Genome-wide identification of the Pinus tabuliformis CONSTANS-like gene family and their potential roles in reproductive cone development.

The CONSTANS-like (COL) genes, as a core transcription factor in the photoperiod regulation pathway, play a key role in plant reproduction development. However, their molecular characterization has rarely been studied in Pinus tabuliformis. Here, 10 PtCOL genes were identified in the P. tabuliformis genome and multiple sequence alignments have indicated that the PtCOL proteins contained highly conserved B-BOX1 and CCT domains. Sequence similarity analysis showed that PtCOL1 and PtCOL3 had the higher similarity with Norway spruce COLs (PaCOL2 and PaCOL1) and Arabidopsis COLs (AtCOL3, 4 and 5), respectively. Phylogeny and gene structure analyses revealed that PtCOLs were divided into three subgroups, each with identical or similar distributions of exons, introns, and motifs. Moreover, 10 PtCOLs were distributed on 6 chromosomes and PtCOL9 has syntenic gene pairs in both Ginkgo biloba and Sequoiadendron giganteum. Interestingly, in transcriptome profiles, most PtCOLs exhibited a diurnal oscillation pattern under both long (LD) and short (SD) day conditions. Additionally, PtCOLs were highly expressed in needles and female cones, and showed different spatial expression patterns. Among the ten PtCOLs, PtCOL1/3 heterologous overexpression Arabidopsis displayed a delayed-flowering phenotype under SD, indicating that they are likely to play a crucial role in the reproductive development. Additionally, PtCOL1 and PtCOL3 were not only capable of interacting with each other, but they were each capable of interacting with themselves. Furthermore, PtCOL1 and PtCOL3 were also involved in the MADS-box protein-protein interaction (PPI) network in P. tabuliformis cone development. Direct interactions of PtDAL11 with PtCOL1/3 impeded PtCOL1/3 translocation into the nucleus. In summary, this study provided comprehensive understanding for the functions of the PtCOL gene family and revealed their biological roles in the photoperiod-dependent P. tabuliformis cone development.

Involvement of CONSTANS-like Proteins in Plant Flowering and Abiotic Stress Response.

The process of flowering in plants is a pivotal stage in their life cycle, and the CONSTANS-like (COL) protein family, known for its photoperiod sensing ability, plays a crucial role in regulating plant flowering. Over the past two decades, homologous genes of COL have been identified in various plant species, leading to significant advancements in comprehending their involvement in the flowering pathway and response to abiotic stress. This article presents novel research progress on the structural aspects of COL proteins and their regulatory patterns within transcription complexes. Additionally, we reviewed recent information about their participation in flowering and abiotic stress response, aiming to provide a more comprehensive understanding of the functions of COL proteins.

Identification of the CONSTANS-like family in Cymbidium sinense, and their functional characterization.

BACKGROUND: Cymbidium sinense is an orchid that is typically used as a potted plant, given its high-grade ornamental characteristics, and is most frequently distributed in China and SE Asia. The inability to strictly regulate flowering in this economically important potted and cut-flower orchid is a bottleneck that limits its industrial development. Studies on C. sinense flowering time genes would help to elucidate the mechanism regulating flowering. There are very few studies on the genetic regulation of flowering pathways in C. sinense. Photoperiod significantly affects the flowering of C. sinense, but it was unknown how the CONSTANS gene family is involved in regulating flowering. RESULTS: In this study, eight CONSTANS-like genes were identified and cloned. They were divided into three groups based on a phylogenetic analysis. Five representative CsCOL genes (CsCOL3/4/6/8/9) were selected from the three groups to perform expression characterization and functional study. CsCOL3/4/6/8/9 are nucleus-localized proteins, and all five CsCOL genes were expressed in all organs, mainly in leaves followed by sepals. The expression levels of CsCOL3/4 (group I) were higher in all organs than other CsCOL genes. Developmental stage specific expression revealed that the expression of CsCOL3/4/9 peaked at the initial flowering stage. In contrast, the transcript level of CsCOL6/8 was highest at the pedicel development stage. Photoperiodic experiments demonstrated that the transcripts of the five CsCOL genes exhibited distinct diurnal rhythms. Under LD conditions, the overexpression of CsCOL3/4 promoted early flowering, and CsCOL6 had little effect on flowering time, whereas CsCOL8 delayed flowering of Arabidopsis thaliana. However, under SD conditions, overexpression of CsCOL4/6/8 promoted early flowering and the rosette leaves growth, and CsCOL3 induced flower bud formation in transgenic Arabidopsis. CONCLUSION: The phylogenetic analysis, temporal and spatial expression patterns, photoperiodic rhythms and functional study indicate that CsCOL family members in C. sinense were involved in growth, development and flowering regulation through different photoperiodic pathway. The results will be useful for future research on mechanisms pertaining to photoperiod-dependent flowering, and will also facilitate genetic engineering-based research that uses Cymbidium flowering time genes.

