Distinct functions of FASCILIN-LIKE ARABINOGALACTAN PROTEINS relate to domain structure.

The role of glycoproteins as key cell surface molecules during development and stress is well established; yet, the relationship between their structural features and functional mechanisms is poorly defined. FASCICLIN-LIKE ARABINOGALACTAN PROTEINs (FLAs), which impact plant growth and development, are an excellent example of a glycoprotein family with a complex multidomain structure. FLAs combine globular fasciclin-like (FAS1) domains with regions that are intrinsically disordered and contain glycomotifs for directing the addition of O-linked arabinogalactan (AG) glycans. Additional posttranslational modifications on FLAs include N-linked glycans in the FAS1 domains, a cleaved signal peptide at the N terminus, and often a glycosylphosphatidylinositol (GPI) anchor signal sequence at the C terminus. The roles of glycosylation, the GPI anchor, and FAS1 domain functions in the polysaccharide-rich extracellular matrix of plants remain unclear, as do the relationships between them. In this study, we examined sequence-structure-function relationships of Arabidopsis (Arabidopsis thaliana) FLA11, demonstrated to have roles in secondary cell wall (SCW) development, by introducing domain mutations and functional specialization through domain swaps with FLA3 and FLA12. We identified FAS1 domains as essential for FLA function, differentiating FLA11/FLA12, with roles in SCW development, from FLA3, specific to flowers and involved in pollen development. The GPI anchor and AG glycosylation co-regulate the cell surface location and release of FLAs into cell walls. The AG glycomotif sequence closest to the GPI anchor (AG2) is a major feature differentiating FLA11 from FLA12. The results of our study show that the multidomain structure of different FLAs influences their subcellular location and biological functions during plant development.

An autoactive NB-LRR gene causes Rht13 dwarfism in wheat.

Semidwarfing genes have greatly increased wheat yields globally, yet the widely used gibberellin (GA)-insensitive genes Rht-B1b and Rht-D1b have disadvantages for seedling emergence. Use of the GA-sensitive semidwarfing gene Rht13 avoids this pleiotropic effect. Here, we show that Rht13 encodes a nucleotide-binding site/leucine-rich repeat (NB-LRR) gene. A point mutation in the semidwarf Rht-B13b allele autoactivates the NB-LRR gene and causes a height reduction comparable with Rht-B1b and Rht-D1b in diverse genetic backgrounds. The autoactive Rht-B13b allele leads to transcriptional up-regulation of pathogenesis-related genes including class III peroxidases associated with cell wall remodeling. Rht13 represents a new class of reduced height (Rht) gene, unlike other Rht genes, which encode components of the GA signaling or metabolic pathways. This discovery opens avenues to use autoactive NB-LRR genes as semidwarfing genes in a range of crop species, and to apply Rht13 in wheat breeding programs using a perfect genetic marker.

Cotton Breeding in Australia: Meeting the Challenges of the 21st Century.

The Commonwealth Scientific and Industrial Research Organisation (CSIRO) cotton breeding program is the sole breeding effort for cotton in Australia, developing high performing cultivars for the local industry which is worth∼AU$3 billion per annum. The program is supported by Cotton Breeding Australia, a Joint Venture between CSIRO and the program's commercial partner, Cotton Seed Distributors Ltd. (CSD). While the Australian industry is the focus, CSIRO cultivars have global impact in North America, South America, and Europe. The program is unique compared with many other public and commercial breeding programs because it focuses on diverse and integrated research with commercial outcomes. It represents the full research pipeline, supporting extensive long-term fundamental molecular research; native and genetically modified (GM) trait development; germplasm enhancement focused on yield and fiber quality improvements; integration of third-party GM traits; all culminating in the release of new commercial cultivars. This review presents evidence of past breeding successes and outlines current breeding efforts, in the areas of yield and fiber quality improvement, as well as the development of germplasm that is resistant to pests, diseases and abiotic stressors. The success of the program is based on the development of superior germplasm largely through field phenotyping, together with strong commercial partnerships with CSD and Bayer CropScience. These relationships assist in having a shared focus and ensuring commercial impact is maintained, while also providing access to markets, traits, and technology. The historical successes, current foci and future requirements of the CSIRO cotton breeding program have been used to develop a framework designed to augment our breeding system for the future. This will focus on utilizing emerging technologies from the genome to phenome, as well as a panomics approach with data management and integration to develop, test and incorporate new technologies into a breeding program. In addition to streamlining the breeding pipeline for increased genetic gain, this technology will increase the speed of trait and marker identification for use in genome editing, genomic selection and molecular assisted breeding, ultimately producing novel germplasm that will meet the coming challenges of the 21st Century.

FLA11 and FLA12 glycoproteins fine-tune stem secondary wall properties in response to mechanical stresses.

Secondary cell walls (SCWs) in stem xylem vessel and fibre cells enable plants to withstand the enormous compressive forces associated with upright growth. It remains unclear if xylem vessel and fibre cells can directly sense mechanical stimuli and modify their SCW during development. We provide evidence that Arabidopsis SCW-specific Fasciclin-Like Arabinogalactan-proteins 11 (FLA11) and 12 (FLA12) are possible cell surface sensors regulating SCW development in response to mechanical stimuli. Plants overexpressing FLA11 (OE-FLA11) showed earlier SCW development compared to the wild-type (WT) and altered SCW properties that phenocopy WT plants under compression stress. By contrast, OE-FLA12 stems showed higher cellulose content compared to WT plants, similar to plants experiencing tensile stress. fla11, OE-FLA11, fla12, and OE-FLA12 plants showed altered SCW responses to mechanical stress compared to the WT. Quantitative polymerase chain reaction (qPCR) and RNA-seq analysis revealed the up-regulation of genes and pathways involved in stress responses and SCW synthesis and regulation. Analysis of OE-FLA11 nst1 nst3 plants suggests that FLA11 regulation of SCWs is reliant on classical transcriptional networks. Our data support the involvement of FLA11 and FLA12 in SCW sensing complexes to fine-tune both the initiation of SCW development and the balance of lignin and cellulose synthesis/deposition in SCWs during development and in response to mechanical stimuli.

Fasciclin-Like Arabinogalactan-Protein 16 (FLA16) Is Required for Stem Development in Arabidopsis.

The predominant Fascilin 1 (FAS1)-containing proteins in plants belong to the Fasciclin-Like Arabinogalactan-protein (FLA) family of extracellular glycoproteins. In addition to FAS1 domains, these multi-domain FLA proteins contain glycomotif regions predicted to direct addition of large arabinogalactan (AG) glycans and many contain signal sequences for addition of a glycosylphosphatidylinositol (GPI)-anchor to tether them to the plasma membrane. FLAs are proposed to play both structural and signaling functions by forming a range of interactions in the plant extracellular matrix, similar to FAS1-containing proteins in animals. FLA group B members contain two FAS1 domains and are not predicted to be GPI-anchored. None of the group B members have been functionally characterized or their sub-cellular location resolved, limiting understanding of their function. We investigated the group B FLA16 in Arabidopsis that is predominantly expressed in inflorescence tissues. FLA16 is the most highly expressed FLA in the stem after Group A members FLA11 and FLA12 that are stem specific. A FLA16-YFP fusion protein driven by the endogenous putative FLA16 promoter in wild type background showed expression in cells with secondary cell walls, and FLA16 displayed characteristics of cell wall glycoproteins with moderate glycosylation. Investigation of a fla16 mutant showed loss of FLA16 leads to reduced stem length and altered biomechanical properties, likely as a result of reduced levels of cellulose. Immuno-labeling indicated support for FLA16 location to the plasma-membrane and (apoplastic) cell wall of interfascicular stem fiber cells. Together these results indicate FLA16, a two-FAS1 domain FLAs, plays a role in plant secondary cell wall synthesis and function.

Correction to: Tissue and cell-specific transcriptomes in cotton reveal the subtleties of gene regulation underlying the diversity of plant secondary cell walls.

Upon publication of the original article [1], the authors had flagged that Fig. 1 had been published twice, as both Fig. 1 and Additional file 3.

Rht18 Semidwarfism in Wheat Is Due to Increased GA 2-oxidaseA9 Expression and Reduced GA Content.

Semidwarfing genes have improved crop yield by reducing height, improving lodging resistance, and allowing plants to allocate more assimilates to grain growth. In wheat (Triticum aestivum), the Rht18 semidwarfing gene was identified and deployed in durum wheat before it was transferred into bread wheat, where it was shown to have agronomic potential. Rht18, a dominant and gibberellin (GA) responsive mutant, is genetically and functionally distinct from the widely used GA-insensitive semidwarfing genes Rht-B1b and Rht-D1b In this study, the Rht18 gene was identified by mutagenizing the semidwarf durum cultivar Icaro (Rht18) and generating mutants with a range of tall phenotypes. Isolating and sequencing chromosome 6A of these "overgrowth" mutants showed that they contained independent mutations in the coding region of GA2oxA9GA2oxA9 is predicted to encode a GA 2-oxidase that metabolizes GA biosynthetic intermediates into inactive products, effectively reducing the amount of bioactive GA (GA1). Functional analysis of the GA2oxA9 protein demonstrated that GA2oxA9 converts the intermediate GA12 to the inactive metabolite GA110 Furthermore, Rht18 showed higher expression of GA2oxA9 and lower GA content compared with its tall parent. These data indicate that the increased expression of GA2oxA9 in Rht18 results in a reduction of both bioactive GA content and plant height. This study describes a height-reducing mechanism that can generate new genetic diversity for semidwarfism in wheat by combining increased expression with mutations of specific amino acid residues in GA2oxA9.

Genomewide analysis of the lateral organ boundaries domain gene family in Eucalyptus grandis reveals members that differentially impact secondary growth.

Lateral Organ Boundaries Domain (LBD) proteins are plant-specific transcription factors playing crucial roles in growth and development. However, the function of LBD proteins in Eucalyptus grandis remains largely unexplored. In this study, LBD genes in E. grandis were identified and characterized using bioinformatics approaches. Gene expression patterns in various tissues and the transcriptional responses of EgLBDs to exogenous hormones were determined by qRT-PCR. Functions of the selected EgLBDs were studied by ectopically overexpressing in a hybrid poplar (Populus alba × Populus glandulosa). Expression levels of genes in the transgenic plants were investigated by RNA-seq. Our results showed that there were forty-six EgLBD members in the E. grandis genome and three EgLBDs displayed xylem- (EgLBD29) or phloem-preferential expression (EgLBD22 and EgLBD37). Confocal microscopy indicated that EgLBD22, EgLBD29 and EgLBD37 were localized to the nucleus. Furthermore, we found that EgLBD22, EgLBD29 and EgLBD37 were responsive to the treatments of indol-3-acetic acid and gibberellic acid. More importantly, we demonstrated EgLBDs exerted different influences on secondary growth. Namely, 35S::EgLBD37 led to significantly increased secondary xylem, 35S::EgLBD29 led to greatly increased phloem fibre production, and 35S::EgLBD22 showed no obvious effects. We revealed that key genes related to gibberellin, ethylene and auxin signalling pathway as well as cell expansion were significantly up- or down-regulated in transgenic plants. Our new findings suggest that LBD genes in E. grandis play important roles in secondary growth. This provides new mechanisms to increase wood or fibre production.

Tissue and cell-specific transcriptomes in cotton reveal the subtleties of gene regulation underlying the diversity of plant secondary cell walls.

BACKGROUND: Knowledge of plant secondary cell wall (SCW) regulation and deposition is mainly based on the Arabidopsis model of a 'typical' lignocellulosic SCW. However, SCWs in other plants can vary from this. The SCW of mature cotton seed fibres is highly cellulosic and lacks lignification whereas xylem SCWs are lignocellulosic. We used cotton as a model to study different SCWs and the expression of the genes involved in their formation via RNA deep sequencing and chemical analysis of stem and seed fibre. RESULTS: Transcriptome comparisons from cotton xylem and pith as well as from a developmental series of seed fibres revealed tissue-specific and developmentally regulated expression of several NAC transcription factors some of which are likely to be important as top tier regulators of SCW formation in xylem and/or seed fibre. A so far undescribed hierarchy was identified between the top tier NAC transcription factors SND1-like and NST1/2 in cotton. Key SCW MYB transcription factors, homologs of Arabidopsis MYB46/83, were practically absent in cotton stem xylem. Lack of expression of other lignin-specific MYBs in seed fibre relative to xylem could account for the lack of lignin deposition in seed fibre. Expression of a MYB103 homolog correlated with temporal expression of SCW CesAs and cellulose synthesis in seed fibres. FLAs were highly expressed and may be important structural components of seed fibre SCWs. Finally, we made the unexpected observation that cell walls in the pith of cotton stems contained lignin and had a higher S:G ratio than in xylem, despite that tissue's lacking many of the gene transcripts normally associated with lignin biosynthesis. CONCLUSIONS: Our study in cotton confirmed some features of the currently accepted gene regulatory cascade for 'typical' plant SCWs, but also revealed substantial differences, especially with key downstream NACs and MYBs. The lignocellulosic SCW of cotton xylem appears to be achieved differently from that in Arabidopsis. Pith cell walls in cotton stems are compositionally very different from that reported for other plant species, including Arabidopsis. The current definition of a 'typical' primary or secondary cell wall might not be applicable to all cell types in all plant species.

Arabidopsis DEFECTIVE KERNEL1 regulates cell wall composition and axial growth in the inflorescence stem.

Axial growth in plant stems requires a fine balance between elongation and stem mechanical reinforcement to ensure mechanical stability. Strength is provided by the plant cell wall, the deposition of which must be coordinated with cell expansion and elongation to ensure that integrity is maintained during growth. Coordination of these processes is critical and yet poorly understood. The plant-specific calpain, DEFECTIVE KERNEL1 (DEK1), plays a key role in growth coordination in leaves, yet its role in regulating stem growth has not been addressed. Using plants overexpressing the active CALPAIN domain of DEK1 (CALPAIN OE) and a DEK1 knockdown line (amiRNA-DEK1), we undertook morphological, biochemical, biophysical, and microscopic analyses of mature inflorescence stems. We identify a novel role for DEK1 in the maintenance of cell wall integrity and coordination of growth during inflorescence stem development. CALPAIN OE plants are significantly reduced in stature and have short, thickened stems, while amiRNA-DEK1 lines have weakened stems that are unable to stand upright. Microscopic analyses of the stems identify changes in cell size, shape and number, and differences in both primary and secondary cell wall thickness and composition. Taken together, our results suggest that DEK1 influences primary wall growth by indirectly regulating cellulose and pectin deposition. In addition, we observe changes in secondary cell walls that may compensate for altered primary cell wall composition. We propose that DEK1 activity is required for the coordination of stem strengthening with elongation during axial growth.

The fasciclin-like arabinogalactan protein family of Eucalyptus grandis contains members that impact wood biology and biomechanics.

Fasciclin-like arabinogalactan protein (FLA) families have been identified and characterised in key plant species, with some members exhibiting functional specialization. Here we identify the FLA family of Eucalyptus grandis, and investigate the roles of three single-FAS domain FLAs, with particular focus on secondary cell-wall formation and wood properties. We use various in-silico approaches to identify and characterise E. grandis genome FLAs, and perform phylogenetic comparisons with other species. For three key FLAs, we perform functional testing including promoter-reporter and overexpression transgenic approaches using eucalypts, poplar and tobacco. Of the 18 eucalypt FLAs identified, several were specifically and highly expressed in stems. The specificity to stem xylem vessel and fibre development was demonstrated with EniFLA1promoter:GUS studies in several species. Testing of select eucalypt FLAs resulted in altered wood development and properties, for example 35S:EgrFLA2 led to a 3 degree reduction in cellulose microfibril angle in eucalypt xylem fibres, and 35S:EgrFLA3 to a reduction in tobacco stem flexural strength. These results indicate that the eucalypt FLA family contains diverse members, and particular members with single FAS domains that are functionally specialized for secondary cell wall growth and properties.

The Arabidopsis wood model-the case for the inflorescence stem.

