researched for any history of previous ACL injury. The NFL games database from USA Today was used to determine the incidence of ACL injuries on artificial turf and grass fields. Databases from Pro Football Focus and Pro Football Reference were used to determine the injury rate for each position. RESULTS: NFL players suffered 219 ACL injuries between 2010 and 2013. Forty players (18.3%) had a history of previous ACL injury, with 27 (12.3%) retears and 16 (7.3%) tears contralateral to a previous ACL injury. Five players (2.28%) suffered their third ACL tear. Receivers (wide receivers and tight ends) and backs (linebackers, fullbacks, and halfbacks) had significantly greater injury risk than the rest of the NFL players, while perimeter linemen (defensive ends and offensive tackles) had significantly lower injury risk than the rest of the players. Interior linemen (offensive guards, centers, and defensive tackles) had significantly greater injury risk compared with perimeter linemen. ACL injury rates per team games played were 0.050 for grass and 0.053 for turf fields (P > .05). CONCLUSION: In this retrospective epidemiological study of ACL tears in NFL players, retears and ACL tears contralateral to a previously torn ACL constituted a substantial portion (18.3%) of total ACL injuries. The significant majority of ACL injuries in players with a history of previous ACL injury were retears. Skilled offensive players and linebackers had the greatest injury risk, and significantly more ACL tears occurred among interior linemen than perimeter linemen. The month of August had the highest incidence of ACL injuries, probably because of expanded roster sizes at that point in the NFL season.

Evolution of VRN2/Ghd7-Like Genes in Vernalization-Mediated Repression of Grass Flowering.

Flowering of many plant species is coordinated with seasonal environmental cues such as temperature and photoperiod. Vernalization provides competence to flower after prolonged cold exposure, and a vernalization requirement prevents flowering from occurring prior to winter. In winter wheat (Triticum aestivum) and barley (Hordeum vulgare), three genes VRN1, VRN2, and FT form a regulatory loop that regulates the initiation of flowering. Prior to cold exposure, VRN2 represses FT. During cold, VRN1 expression increases, resulting in the repression of VRN2, which in turn allows activation of FT during long days to induce flowering. Here, we test whether the circuitry of this regulatory loop is conserved across Pooideae, consistent with their niche transition from the tropics to the temperate zone. Our phylogenetic analyses of VRN2-like genes reveal a duplication event occurred before the diversification of the grasses that gave rise to a CO9 and VRN2/Ghd7 clade and support orthology between wheat/barley VRN2 and rice (Oryza sativa) Ghd7 Our Brachypodium distachyon VRN1 and VRN2 knockdown and overexpression experiments demonstrate functional conservation of grass VRN1 and VRN2 in the promotion and repression of flowering, respectively. However, expression analyses in a range of pooids demonstrate that the cold repression of VRN2 is unique to core Pooideae such as wheat and barley. Furthermore, VRN1 knockdown in B. distachyon demonstrates that the VRN1-mediated suppression of VRN2 is not conserved. Thus, the VRN1-VRN2 feature of the regulatory loop appears to have evolved late in the diversification of temperate grasses.