Lack of the CCT domain changes the ability of mango MiCOL14A to resist salt and drought stress in Arabidopsis.

CONSTANS (CO) is the key gene in the photoperiodic pathway that regulates flowering in plants. In this paper, a CONSTANS-like 14A (COL14A) gene was obtained from mango, and its expression patterns and functions were characterized. Sequence analysis shows that MiCOL14A-JH has an additional A base, which leads to code shifting in subsequent coding boxes and loss of the CCT domain. The MiCOL14A-JH and MiCOL14A-GQ genes both belonged to group Ⅲ of the CO/COL gene family. Analysis of tissue expression patterns showed that MiCOL14A was expressed in all tissues, with the highest expression in the leaves of seedling, followed by lower expression levels in the flowers and stems of adult leaves. However, there was no significant difference between different mango varieties. At different development stages of flowering, the expression level of MiCOL14A-GQ was the highest in the leaves before floral induction period, and the lowest at flowering stage, while the highest expression level of MiCOL14A-JH appeared in the leaves at flowering stage. The transgenic functional analysis showed that both MiCOL14A-GQ and MiCOL14A-JH induced delayed flowering of transgenic Arabidopsis. In addition, MiCOL14A-JH enhanced the resistance of transgenic Arabidopsis to drought stress, while MiCOL14A-GQ increased the sensitivity of transgenic Arabidopsis to salt stress. Further proteinprotein interaction analysis showed that MiCOL14A-JH directly interacted with MYB30-INTERACTING E3 LIGASE 1 (MiMIEL1), CBL-interacting protein kinase 9 (MiCIPK9) and zinc-finger protein 4 (MiZFP4), but MiCOL14A-GQ could not interact with these three stress-related proteins. Together, our results demonstrated that MiCOL14A-JH and MiCOL14A-GQ not only regulate flowering but also play a role in the abiotic stress response in mango, and the lack of the CCT domain affects the proteinprotein interaction, thus affecting the gene response to stress. The insertion of an A base can provide a possible detection site for mango resistance breeding.

Overexpression of two CONSTANS-like 2 (MiCOL2) genes from mango delays flowering and enhances tolerance to abiotic stress in transgenic Arabidopsis.

The CO/COL gene family plays an important role in regulating photoperiod-dependent flowering time in plants. In this study, two COL2 gene homologs, MiCOL2A and MiCOL2B, were isolated from 'SiJiMi' mango, and their expression patterns and functions were characterized. The MiCOL2A and MiCOL2B genes both belonged to the group Ⅰ of CO/COL gene family. MiCOL2A and MiCOL2B exhibited distinct circadian rhythms and were highly expressed in leaves during the flowering induction period. Subcellular localization analysis revealed that MiCOL2A and MiCOL2B are localized in the nucleus. The overexpression of MiCOL2A and MiCOL2B significantly delayed flowering time in Arabidopsis under both long-day (LD) and short-day (SD) conditions. The MiCOL2A and MiCOL2B overexpression Arabidopsis plants exhibited more tolerance to slat and drought stress after abiotic stress treatments, with greater ROS scavenging capacity and protective enzyme activity, less cell damage and death and higher expression of stress response genes than wild type plants. Bimolecular fluorescence complementation (BiFC) analysis showed that MiCOL2A and MiCOL2B interacted with several stress-related proteins, including zinc finger protein 4 (MiZFP4), MYB30-INTERACTING E3 LIGASE 1 (MiMIEL1) and RING zinc finger protein 34 (MiRZFP34). The results indicate that MiCOL2A and MiCOL2B are not only involved in flowering time but also play a positive role in abiotic stress responses in plants.

The tomato CONSTANS-LIKE protein SlCOL1 regulates fruit yield by repressing SFT gene expression.

