Photoperiod-1 regulates the wheat inflorescence transcriptome to influence spikelet architecture and flowering time.

Photoperiod insensitivity has been selected by breeders to help adapt crops to diverse environments and farming practices. In wheat, insensitive alleles of Photoperiod-1 (Ppd-1) relieve the requirement of long daylengths to flower by promoting expression of floral promoting genes early in the season; however, these alleles also limit yield by reducing the number and fertility of grain-producing florets through processes that are poorly understood. Here, we performed transcriptome analysis of the developing inflorescence using near-isogenic lines that contain either photoperiod-insensitive or null alleles of Ppd-1, during stages when spikelet number is determined and floret development initiates. We report that Ppd-1 influences the stage-specific expression of genes with roles in auxin signaling, meristem identity, and protein turnover, and analysis of differentially expressed transcripts identified bZIP and ALOG transcription factors, namely PDB1 and ALOG1, which regulate flowering time and spikelet architecture. These findings enhance our understanding of genes that regulate inflorescence development and introduce new targets for improving yield potential.

Climate change enhances stability of wheat-flowering-date.

The stability of winter wheat-flowering-date is crucial for ensuring consistent and robust crop performance across diverse climatic conditions. However, the impact of climate change on wheat-flowering-dates remains uncertain. This study aims to elucidate the influence of climate change on wheat-flowering-dates, predict how projected future climate conditions will affect flowering date stability, and identify the most stable wheat genotypes in the study region. We applied a multi-locus genotype-based (MLG-based) model for simulating wheat-flowering-dates, which we calibrated and evaluated using observed data from the Northern China winter wheat region (NCWWR). This MLG-based model was employed to project flowering dates under different climate scenarios. The simulated flowering dates were then used to assess the stability of flowering dates under varying allelic combinations in projected climatic conditions. Our MLG-based model effectively simulated flowering dates, with a root mean square error (RMSE) of 2.3 days, explaining approximately 88.5 % of the genotypic variation in flowering dates among 100 wheat genotypes. We found that, in comparison to the baseline climate, wheat-flowering-dates are expected to shift earlier within the target sowing window by approximately 11 and 14 days by 2050 under the Representative Concentration Pathways 4.5 (RCP4.5) and RCP8.5 climate scenarios, respectively. Furthermore, our analysis revealed that wheat-flowering-date stability is likely to be further strengthened under projected climate scenarios due to early flowering trends. Ultimately, we demonstrate that the combination of Vrn and Ppd genes, rather than individual Vrn or Ppd genes, plays a critical role in wheat-flowering-date stability. Our results suggest that the combination of Ppd-D1a with winter genotypes carrying the vrn-D1 allele significantly contributes to flowering date stability under current and projected climate scenarios. These findings provide valuable insights for wheat breeders and producers under future climatic conditions.

Speed vernalization to accelerate generation advance in winter cereal crops.

There are many challenges facing the development of high-yielding, nutritious crops for future environments. One limiting factor is generation time, which prolongs research and plant breeding timelines. Recent advances in speed breeding protocols have dramatically reduced generation time for many short-day and long-day species by optimizing light and temperature conditions during plant growth. However, winter crops with a vernalization requirement still require up to 6-10 weeks in low-temperature conditions before the transition to reproductive development. Here, we tested a suite of environmental conditions and protocols to investigate whether the vernalization process can be accelerated. We identified a vernalization method consisting of exposing seeds at the soil surface to an extended photoperiod of 22 h day:2 h night at 10°C with transfer to speed breeding conditions that dramatically reduces generation time in both winter wheat (Triticum aestivum) and winter barley (Hordeum vulgare). Implementation of the speed vernalization protocol followed by speed breeding allowed the completion of up to five generations per year for winter wheat or barley, whereas only two generations can be typically completed under standard vernalization and plant growth conditions. The speed vernalization protocol developed in this study has great potential to accelerate biological research and breeding outcomes for winter crops.

MicroRNA-resistant alleles of HOMEOBOX DOMAIN-2 modify inflorescence branching and increase grain protein content of wheat.

Plant and inflorescence architecture determine the yield potential of crops. Breeders have harnessed natural diversity for inflorescence architecture to improve yields, and induced genetic variation could provide further gains. Wheat is a vital source of protein and calories; however, little is known about the genes that regulate the development of its inflorescence. Here, we report the identification of semidominant alleles for a class III homeodomain-leucine zipper transcription factor, HOMEOBOX DOMAIN-2 (HB-2), on wheat A and D subgenomes, which generate more flower-bearing spikelets and enhance grain protein content. These alleles increase HB-2 expression by disrupting a microRNA 165/166 complementary site with conserved roles in plants; higher HB-2 expression is associated with modified leaf and vascular development and increased amino acid supply to the inflorescence during grain development. These findings enhance our understanding of genes that control wheat inflorescence development and introduce an approach to improve the nutritional quality of grain.

The Roles of Temperature-Related Post-Transcriptional Regulation in Cereal Floral Development.

Temperature is a critical environmental signal in the regulation of plant growth and development. The temperature signal varies across a daily 24 h period, between seasons and stochastically depending on local environmental events. Extracting important information from these complex signals has led plants to evolve multiple temperature responsive regulatory mechanisms at the molecular level. In temperate cereals, we are starting to identify and understand these molecular mechanisms. In addition, we are developing an understanding of how this knowledge can be used to increase the robustness of crop yield in response to significant changes in local and global temperature patterns. To enable this, it is becoming apparent that gene regulation, regarding expression and post-transcriptional regulation, is crucial. Large transcriptomic studies are identifying global changes in spliced transcript variants and regulatory non-coding RNAs in response to seasonal and stress temperature signals in many of the cereal crops. Understanding the functions of these variants and targets of the non-coding RNAs will greatly increase how we enable the adaptation of crops. This review considers our current understanding and areas for future development.

Asymmetric expansions of FT and TFL1 lineages characterize differential evolution of the EuPEBP family in the major angiosperm lineages.

BACKGROUND: In flowering plants, precise timing of the floral transition is crucial to maximize chances of reproductive success, and as such, this process has been intensively studied. FLOWERING LOCUS T (FT) and TERMINAL FLOWER1 (TFL1) have been identified as closely related eukaryotic phosphatidylethanolamine-binding proteins ('EuPEBPs') that integrate multiple environmental stimuli, and act antagonistically to determine the optimal timing of the floral transition. Extensive research has demonstrated that FT acts similar to hormonal signals, being transported in the phloem from its primary site of expression in leaves to its primary site of action in the shoot meristem; TFL1 also appears to act as a mobile signal. Recent work implicates FT, TFL1, and the other members of the EuPEBP family, in the control of other important processes, suggesting that the EuPEBP family may be key general regulators of developmental transitions in flowering plants. In eudicots, there are a small number of EuPEBP proteins, but in monocots, and particularly grasses, there has been a large, but uncharacterized expansion of EuPEBP copy number, with unknown consequences for the EuPEBP function. RESULTS: To systematically characterize the evolution of EuPEBP proteins in flowering plants, and in land plants more generally, we performed a high-resolution phylogenetic analysis of 701 PEBP sequences from 208 species. We refine previous models of EuPEBP evolution in early land plants, demonstrating the algal origin of the family, and pin-pointing the origin of the FT/TFL1 clade at the base of monilophytes. We demonstrate how a core set of genes (MFT1, MFT2, FT, and TCB) at the base of flowering plants has undergone differential evolution in the major angiosperm lineages. This includes the radical expansion of the FT family in monocots into 5 core lineages, further re-duplicated in the grass family to 12 conserved clades. CONCLUSIONS: We show that many grass FT proteins are strongly divergent from other FTs and are likely neo-functional regulators of development. Our analysis shows that monocots and eudicots have strongly divergent patterns of EuPEBP evolution.

Multi-parent populations in crops: a toolbox integrating genomics and genetic mapping with breeding.

Crop populations derived from experimental crosses enable the genetic dissection of complex traits and support modern plant breeding. Among these, multi-parent populations now play a central role. By mixing and recombining the genomes of multiple founders, multi-parent populations combine many commonly sought beneficial properties of genetic mapping populations. For example, they have high power and resolution for mapping quantitative trait loci, high genetic diversity and minimal population structure. Many multi-parent populations have been constructed in crop species, and their inbred germplasm and associated phenotypic and genotypic data serve as enduring resources. Their utility has grown from being a tool for mapping quantitative trait loci to a means of providing germplasm for breeding programmes. Genomics approaches, including de novo genome assemblies and gene annotations for the population founders, have allowed the imputation of rich sequence information into the descendent population, expanding the breadth of research and breeding applications of multi-parent populations. Here, we report recent successes from crop multi-parent populations in crops. We also propose an ideal genotypic, phenotypic and germplasm 'package' that multi-parent populations should feature to optimise their use as powerful community resources for crop research, development and breeding.

TEOSINTE BRANCHED1 regulates height and stem internode length in bread wheat.

Regulation of plant height and stem elongation has contributed significantly to improvement of cereal productivity by reducing lodging and improving distribution of assimilates to the inflorescence and grain. In wheat, genetic control of height has been largely contributed by the Reduced height-1 alleles that confer gibberellin insensitivity; the beneficial effects of these alleles are associated with less favourable effects involving seedling emergence, grain quality, and inflorescence architecture that have driven new research investigating genetic variation of stem growth. Here, we show that TEOSINTE BRANCHED1 (TB1) regulates height of wheat, with TB1 being expressed at low levels in nodes of the main culm prior to elongation, and increased dosage of TB1 restricting elongation of stem internodes. The effect of TB1 on stem growth is not accompanied by poor seedling emergence, as transgenic lines with increased activity of TB1 form longer coleoptiles than null transgenic controls. Analysis of height in a multiparent mapping population also showed that allelic variation for TB1 on the B genome influences height, with plants containing the variant TB-B1b allele being taller than those with the wild-type TB-B1a allele. Our results show that TB1 restricts height and stem elongation in wheat, suggesting that variant alleles that alter the expression or function of TB1 could be used as a new source of genetic diversity for optimizing architecture of wheat in breeding programmes.

VERNALIZATION1 controls developmental responses of winter wheat under high ambient temperatures.

Low temperatures are required to regulate the transition from vegetative to reproductive growth via a pathway called vernalization. In wheat, vernalization predominantly involves the cold upregulation of the floral activator VERNALIZATION1 (VRN1). Here, we have used an extreme vernalization response, identified through studying ambient temperature responses, to reveal the complexity of temperature inputs into VRN-A1, with allelic inter-copy variation at a gene expansion of VRN-A1 modulating these effects. We find that the repressors of the reproductive transition, VERNALIZATION2 (VRN2) and ODDSOC2, are re-activated when plants experience high temperatures during and after vernalization. In addition, this re-activation is regulated by photoperiod for VRN2 but was independent of photoperiod for ODDSOC2 We also find this warm temperature interruption affects flowering time and floret number and is stage specific. This research highlights the important balance between floral activators and repressors in coordinating the response of a plant to temperature, and that the absence of warmth is essential for the completion of vernalization. This knowledge can be used to develop agricultural germplasm with more predictable vernalization responses that will be more resilient to variable growth temperatures.

TEOSINTE BRANCHED1 Regulates Inflorescence Architecture and Development in Bread Wheat (Triticum aestivum).

The flowers of major cereals are arranged on reproductive branches known as spikelets, which group together to form an inflorescence. Diversity for inflorescence architecture has been exploited during domestication to increase crop yields, and genetic variation for this trait has potential to further boost grain production. Multiple genes that regulate inflorescence architecture have been identified by studying alleles that modify gene activity or dosage; however, little is known in wheat. Here, we show TEOSINTE BRANCHED1 (TB1) regulates inflorescence architecture in bread wheat (Triticum aestivum) by investigating lines that display a form of inflorescence branching known as "paired spikelets." We show that TB1 interacts with FLOWERING LOCUS T1 and that increased dosage of TB1 alters inflorescence architecture and growth rate in a process that includes reduced expression of meristem identity genes, with allelic diversity for TB1 found to associate genetically with paired spikelet development in modern cultivars. We propose TB1 coordinates formation of axillary spikelets during the vegetative to floral transition and that alleles known to modify dosage or function of TB1 could help increase wheat yields.

Developmental responses of bread wheat to changes in ambient temperature following deletion of a locus that includes FLOWERING LOCUS T1.

FLOWERING LOCUS T (FT) is a central integrator of environmental signals that regulates the timing of vegetative to reproductive transition in flowering plants. In model plants, these environmental signals have been shown to include photoperiod, vernalization, and ambient temperature pathways, and in crop species, the integration of the ambient temperature pathway remains less well understood. In hexaploid wheat, at least 5 FT-like genes have been identified, each with a copy on the A, B, and D genomes. Here, we report the characterization of FT-B1 through analysis of FT-B1 null and overexpression genotypes under different ambient temperature conditions. This analysis has identified that the FT-B1 alleles perform differently under diverse environmental conditions; most notably, the FT-B1 null produces an increase in spikelet and tiller number when grown at lower temperature conditions. Additionally, absence of FT-B1 facilitates more rapid germination under both light and dark conditions. These results provide an opportunity to understand the FT-dependent pathways that underpin key responses of wheat development to changes in ambient temperature. This is particularly important for wheat, for which development and grain productivity are sensitive to changes in temperature.

Speed breeding is a powerful tool to accelerate crop research and breeding.

The growing human population and a changing environment have raised significant concern for global food security, with the current improvement rate of several important crops inadequate to meet future demand 1 . This slow improvement rate is attributed partly to the long generation times of crop plants. Here, we present a method called 'speed breeding', which greatly shortens generation time and accelerates breeding and research programmes. Speed breeding can be used to achieve up to 6 generations per year for spring wheat (Triticum aestivum), durum wheat (T. durum), barley (Hordeum vulgare), chickpea (Cicer arietinum) and pea (Pisum sativum), and 4 generations for canola (Brassica napus), instead of 2-3 under normal glasshouse conditions. We demonstrate that speed breeding in fully enclosed, controlled-environment growth chambers can accelerate plant development for research purposes, including phenotyping of adult plant traits, mutant studies and transformation. The use of supplemental lighting in a glasshouse environment allows rapid generation cycling through single seed descent (SSD) and potential for adaptation to larger-scale crop improvement programs. Cost saving through light-emitting diode (LED) supplemental lighting is also outlined. We envisage great potential for integrating speed breeding with other modern crop breeding technologies, including high-throughput genotyping, genome editing and genomic selection, accelerating the rate of crop improvement.

Effects of ambient temperature in association with photoperiod on phenology and on the expressions of major plant developmental genes in wheat (Triticum aestivum L.).

In addition to its role in vernalization, temperature is an important environmental stimulus in determining plant growth and development. We used factorial combinations of two photoperiods (16H, 12H) and three temperature levels (11, 18 and 25 °C) to study the temperature responses of 19 wheat cultivars with established genetic relationships. Temperature produced more significant effects on plant development than photoperiod, with strong genotypic components. Wheat genotypes with PPD-D1 photoperiod sensitive allele were sensitive to temperature; their development was delayed by higher temperature, which intensified under non-inductive conditions. The effect of temperature on plant development was not proportional; it influenced the stem elongation to the largest extent, and warmer temperature lengthened the lag phase between the detection of first node and the beginning of intensive stem elongation. The gene expression patterns of VRN1, VRN2 and PPD1 were also significantly modified by temperature, while VRN3 was more chronologically regulated. The associations between VRN1 and VRN3 gene expression with early apex development were significant in all treatments but were only significant for later plant developmental phases under optimal conditions (16H and 18 °C). Under 16H, the magnitude of the transient peak expression of VRN2 observed at 18 and 25 °C associated with the later developmental phases.

Light and circadian regulation of clock components aids flexible responses to environmental signals.

The circadian clock measures time across a 24 h period, increasing fitness by phasing biological processes to the most appropriate time of day. The interlocking feedback loop mechanism of the clock is conserved across species; however, the number of loops varies. Mathematical and computational analyses have suggested that loop complexity affects the overall flexibility of the oscillator, including its responses to entrainment signals. We used a discriminating experimental assay, at the transition between different photoperiods, in order to test this proposal in a minimal circadian network (in Ostreococcus tauri) and a more complex network (in Arabidopsis thaliana). Transcriptional and translational reporters in O. tauri primarily tracked dawn or dusk, whereas in A. thaliana, a wider range of responses were observed, consistent with its more flexible clock. Model analysis supported the requirement for this diversity of responses among the components of the more complex network. However, these and earlier data showed that the O. tauri network retains surprising flexibility, despite its simple circuit. We found that models constructed from experimental data can show flexibility either from multiple loops and/or from multiple light inputs. Our results suggest that O. tauri has adopted the latter strategy, possibly as a consequence of genomic reduction.

Proteasome function is required for biological timing throughout the twenty-four hour cycle.

Circadian clocks were, until recently, seen as a consequence of rhythmic transcription of clock components, directed by transcriptional/translational feedback loops (TTFLs). Oscillations of protein modification were then discovered in cyanobacteria. Canonical posttranslational signaling processes have known importance for clocks across taxa. More recently, evidence from the unicellular eukaryote Ostreococcus tauri revealed a transcription-independent, rhythmic protein modification shared in anucleate human cells. In this study, the Ostreococcus system reveals a central role for targeted protein degradation in the mechanism of circadian timing. The Ostreococcus clockwork contains a TTFL involving the morning-expressed CCA1 and evening-expressed TOC1 proteins. Cellular CCA1 and TOC1 protein content and degradation rates are analyzed qualitatively and quantitatively using luciferase reporter fusion proteins. CCA1 protein degradation rates, measured in high time resolution, feature a sharp clock-regulated peak under constant conditions. TOC1 degradation peaks in response to darkness. Targeted protein degradation, unlike transcription and translation, is shown to be essential to sustain TTFL rhythmicity throughout the circadian cycle. Although proteasomal degradation is not necessary for sustained posttranslational oscillations in transcriptionally inactive cells, TTFL and posttranslational oscillators are normally coupled, and proteasome function is crucial to sustain both.

Circadian rhythms persist without transcription in a eukaryote.

Circadian rhythms are ubiquitous in eukaryotes, and coordinate numerous aspects of behaviour, physiology and metabolism, from sleep/wake cycles in mammals to growth and photosynthesis in plants. This daily timekeeping is thought to be driven by transcriptional-translational feedback loops, whereby rhythmic expression of 'clock' gene products regulates the expression of associated genes in approximately 24-hour cycles. The specific transcriptional components differ between phylogenetic kingdoms. The unicellular pico-eukaryotic alga Ostreococcus tauri possesses a naturally minimized clock, which includes many features that are shared with plants, such as a central negative feedback loop that involves the morning-expressed CCA1 and evening-expressed TOC1 genes. Given that recent observations in animals and plants have revealed prominent post-translational contributions to timekeeping, a reappraisal of the transcriptional contribution to oscillator function is overdue. Here we show that non-transcriptional mechanisms are sufficient to sustain circadian timekeeping in the eukaryotic lineage, although they normally function in conjunction with transcriptional components. We identify oxidation of peroxiredoxin proteins as a transcription-independent rhythmic biomarker, which is also rhythmic in mammals. Moreover we show that pharmacological modulators of the mammalian clock mechanism have the same effects on rhythms in Ostreococcus. Post-translational mechanisms, and at least one rhythmic marker, seem to be better conserved than transcriptional clock regulators. It is plausible that the oldest oscillator components are non-transcriptional in nature, as in cyanobacteria, and are conserved across kingdoms.

Temporal repression of core circadian genes is mediated through EARLY FLOWERING 3 in Arabidopsis.

The circadian clock provides robust, ∼24 hr biological rhythms throughout the eukaryotes. The clock gene circuit in plants comprises interlocking transcriptional feedback loops, reviewed in [1], whereby the morning-expressed transcription factors CIRCADIAN CLOCK-ASSOCIATED 1 (CCA1) and LATE ELONGATED HYPOCOTYL (LHY) repress the expression of evening genes, notably TIMING OF CAB EXPRESSION 1 (TOC1). EARLY FLOWERING 3 (ELF3) has been implicated as a repressor of light signaling to the clock [2, 3] and, paradoxically, as an activator of the light-induced genes CCA1 and LHY [4, 5]. We use cca1-11 lhy-21 elf3-4 plants to separate the repressive function of ELF3 from its downstream targets CCA1 and LHY. We further demonstrate that ELF3 associates physically with the promoter of PSEUDO-RESPONSE REGULATOR 9 (PRR9), a repressor of CCA1 and LHY expression, in a time-dependent fashion. The repressive function of ELF3 is thus consistent with indirect activation of LHY and CCA1, in a double-negative connection via a direct ELF3 target, PRR9. This mechanism reconciles the functions of ELF3 in the clock network during the night and points to further effects of ELF3 during the day.

Multiple light inputs to a simple clock circuit allow complex biological rhythms.

Circadian clocks are biological timekeepers that allow living cells to time their activity in anticipation of predictable environmental changes. Detailed understanding of the circadian network of higher plants, such as Arabidopsis thaliana, is hampered by the high number of partially redundant genes. However, the picoeukaryotic alga Ostreococcus tauri, which was recently shown to possess a small number of non-redundant clock genes, presents an attractive alternative target for detailed modelling of circadian clocks in the green lineage. Based on extensive time-series data from in vivo reporter gene assays, we developed a model of the Ostreococcus clock as a feedback loop between the genes TOC1 and CCA1. The model reproduces the dynamics of the transcriptional and translational reporters over a range of photoperiods. Surprisingly, the model is also able to predict the transient behaviour of the clock when the light conditions are altered. Despite the apparent simplicity of the clock circuit, it displays considerable complexity in its response to changing light conditions. Systematic screening of the effects of altered day length revealed a complex relationship between phase and photoperiod, which is also captured by the model. The complex light response is shown to stem from circadian gating of light-dependent mechanisms. This study provides insights into the contributions of light inputs to the Ostreococcus clock. The model suggests that a high number of light-dependent reactions are important for flexible timing in a circadian clock with only one feedback loop.