PHYTOCHROME-DEPENDENT LATE-FLOWERING accelerates flowering through physical interactions with phytochrome B and CONSTANS.
ABSTRACT: In flowering plants, light is one of the major environmental stimuli that determine the timing of the transition from the vegetative to reproductive phase. In Arabidopsis, phytochrome B (phyB); phyA; cryptochrome 2; and flavin-binding, KELCH repeat, F-BOX 1 are major photoreceptors that regulate flowering. Unlike phyA; cryptochrome 2; and flavin-binding, KELCH repeat, F-BOX 1, phyB delays flowering mainly by destabilizing the CONSTANS (CO) protein, whose reduction leads to decreased expression of a florigen gene, flowering locus T. However, it remains unclear how the phyB-mediated CO destabilization is mechanistically regulated. Here, we identify a unique phytochrome-dependent late-flowering (PHL) gene, which is mainly involved in the phyB-dependent regulation of flowering. Plants with mutant phl exhibited a late-flowering phenotype, especially under long-day conditions. The late-flowering phenotype of the phl mutant was completely overridden by a phyB mutation, indicating that PHL normally accelerates flowering by countering the inhibitory effect of phyB on flowering. Accordingly, PHL physically interacted with phyB both in vitro and in vivo in a red light-dependent manner. Furthermore, in the presence of phyB under red light, PHL interacted with CO as well. Taken together, we propose that PHL regulates photoperiodic flowering by forming a phyB-PHL-CO tripartite complex.
Project description:The psi2 mutant of Arabidopsis displays amplification of the responses controlled by the red/far red light photoreceptors phytochrome A (phyA) and phytochrome B (phyB) but no apparent defect in blue light perception. We found that loss-of-function alleles of the protein phosphatase 7 (AtPP7) are responsible for the light hypersensitivity in psi2 demonstrating that AtPP7 controls the levels of phytochrome signaling. Plants expressing reduced levels of AtPP7 mRNA display reduced blue-light induced cryptochrome signaling but no noticeable deficiency in phytochrome signaling. Our genetic analysis suggests that phytochrome signaling is enhanced in the AtPP7 loss of function alleles, including in blue light, which masks the reduced cryptochrome signaling. AtPP7 has been found to interact both in yeast and in planta assays with nucleotide-diphosphate kinase 2 (NDPK2), a positive regulator of phytochrome signals. Analysis of ndpk2-psi2 double mutants suggests that NDPK2 plays a critical role in the AtPP7 regulation of the phytochrome pathway and identifies NDPK2 as an upstream element involved in the modulation of the salicylic acid (SA)-dependent defense pathway by light. Thus, cryptochrome- and phytochrome-specific light signals synchronously control their relative contribution to the regulation of plant development. Interestingly, PP7 and NDPK are also components of animal light signaling systems.
Project description:Plants have developed sophisticated systems to monitor and rapidly acclimate to environmental fluctuations. Light is an essential source of environmental information throughout the plant's life cycle. The model plant Arabidopsis thaliana possesses five phytochromes (phyA-phyE) with important roles in germination, seedling establishment, shade avoidance, and flowering. However, our understanding of the phytochrome signaling network is incomplete, and little is known about the individual roles of phytochromes and how they function cooperatively to mediate light responses. Here, we used a bottom-up approach to study the phytochrome network. We added each of the five phytochromes to a phytochrome-less background to study their individual roles and then added the phytochromes by pairs to study their interactions. By analyzing the 16 resulting genotypes, we revealed unique roles for each phytochrome and identified novel phytochrome interactions that regulate germination and the onset of flowering. Furthermore, we found that ambient temperature has both phytochrome-dependent and -independent effects, suggesting that multiple pathways integrate temperature and light signaling. Surprisingly, none of the phytochromes alone conferred a photoperiodic response. Although phyE and phyB were the strongest repressors of flowering, both phyB and phyC were needed to confer a flowering response to photoperiod. Thus, a specific combination of phytochromes is required to detect changes in photoperiod, whereas single phytochromes are sufficient to respond to light quality, indicating how phytochromes signal different light cues.