BACKGROUND: CONSTANS (CO) and CONSTANS-LIKE (COL) transcription factors have been known to regulate a series of cellular processes including the transition from the vegetative growth to flower development in plants. However, their role in regulating fruit yield in tomato is poorly understood. RESULT: In this study, the tomato ortholog of Arabidopsis CONSTANS, SlCOL1, was shown to play key roles in the control of flower development and fruit yield. Suppression of SlCOL1 expression in tomato was found to lead to promotion of flower and fruit development, resulting in increased tomato fruit yield. On the contrary, overexpression of SlCOL1 disturbed flower and fruit development, and significantly reduced tomato fruit yield. Genetic and biochemical evidence indicated that SlCOL1 controls inflorescence development by directly binding to the promoter region of tomato inflorescence-associated gene SINGLE-FLOWER TRUSS (SFT) and negatively regulating its expression. Additionally, we found that SlCOL1 can also negatively regulate fruit size in tomato. CONCLUSIONS: Tomato SlCOL1 binds to the promoter of the SFT gene, down-regulates its expression, and plays a key role in reducing the fruit size.

Genome-Wide Identification, Characterization, and Expression Profile Analysis of CONSTANS-like Genes in Woodland Strawberry (Fragaria vesca).

CONSTANS-like (CO-like) gene is one of the most important regulators in the flowering process of the plant, playing a core role in the photoperiodic flowering induction pathway. In this study, we identified 10 distinct CO-like genes (FveCOs) in woodland strawberry (Fragaria vesca). They were classified into three groups with specific gene structure characteristics or protein domains in each group. The effect of selection pressure on the FveCOs in the woodland strawberry was tested by Ka/Ks, and it was shown that the evolution rate of FveCOs was controlled by purification selection factors. Intraspecific synteny analysis of woodland strawberry FveCOs showed that at least one duplication event existed in the gene family members. Collinearity analysis of woodland strawberry genome with genomes of Arabidopsis, rice (Oryza sativa), and apple (Malus × domestica) showed that CO-like genes of F. vesca and Malus × domestica owned higher similarity for their similar genomes compared with those of other two species. The FveCOs showed different tissue-specific expression patterns. Moreover, real-time quantitative PCR results revealed that the expressions of the most FveCOs followed a 24-h rhythm oscillation under both long-day (LD) and short-day (SD) conditions. Further expression analysis showed that the individual expression changing profile of FveCO3 and FveCO5 was opposite to each other under both LD and SD conditions. Moreover, the expression of FveCO3 and FveCO5 was both negatively correlated with the flowering time variation of the woodland strawberry grown under LD and SD conditions, indicating their potential vital roles in the photoperiodic flowering regulation. Further protein interaction network analysis also showed that most of the candidate interaction proteins of FveCO3 and FveCO5 were predicted to be the flowering regulators. Finally, LUC assay indicated that both FveCO3 and FveCO5 could bind to the promoter of FveFT1, the key regulator of flowering regulation in the woodland strawberry, and thus activate its expression. Taken together, this study laid a foundation for understanding the exact roles of FveCOs in the reproductive development regulation of the woodland strawberry, especially in the photoperiodic flowering process.

Major niche transitions in Pooideae correlate with variation in photoperiodic flowering and evolution of CCT domain genes.

The external cues that trigger timely flowering vary greatly across tropical and temperate plant taxa, the latter relying on predictable seasonal fluctuations in temperature and photoperiod. In the grass family (Poaceae) for example, species of the subfamily Pooideae have become specialists of the northern temperate hemisphere, generating the hypothesis that their progenitor evolved a flowering response to long days from a short-day or day-neutral ancestor. Sampling across the Pooideae, we found support for this hypothesis, and identified several secondary shifts to day-neutral flowering and one to short-day flowering in a tropical highland clade. To explain the proximate mechanisms for the secondary transition back to short-day-regulated flowering, we investigated the expression of CCT domain genes, some of which are known to repress flowering in cereal grasses under specific photoperiods. We found a shift in CONSTANS 1 and CONSTANS 9 expression that coincides with the derived short-day photoperiodism of our exemplar species Nassella pubiflora. This sets up the testable hypothesis that trans- or cis-regulatory elements of these CCT domain genes were the targets of selection for major niche shifts in Pooideae grasses.

Isolation and functional analysis of two CONSTANS-like 1 genes from mango.

The CONSTANS-LIKE1 (COL1) gene plays an important role in the regulation of photoperiodic flowering in plants. In this study, two COL1 homolog genes, MiCOL1A and MiCOL1B, were isolated from mango (Mangifera indica L.). The open reading frames of MiCOL1A and MiCOL1B are 852 and 822 bp in length and encode 284 and 274 amino acids, respectively. The MiCOL1A and MiCOL1B proteins contain only one CCT domain and belong to the CO/COL group IV protein family. MiCOL1A and MiCOL1B were expressed both in vegetative and reproductive organs but with expression level differences. MiCOL1A was highly expressed in juvenile and adult leaves, but MiCOL1B was highly expressed in flowers. Seasonal expression analysis showed that MiCOL1A and MiCOL1B have similar expression patterns and higher expression levels during flower induction and flower organ differentiation periods. However, MiCOL1A and MiCOL1B exhibited unstable patterns in circadian expression analysis. MiCOL1A and MiCOL1B were localized in the nucleus and had transcriptional activation activity in yeast. Overexpression of MiCOL1A and MiCOL1B resulted in significantly delayed flowering time in Arabidopsis. Furthermore, we also found that overexpression of MiCOL1A and MiCOL1B enhanced drought tolerance in transgenic Arabidopsis. The results demonstrated that MiCOL1A and MiCOL1B are not only involved in flowering regulation but also play a role in the stress response of plants.

Conservation and Divergence of the CONSTANS-Like (COL) Genes Related to Flowering and Circadian Rhythm in Brassica napus.

The CONSTANS-LIKE (COL) genes are important signaling component in the photoperiod pathway and flowering regulation pathway. However, people still know little about their role in Brassica napus. To achieve a better understanding of the members of the BnaCOL gene family, reveal their evolutionary relationship and related functions involved in photoperiod regulation, we systematically analyzed the BnaCOL family members in B. napus genome. A total of 33 BnaCOL genes distributed unevenly on 16 chromosomes were identified in B. napus and could be classified into three subfamilies. The same subfamilies have relatively conservative gene structures, three-dimensional protein structures and promoter motifs such as light-responsive cis-elements. The collinearity analysis detected 37 pairs of repetitive genes in B. napus genome. A 67.7% of the BnaCOL genes were lost after B. napus genome polyploidization. In addition, the BnaCOL genes showed different tissue-specific expression patterns. A 81.8% of the BnaCOL genes were mainly expressed in leaves, indicating that they may play a conservative role in leaves. Subsequently, we tested the circadian expression profiles of nine homologous genes that regulate flowering in Arabidopsis. Most BnaCOL genes exhibit several types of circadian rhythms, indicating that these BnaCOL genes are involved in the photoperiod pathway. As such, our research has laid the foundation for understanding the exact role of the BnaCOL family in the growth and development of rapeseed, especially in flowering.

Genome-wide identification, genomic organization, and expression profiling of the CONSTANS-like (COL) gene family in petunia under multiple stresses.

BACKGROUND: CONSTANS-like (CO-like, COL) are putative zinc-finger transcription factors known to play vital role in various plant biological processes such as control of flowering time, regulation of plant growth and development and responses to stresses. However, no systematic analysis of COL family gene regarding the plant development and stress response has been previously performed in any solanaceous crop. In the present study, a comprehensive genome-wide analysis of COL family genes in petunia has been conducted to figure out their roles in development of organs and stress response. RESULTS: A total of 33 COL genes, 15 PaCOL genes in P. axillaris and 18 PiCOL genes in P. inflata, were identified in petunia. Subsequently, a genome-wide systematic analysis was performed in 15 PaCOL genes. Considering the domain composition and sequence similarity the 15 PaCOL and 18 PiCOL genes were phylogenetically classified into three groups those are conserved among the flowering plants. Moreover, all of the 15 PaCOL proteins were localized in nucleus. Furthermore, differential expression patterns of PaCOL genes were observed at different developmental stages of petunia. Additionally, transcript expression of 15 PaCOL genes under various abiotic and phytohormone treatments showed their response against stresses. Moreover, several cis-elements related to stress, light-responsive, hormone signaling were also detected in different PaCOL genes. CONCLUSION: The phylogenetic clustering, organ specific expression pattern and stress responsive expression profile of conserved petunia COL genes indicating their involvement in plant growth and development and stress response mechanism. This work provide a significant foundation for understanding the biological roles of petunia COL genes in plant growth, development and in stress response.

Genome-wide identification, expression, and sequence analysis of CONSTANS-like gene family in cannabis reveals a potential role in plant flowering time regulation.

BACKGROUND: Cannabis, an important industrial crop, has a high sensitivity to photoperiods. The flowering time of cannabis is one of its important agronomic traits, and has a significant effect on its yield and quality. The CONSTANS-like (COL) gene plays a key role in the regulation of flowering in this plant. However, the specific roles of the COL gene family in cannabis are still unknown. RESULTS: In this study, 13 CsCOL genes were identified in the cannabis genome. Phylogenetic analysis implied that the CsCOL proteins were divided into three subgroups, and each subgroup included conserved intron/exon structures and motifs. Chromosome distribution analysis showed that 13 CsCOL genes were unevenly distributed on 7 chromosomes, with chromosome 10 having the most CsCOL members. Collinearity analysis showed that two syntenic gene pairs of CsCOL4 and CsCOL11 were found in both rice and Gossypium raimondii. Of the 13 CsCOL genes, CsCOL6 and CsCOL12 were a pair of tandem duplicated genes, whereas CsCOL8 and CsCOL11 may have resulted from segmental duplication. Furthermore, tissue-specific expression showed that 10 CsCOL genes were preferentially expressed in the leaves, 1 CsCOL in the stem, and 2 CsCOL in the female flower. Most CsCOL exhibited a diurnal oscillation pattern under different light treatment. Additionally, sequence analysis showed that CsCOL3 and CsCOL7 exhibited amino acid differences among the early-flowering and late flowering cultivars. CONCLUSION: This study provided insight into the potential functions of CsCOL genes, and highlighted their roles in the regulation of flowering time in cannabis. Our results laid a foundation for the further elucidation of the functions of COL genes in cannabis.

A CONSTANS-LIKE gene of Nelumbo nucifera could promote potato tuberization.

MAIN CONCLUSION: CONSTANS-LIKE 5 of Nelumbo nucifera is capable of promoting potato tuberization through CONSTANS-FLOWERING LOCUS T and gibberellin signaling pathways with a probable association with lotus rhizome enlargement. Lotus (Nelumbo nucifera) is an aquatic plant that is affiliated to the Nelumbonaceace family. It is widely used as an ornamental, vegetable, and medicinal herb with its rhizome being a popular vegetable. To explore the molecular mechanism underlying its rhizome enlargement, we conducted a systematic analysis on the CONSTANS-LIKE (COL) gene family, with the results, indicating that this gene plays a role in regulating potato tuber expansion. These analyses included phylogenetic relationships, gene structure, and expressional patterns of lotus COL family genes. Based on these analyses, NnCOL5 was selected for further study on its potential function in lotus rhizome formation. NnCOL5 was shown to be located in the nucleus, and its expression was positively associated with the enlargement of lotus rhizome. Besides, the overexpression of NnCOL5 in potato led to increased tuber weight and starch content under short-day conditions without changing the number of tubers. Further analysis suggested that the observed tuber changes might be mediated by affecting the expression of genes in CO-FT and GA signaling pathways. These results provide valuable insight in understanding the functions of COL gene as well as the enlargement of lotus rhizome.

A HY5-COL3-COL13 regulatory chain for controlling hypocotyl elongation in Arabidopsis.

CONSTANS-LIKE (COL) family members are commonly implicated in light signal transduction during early photomorphogenesis. However, some of their functions remain unclear. Here, we propose a role for COL13 in hypocotyl elongation in Arabidopsis thaliana. We found that COL13 RNA accumulates at high levels in hypocotyls and that a disruption in the COL13 function via a T-DNA insertion or RNAi led to the formation of longer hypocotyls of Arabidopsis seedlings under red light. On the contrary, overexpression of COL13 resulted in the formation of shorter hypocotyls. Using various genetic, genomic, and biochemical assays, we proved that another COL protein, COL3, directly binds to the promoter of COL13, and the promoter region of COL3 was targeted by the transcription factor LONG HYPOCOTYL 5 (HY5), to form an HY5-COL3-COL13 regulatory chain for regulating hypocotyl elongation under red light. Additionally, further study demonstrated that COL13 interacts with COL3, and COL13 promotes the interaction between COL3 and CONSTITUTIVE PHOTOMORPHOGENIC1 (COP1), suggesting a possible COP1-dependent COL3-COL13 feedback pathway. