Rod W. King

Abscisic acid in developing wheat grains and its relationship to grain growth and maturation.

SummaryDuring the later stages of growth of grains of wheat (Triticum aestivum L. cvs. WW15 and Gabo) there is a dramatic increase (up to 40fold) in the content of abscisic acid (ABA) to 4–6 ng per grain. This level remains high from 25 to 40 days after anthesis. Then, in association with natural or forced drying of the grain, there is a rapid drop (5–10 fold) in the ABA content and a brief increase in the content of bound ABA. The bulk of ABA in an ear was in the grain (95%) and although the embryo contributed 19% of this ABA it was less than 5% of the grain by weight. There was no clear relationship between ABA content and the growth of grains in various spikelet or floret positions. Application of (±)-ABA to the ear had no effect on grain growth rate but led to an earlier cessation of grain growth and hastened the drying of the grain. Isolated embryos and whole grains were capable of germinating during the mid grain growth period (15–25 days), but germination capacity declined subsequently as ABA accumulated. Later, still, with grain drying and loss of ABA, embryo and grain became germinable again. At this time there was also a dramatic increase in the ability of the grain to synthesize α-amylase. It is suggested that the accumulation of ABA at the later stages of grain growth prevents precocious germination and premature hydrolysis of starch reserves of the morphologically mature but still unripe grain. An inevitable consequence of such action may be in triggering grain maturation.

The Involvement of Gibberellin 20-Oxidase Genes in Phytochrome-Regulated Petiole Elongation of Arabidopsis

Long day (LD) exposure of rosette plants causes rapid stem/petiole elongation, a more vertical growth habit, and flowering; all changes are suggestive of a role for the gibberellin (GA) plant growth regulators. For Arabidopsis (Arabidopsis thaliana) L. (Heynh), we show that enhancement of petiole elongation by a far-red (FR)-rich LD is mimicked by a brief (10 min) end-of-day (EOD) FR exposure in short day (SD). The EOD response shows red (R)/FR photoreversibility and is not affected in a phytochrome (PHY) A mutant so it is mediated by PHYB and related PHYs. FR photoconversion of PHYB to an inactive form activates a signaling pathway, leading to increased GA biosynthesis. Of 10 GA biosynthetic genes, expression of the 20-oxidase, AtGA20ox2, responded most to FR (up to a 40-fold increase within 3 h). AtGA20ox1 also responded but to a lesser extent. Stimulation of petiole elongation by EOD FR is reduced in a transgenic AtGA20ox2 hairpin gene silencing line. By contrast, it was only in SD that a T-DNA insertional mutant of AtGA20ox1 (ga5-3) showed reduced response. Circadian entrainment to a daytime pattern provides an explanation for the SD expression of AtGA20ox1. Conversely, the strong EOD/LD FR responses of AtGA20ox2 may reflect its independence of circadian regulation. While FR acting via PHYB increases expression of AtGA20ox2, other GA biosynthetic genes are known to respond to R rather than FR light and/or to other PHYs. Thus, there must be different signal transduction pathways, one at least showing a positive response to active PHYB and another showing a negative response.

Flowering Responses to Altered Expression of Phytochrome in Mutants and Transgenic Lines of Arabidopsis thaliana (L.) Heynh.

The long-day plant Arabidopsis thaliana (L.) Heynh. flowers early in response to brief end-of-day (EOD) exposures to far-red light (FR) following a fluorescent short day of 8 h. FR promotion of flowering was nullified by subsequent brief red light (R) EOD exposure, indicating phytochrome involvement. The EOD response to R or FR is a robust measure of phytochrome action. Along with their wild-type (WT) parents, mutants deficient in either phytochrome A or B responded similarly to the EOD treatments. Thus, neither phytochrome A nor B exclusively regulated flowering, although phytochrome B controlled hypocotyl elongation. Perhaps a third phytochrome species is important for the EOD responses of the mutants and/or their flowering is regulated by the amount of the FR-absorbing form of phytochrome, irrespective of the phytochrome species. Overexpression of phytochrome A or phytochrome B resulted in differing photoperiod and EOD responses among the genotypes. The day-neutral overexpressor of phytochrome A had an EOD response similar to all of the mutants and WTs, whereas R EOD exposure promoted flowering in the overexpressor of phytochrome B and FR EOD exposure inhibited this promotion. The comparisons between relative flowering times and leaf numbers at flowering of the overexpressors and their WTs were not consistent across photoperiods and light treatments, although both phytochromes A and B contributed to regulating flowering of the transgenic plants.

Regulation of Flowering in the Long-Day Grass Lolium temulentum by Gibberellins and the FLOWERING LOCUS T Gene

Seasonal control of flowering often involves leaf sensing of daylength coupled to time measurement and generation and transport of florigenic signals to the shoot apex. We show that transmitted signals in the grass Lolium temulentum may include gibberellins (GAs) and the FLOWERING LOCUS T (FT) gene. Within 2 h of starting a florally inductive long day (LD), expression of a 20-oxidase GA biosynthetic gene increases in the leaf; its product, GA20, then increases 5.7-fold versus short day; its substrate, GA19, decreases equivalently; and a bioactive product, GA5, increases 4-fold. A link between flowering, LD, GAs, and GA biosynthesis is shown in three ways: (1) applied GA19 became florigenic on exposure to LD; (2) expression of LtGA20ox1, an important GA biosynthetic gene, increased in a florally effective LD involving incandescent lamps, but not with noninductive fluorescent lamps; and (3) paclobutrazol, an inhibitor of an early step of GA biosynthesis, blocked flowering, but only if applied before the LD. Expression studies of a 2-oxidase catabolic gene showed no changes favoring a GA increase. Thus, the early LD increase in leaf GA5 biosynthesis, coupled with subsequent doubling in GA5 content at the shoot apex, provides a substantial trail of evidence for GA5 as a LD florigen. LD signaling may also involve transport of FT mRNA or protein because expression of LtFT and LtCONSTANS increased rapidly, substantially (>80-fold for FT), and independently of GA. However, because a LD from fluorescent lamps induced LtFT expression but not flowering, the nature of the light response of FT requires clarification.

The nature of floral signals in Arabidopsis. II. Roles for FLOWERING LOCUS T (FT) and gibberellin

Signals produced in leaves are transported to the shoot apex where they cause flowering. Protein of the gene FLOWERING LOCUS T (FT) is probably a long day (LD) signal in Arabidopsis. In the companion paper, rapid LD increases in FT expression associated with flowering driven photosynthetically in red light were documented. In a far red (FR)-rich LD, along with FT there was a potential role for gibberellin (GA). Here, with the GA biosynthesis dwarf mutant ga1-3, GA4-treated plants flowered after 26 d in short days (SD) but untreated plants were still vegetative after 6 months. Not only was FT expression low in SD but applied GA bypassed some of the block to flowering in ft-1. On transfer to LD, ga1-3 only flowered when treated simultaneously with GA, and FT expression increased rapidly (<19.5 h) and dramatically (15-fold). In contrast, in the wild type in LD there was little requirement for GA for FT increase and flowering so its endogenous GA content was near to saturating. Despite this permissive role for endogenous GA in Columbia, RNA interference (RNAi) silencing of the GA biosynthesis gene, GA 20-OXIDASE2, revealed an additional, direct role for GA in LD. Flowering took twice as long after silencing the LD-regulated gene, GA 20-OXIDASE2. Such independent LD input by FT and GA reflects their non-sympatric expression (FT in the leaf blade and GA 20-OXIDASE2 in the petiole). Overall, FT acts as the main LD floral signal in Columbia and GA acts on flowering both via and independently of FT.

Blue-light promotion of flowering is absent in hy4 mutants of Arabidopsis

Loss of a blue-light photoreceptor in the hy4 mutants of Arabidopsis thaliana (L.) Heynh substantially delayed flowering (>100 d to flower vs. 40–50 d), especially with blue light exposure from lamps lacking much red (R) and/or far-red (FR) light. Red night breaks were promotory but flowering was still later for the hy4-101 mutant. However, with exposure to light from FR-rich lamps, flowering of all mutants was early and no different from the wild type. Thus, flowering of Arabidopsis involves a blue-light photoreceptor and other, often more effective photoreceptors. The latter may involve phytochrome photoresponses to R and FR, but with little or no phytochrome response to blue wavelengths.

Inhibition of tiller bud outgrowth in the tin mutant of wheat is associated with precocious internode development

Tillering (branching) is a major yield component and, therefore, a target for improving the yield of crops. However, tillering is regulated by complex interactions of endogenous and environmental signals, and the knowledge required to achieve optimal tiller number through genetic and agronomic means is still lacking. Regulatory mechanisms may be revealed through physiological and molecular characterization of naturally occurring and induced tillering mutants in the major crops. Here we characterize a reduced tillering (tin, for tiller inhibition) mutant of wheat (Triticum aestivum). The reduced tillering in tin is due to early cessation of tiller bud outgrowth during the transition of the shoot apex from the vegetative to the reproductive stage. There was no observed difference in the development of the main stem shoot apex between tin and the wild type. However, tin initiated internode development earlier and, unlike the wild type, the basal internodes in tin were solid rather than hollow. We hypothesize that tin represents a novel type of reduced tillering mutant associated with precocious internode elongation that diverts sucrose (Suc) away from developing tillers. Consistent with this hypothesis, we have observed upregulation of a gene induced by Suc starvation, downregulation of a Suc-inducible gene, and a reduced Suc content in dormant tin buds. The increased expression of the wheat Dormancy-associated (DRM1-like) and Teosinte Branched1 (TB1-like) genes and the reduced expression of cell cycle genes also indicate bud dormancy in tin. These results highlight the significance of Suc in shoot branching and the possibility of optimizing tillering by manipulating the timing of internode elongation.

The nature of floral signals in Arabidopsis. I. Photosynthesis and a far-red photoresponse independently regulate flowering by increasing expression of FLOWERING LOCUS T (FT)

Arabidopsis flowers in long day (LD) in response to signals transported from the photoinduced leaf to the shoot apex. These LD signals may include protein of the gene FLOWERING LOCUS T (FT) while in short day (SD) with its slower flowering, signalling may involve sucrose and gibberellin. Here, it is shown that after 5 weeks growth in SD, a single LD up-regulated leaf blade expression of FT and CONSTANS (CO) within 4–8 h, and flowers were visible within 2–3 weeks. Plants kept in SDs were still vegetative 7 weeks later. This LD response was blocked in ft-1 and a co mutant. Exposure to different LD light intensities and spectral qualities showed that two LD photoresponses are important for up-regulation of FT and for flowering. Phytochrome is effective at a low intensity from far-red (FR)-rich incandescent lamps. Independently, photosynthesis is active in an LD at a high intensity from red (R)-rich fluorescent lamps. The photosynthetic role of a single high light LD is demonstrated here by the blocking of the flowering and FT increase on removal of atmospheric CO2 or by decreasing the LD light intensity by 10-fold. These conditions also reduced leaf blade sucrose content and photosynthetic gene expression. An SD light integral matching that in a single LD was not effective for flowering, although there was reasonable FT-independent flowering after 12 SD at high light. While a single photosynthetic LD strongly amplified FT expression, the ability to respond to the LD required an additional but unidentified photoresponse. The implications of these findings for studies with mutants and for flowering in natural conditions are discussed.

Temperature-dependent feedback inhibition of photosynthesis in peanut

Arachis hypogaea L. is a tropical crop that is slow-growing at temperatures below 25°C. Unadapted CO2-assimilation rate (A) showed insufficient variation between 15 and 30°C in the short term (hours) to explain this marked reduction in growth. However, at longer periods (12 d), A was depressed as were growth rate and leafproduction rate. To examine the possible relationship between growth, A and sink demand plants were transferred from 30°C, which is near the optimum for growth, to a suboptimal temperature (19°C). In the first 2 d of cooling, A decreased by 50–70%, the stomata stayed open, and the intercellular CO2 concentration (ci) rose, i.e. the decrease in A of the cooled plants was the result of non-stomatal factors. Changes in dark respiration did not account for the decline in A.Clear evidence was obtained of sink control of A by independently manipulating the temperature of different leaves on the plant. Cooling (to 19°C) most of the plant (the sink) led to a 70% decline in A of the remaining leaves at 30°C after 3 d, whereas the converse treatments (30°C sink, 19°C source) resulted in small changes (17%). In plants at 19°C which were exposed to low CO2 concentration to prevent photosynthesis, A was not reduced when measured at normal CO2 concentrations, indicating that carbohydrate accumulation was responsible for the decline in A. Dry-matter build-up at suboptimal temperature was also consistent with end-product inhibition of photosynthesis.

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.