Project description:BACKGROUND:Phytochromes are a family of red/far-red photoreceptors that regulate a number of important developmental traits in cotton (Gossypium spp.), including plant architecture, fiber development, and photoperiodic flowering. Little is known about the composition and evolution of the phytochrome gene family in diploid (G. herbaceum, G. raimondii) or allotetraploid (G. hirsutum, G. barbadense) cotton species. The objective of this study was to obtain a preliminary inventory and molecular-evolutionary characterization of the phytochrome gene family in cotton. RESULTS:We used comparative sequence resources to design low-degeneracy PCR primers that amplify genomic sequence tags (GSTs) for members of the PHYA, PHYB/D, PHYC and PHYE gene sub-families from A- and D-genome diploid and AD-genome allotetraploid Gossypium species. We identified two paralogous PHYA genes (designated PHYA1 and PHYA2) in diploid cottons, the result of a Malvaceae-specific PHYA gene duplication that occurred approximately 14 million years ago (MYA), before the divergence of the A- and D-genome ancestors. We identified a single gene copy of PHYB, PHYC, and PHYE in diploid cottons. The allotetraploid genomes have largely retained the complete gene complements inherited from both of the diploid genome ancestors, with at least four PHYA genes and two genes encoding PHYB, PHYC and PHYE in the AD-genomes. We did not identify a PHYD gene in any cotton genomes examined. CONCLUSIONS:Detailed sequence analysis suggests that phytochrome genes retained after duplication by segmental duplication and allopolyploidy appear to be evolving independently under a birth-and-death-process with strong purifying selection. Our study provides a preliminary phytochrome gene inventory that is necessary and sufficient for further characterization of the biological functions of each of the cotton phytochrome genes, and for the development of 'candidate gene' markers that are potentially useful for cotton improvement via modern marker-assisted selection strategies.
Project description:Phytochromes are the main plant photoreceptors regulating multiple developmental processes. However, the regulatory network of phytochrome-mediated plant reproduction has remained largely unexplored. There are three phytochromes in rice, phyA, phyB and phyC. No changes in fertility are observed in the single mutants, whereas the seed-setting rate of the phyA phyB double mutant is significantly reduced. Histological and cytological analyses showed that the reduced fertility of the phyA phyB mutant was due to defects in both anther and pollen development. The four anther lobes in the phyA phyB mutant were developed at different stages with fewer pollen grains, most of which were aborted. At the mature stage, more than one lobe in the double mutant was just consisted of several cell layers. To identify genes involved in phytochrome-mediated anther development, anther transcriptomes of phyA, phyB and phyA phyB mutants were compared to that of wild-type rice respectively. Analysis of 2,241 double-mutant-specific differentially expressed transcripts revealed that the metabolic profiles, especially carbohydrate metabolism, were altered greatly, and heat-shock responses were activated in the double mutant. This study firstly provides valuable insight into the complex regulatory networks underlying phytochrome-mediated anther and pollen development in plants, and offers novel clues for hybrid rice breeding.
Project description:<h4>Background</h4>PhyC levels have been observed to be markedly lower in phyB mutants than in Arabidopsis or rice wild type etiolated seedlings, but the mechanism of this phenomenon has not been fully elucidated.<h4>Results</h4>In the present study, we investigated the mechanism by which phyB affects the protein concentration and photo-sensing abilities of phyC and demonstrated that rice phyC exists predominantly as phyB/phyC heterodimers in etiolated seedlings. PHYC-GFP protein was detected when expressed in phyA phyC mutants, but not in phyA phyB mutants, suggesting that phyC requires phyB for its photo-sensing abilities. Interestingly, when a mutant PHYB gene that has no chromophore binding site, PHYB(C364A), was introduced into phyB mutants, the phyC level was restored. Moreover, when PHYB(C364A) was introduced into phyA phyB mutants, the seedlings exhibited de-etiolation under both far-red light (FR) and red light (R) conditions, while the phyA phyB mutants were blind to both FR and R. These results are the first direct evidence that phyC is responsible for regulating seedling de-etiolation under both FR and R. These findings also suggest that phyB is indispensable for the expression and function of phyC, which depends on the formation of phyB/phyC heterodimers.<h4>Significance</h4>The present report clearly demonstrates the similarities and differences in the properties of phyC between Arabidopsis and rice and will advance our understanding of phytochrome functions in monocots and dicots.