Arabidopsis thaliana has successfully served as a model to discover genes and proteins that have roles in a wide range of plant traits, including wood-related traits, such as lignin, cellulose and hemicellulose biosynthesis, secondary growth regulation, and secondary cell wall synthesis. Both the radially thickened hypocotyl and the inflorescence stem (flower stalk) have been studied. In this review, we address lingering doubts regarding the utility of Arabidopsis as a model for wood development by highlighting studies that provide new biochemical and biophysical evidence that extend support for the Arabidopsis inflorescence stem as a model for wood development beyond what is currently thought. We describe different aspects of Arabidopsis that make it a highly versatile tool for the study of wood development. One would likely utilise the radially thickened hypocotyl because of its more fully developed vascular cambium for traits related specifically to secondary (i.e. cambial) growth. It is more productive to utilise the inflorescence stem for wood-like biophysical traits. Accession variation has been underexploited as a powerful method to discover genes governing wood-like traits. We discuss recent findings that survey the accession variation in Arabidopsis for biochemical and biophysical properties of various wood traits, such as microfibril angle, tensile strength and cellulose/hemicellulose content. Furthermore we discuss how larger-scale studies of this nature using plants grown in long days (as opposed to the current short-day paradigm) could accelerate gene discovery and our understanding of cell wall and wood development. We highlight some relatively unexplored areas of research relating to the secondary cell wall composition, architecture and biophysical properties of the inflorescence stem, and how these traits are relevant to wood formation. The Arabidopsis inflorescence stem has other characteristics, expressed genes and traits held in common with woody species that have not been widely characterised or discussed to date. We discuss how this conservation may indicate the more general potential for "true" woodiness in herbaceous species, in the context of so-called secondary woodiness.

A survey of the natural variation in biomechanical and cell wall properties in inflorescence stems reveals new insights into the utility of Arabidopsis as a wood model.

The natural trait variation in Arabidopsis thaliana (L.) Heynh. accessions is an important resource for understanding many biological processes but it is underexploited for wood-related properties. Twelve A. thaliana accessions from diverse geographical locations were examined for variation in secondary growth, biomechanical properties, cell wall glycan content, cellulose microfibril angle (MFA) and flowering time. The effect of daylength was also examined. Secondary growth in rosette and inflorescence stems was observed in all accessions. Organised cellulose microfibrils in inflorescence stems were found in plants grown under long and short days. A substantial range of phenotypic variation was found in biochemical and wood-related biophysical characteristics, particularly for tensile strength, tensile stiffness, MFA and some cell wall components. The four monosaccharides galactose, arabinose, rhamnose and fucose strongly correlated with each other as well as with tensile strength and MFA, consistent with mutations in arabinogalactan protein and fucosyl- and xyloglucan galactosyl-transferase genes that result in decreases in strength. Conversely, these variables showed negative correlations with lignin content. Our data support the notion that large-scale natural variation studies of wood-related biomechanical and biochemical properties of inflorescence stems will be useful for the identification of novel genes important for wood formation and quality, and therefore biomaterial and renewable biofuel production.

Fasciclin-like arabinogalactan proteins: specialization for stem biomechanics and cell wall architecture in Arabidopsis and Eucalyptus.

The ancient cell adhesion fasciclin (FAS) domain is found in bacteria, fungi, algae, insects and animals, and occurs in a large family of fasciclin-like arabinogalactan proteins (FLAs) in higher plants. Functional roles for FAS-containing proteins have been determined for insects, algae and vertebrates; however, the biological functions of the various higher-plant FLAs are not clear. Expression of some FLAs has been correlated with the onset of secondary-wall cellulose synthesis in Arabidopsis stems, and also with wood formation in the stems and branches of trees, suggesting a biological role in plant stems. We examined whether FLAs contribute to plant stem biomechanics. Using phylogenetic, transcript abundance and promoter-GUS fusion analyses, we identified a conserved subset of single FAS domain FLAs (group A FLAs) in Eucalyptus and Arabidopsis that have specific and high transcript abundance in stems, particularly in stem cells undergoing secondary-wall deposition, and that the phylogenetic conservation appears to extend to other dicots and monocots. Gene-function analyses revealed that Arabidopsis T-DNA knockout double mutant stems had altered stem biomechanics with reduced tensile strength and a reduced tensile modulus of elasticity, as well as altered cell-wall architecture and composition, with increased cellulose microfibril angle and reduced arabinose, galactose and cellulose content. Using materials engineering concepts, we relate the effects of these FLAs on cell-wall composition with stem biomechanics. Our results suggest that a subset of single FAS domain FLAs contributes to plant stem strength by affecting cellulose deposition, and to the stem modulus of elasticity by affecting the integrity of the cell-wall matrix.

Selective deactivation of gibberellins below the shoot apex is critical to flowering but not to stem elongation of Lolium.

Gibberellins (GAs) cause dramatic increases in plant height and a genetic block in the synthesis of GA(1) explains the dwarfing of Mendel's pea. For flowering, it is GA(5) which is important in the long-day (LD) responsive grass, Lolium. As we show here, GA(1) and GA(4) are restricted in their effectiveness for flowering because they are deactivated by C-2 hydroxylation below the shoot apex. In contrast, GA(5) is effective because of its structural protection at C-2. Excised vegetative shoot tips rapidly degrade [14C]GA(1), [14C]GA(4), and [14C]GA(20) (>80% in 6 h), but not [14C]GA(5). Coincidentally, genes encoding two 2beta-oxidases and a putative 16-17-epoxidase were most expressed just below the shoot apex (<3 mm). Further down the immature stem (>4 mm), expression of these GA deactivation genes is reduced, so allowing GA(1) and GA(4) to promote sub-apical stem elongation. Subsequently, GA degradation declines in florally induced shoot tips and these GAs can become active for floral development. Structural changes which stabilize GA(4) confirm the link between florigenicity and restricted GA 2beta-hydroxylation (e.g. 2alpha-hydroxylation and C-2 di-methylation). Additionally, a 2-oxidase inhibitor (Trinexapac Ethyl) enhanced the activity of applied GA(4), as did limiting C-16,17 epoxidation in 16,17-dihydro GAs or after C-13 hydroxylation. Overall, deactivation of GA(1) and GA(4) just below the shoot apex effectively restricts their florigenicity in Lolium and, conversely, with GA(5), C-2 and C-13 protection against deactivation allows its high florigenicity. Speculatively, such differences in GA access to the shoot apex of grasses may be important for separating floral induction from inflorescence emergence and thus could influence their survival under conditions of herbivore predation.

beta-tubulin affects cellulose microfibril orientation in plant secondary fibre cell walls.

Cellulose microfibrils are the major structural component of plant secondary cell walls. Their arrangement in plant primary cell walls, and its consequent influence on cell expansion and cellular morphology, is directed by cortical microtubules; cylindrical protein filaments composed of heterodimers of alpha- and beta-tubulin. In secondary cell walls of woody plant stems the orientation of cellulose microfibrils influences the strength and flexibility of wood, providing the physical support that has been instrumental in vascular plant colonization of the troposphere. Here we show that a Eucalyptus grandisbeta-tubulin gene (EgrTUB1) is involved in determining the orientation of cellulose microfibrils in plant secondary fibre cell walls. This finding is based on RNA expression studies in mature trees, where we identified and isolated EgrTUB1 as a candidate for association with wood-fibre formation, and on the analysis of somatically derived transgenic wood sectors in Eucalyptus. We show that cellulose microfibril angle (MFA) is correlated with EgrTUB1 expression, and that MFA was significantly altered as a consequence of stable transformation with EgrTUB1. Our findings present an important step towards the production of fibres with altered tensile strength, stiffness and elastic properties, and shed light on one of the molecular mechanisms that has enabled trees to dominate terrestrial ecosystems.

Flowering of the grass Lolium perenne: effects of vernalization and long days on gibberellin biosynthesis and signaling.

Almost 50 years ago, it was shown that gibberellin (GA) applications caused flowering in species normally responding to cold (vernalization) and long day (LD). The implication that GAs are involved with vernalization and LD responses is examined here with the grass Lolium perenne. This species has an obligatory requirement for exposure to both vernalization and LD for its flowering (inflorescence initiation). Specific effects of vernalization or LD on GA synthesis, content, and action have been documented using four treatment pairs: nonvernalized or vernalized plants exposed to short days (SDs) or LDs. Irrespective of vernalization status, exposure to two LDs increased expression of L. perenne GA 20-oxidase-1 (LpGA20ox1), a critical GA biosynthetic gene, with endogenous GAs increasing by up to 5-fold in leaf and shoot. In parallel, LD led to degradation of a DELLA protein, SLENDER (within 48 h of LD or within 2 h of GA application). There was no effect on GA catabolism or abscisic acid content. Loss of SLENDER, which is a repressor of GA signaling, confirms the physiological relevance of increased GA content in LD. For flowering, applied GA replaced the need for LD but not that for vernalization. Thus, GAs may be an LD, leaf-sourced hormonal signal for flowering of L. perenne. By contrast, vernalization had little impact on GA or SLENDER levels or on SLENDER degradation following GA application. Thus, although vernalization and GA are both required for flowering of L. perenne, GA signaling is independent of vernalization that apparently impacts on unrelated processes.