Project description:It has been reported that Arabidopsis phytochrome (phy) A and phyB are crucial photoreceptors that display synergistic and antagonistic action during seedling de-etiolation in multiple light signaling pathways. However, the functional relationship between phyA and phyB is not fully understood under different kinds of light and in response to different intensities of such light. In this work, we compared hypocotyl elongation of the phyA-211 phyB-9 double mutant with the wild type, the phyA-211 and phyB-9 single mutants under different intensities of far-red (FR), red (R), blue (B) and white (W) light. We confirmed that phyA and phyB synergistically promote seedling de-etiolation in B-, B plus R-, W- and high R-light conditions. The correlation of endogenous ELONGATED HYPOCOTYL 5 (HY5) protein levels with the trend of hypocotyl elongation of all lines indicate that both phyA and phyB promote seedling photomorphogenesis in a synergistic manner in high-irradiance white light. Gene expression analyses of RBCS members and HY5 suggest that phyB and phyA act antagonistically on seedling development under FR light.
Project description:To begin the study of phytochrome signaling in maize, we have cloned and characterized the phytochrome gene family from the inbred B73. Through DNA gel blot analysis of maize genomic DNA and BAC library screens, we show that the PhyA, PhyB, and PhyC genes are each duplicated once in the genome of maize. Each gene pair was positioned to homeologous regions of the genome using recombinant inbred mapping populations. These results strongly suggest that the duplication of the phytochrome gene family in maize arose as a consequence of an ancient tetraploidization in the maize ancestral lineage. Furthermore, sequencing of Phy genes directly from BAC clones indicates that there are six functional phytochrome genes in maize. Through Northern gel blot analysis and a semiquantitative reverse transcriptase polymerase chain reaction assay, we determined that all six phytochrome genes are transcribed in several seedling tissues. However, expression from PhyA1, PhyB1, and PhyC1 predominate in all seedling tissues examined. Dark-grown seedlings express higher levels of PhyA and PhyB than do light-grown plants but PhyC genes are expressed at similar levels under light and dark growth conditions. These results are discussed in relation to phytochrome gene regulation in model eudicots and monocots and in light of current genome sequencing efforts in maize.
Project description:Light signaling by phytochrome B in long days inhibits flowering in sorghum by increasing expression of the long day floral repressors PSEUDORESPONSE REGULATOR PROTEIN (SbPRR37, Ma1) and GRAIN NUMBER, PLANT HEIGHT AND HEADING DATE 7 (SbGHD7, Ma6). SbPRR37 and SbGHD7 RNA abundance peaks in the morning and in the evening of long days through coordinate regulation by light and output from the circadian clock. 58 M, a phytochrome B deficient (phyB-1, ma3R) genotype, flowered ?60 days earlier than 100 M (PHYB, Ma3) in long days and ?11 days earlier in short days. Populations derived from 58 M (Ma1, ma3R, Ma5, ma6) and R.07007 (Ma1, Ma3, ma5, Ma6) varied in flowering time due to QTL aligned to PHYB/phyB-1 (Ma3), Ma5, and GHD7/ghd7-1 (Ma6). PHYC was proposed as a candidate gene for Ma5 based on alignment and allelic variation. PHYB and Ma5 (PHYC) were epistatic to Ma1 and Ma6 and progeny recessive for either gene flowered early in long days. Light signaling mediated by PhyB was required for high expression of the floral repressors SbPRR37 and SbGHD7 during the evening of long days. In 100 M (PHYB) the floral activators SbEHD1, SbCN8 and SbCN12 were repressed in long days and de-repressed in short days. In 58 M (phyB-1) these genes were highly expressed in long and short days. Furthermore, SbCN15, the ortholog of rice Hd3a (FT), is expressed at low levels in 100 M but at high levels in 58 M (phyB-1) regardless of day length, indicating that PhyB regulation of SbCN15 expression may modify flowering time in a photoperiod-insensitive manner.
Project description:Plants sense and respond to red and far-red light using the phytochrome (phy) family of photoreceptors. However, the mechanism of light signal transduction is not well defined. Here, we report the identification of a new mutant Arabidopsis locus, srl2 (short under red-light 2), which confers selective hypersensitivity to continuous red, but not far-red, light. This hypersensitivity is eliminated in srl2phyB, but not srl2phyA, double mutants, indicating that this locus functions selectively and negatively in phyB signaling. The SRL2 gene encodes a bHLH factor, designated PIF4 (phytochrome-interacting factor 4), which binds selectively to the biologically active Pfr form of phyB, but has little affinity for phyA. Despite its hypersensitive morphological phenotype, the srl2 mutant displays no perturbation of light-induced expression of marker genes for chloroplast development. These data suggest that PIF4 may function specifically in a branch of the phyB signaling network that regulates a subset of genes involved in cell expansion. Consistent with this proposal, PIF4 localizes to the nucleus and can bind to a G-box DNA sequence motif found in various light-regulated promoters.
Project description:Among the various abiotic stresses, water and nitrogen are major stress factors that limit crop productivity worldwide. Since leaf nutrients remobilization during leaf senescence might impact response to abiotic stress in crops, we undertook a forward screen of the <i>Mutator</i>-active approach to identify <i>premature senescence</i> loci in maize. A mutant line isolated from a cross between a Pioneer Brand elite line and a public <i>Mutator</i>-active material, designated <i>premature senescence2</i> (<i>pre2</i>), expressed leaf senescence during flower initiation. The <i>Pre2</i> gene encodes PHYTOCHROME-DEPENDENT LATE-FLOWERING (PHL) protein, a nuclear receptor coactivator. The <i>pre2-1</i> mutant allele was not a null mutation but produced a functional wild-type transcript along with multiple mRNA species of varying lengths resulting from the alternate splicing of the <i>Pre2</i> gene. The PHL accelerates flowering by suppressing the inhibitory effect of <i>phyB</i> on flowering in Arabidopsis (Endo et al., 2013). The ZmPRE2 polypeptide is highly conserved in plant species and has two identifiable motifs namely SPT20 and MED15. The Spt20 domain, which is a part of the SAGA (Spt-Ada-Gcn5 acetyltransferase) complex, is involved in histone deacetylation and MED15 proteins have nuclear functions in mediating DNA Pol II transcription. The differential spliced mature transcripts in both the <i>pre2</i> alleles, as a result of transposon interference, were producing truncated proteins that lacked polyglutamine (Q) tract near the C-terminus and might be causative of the premature senescence phenotype in maize. Endogenous gene suppression of <i>ZmPre2</i> by RNAi improves maize agronomic performance under both water stress and suboptimal nitrogen conditions. The homozygous T-DNA knockout of the <i>pre2</i> homolog in Arabidopsis (At1G72390; the same insertional allele used by Endo et al., 2013) results in higher biomass, delayed maturity, enhanced tolerance to drought, and improved nitrogen utilization efficiency. The Arabidopsis mutant also showed hypersensitive response to 1 µM ABA (abscisic acid) concentration. These results indicate that the PHL protein plays a direct or indirect role in ABA-dependent drought and N signaling pathways.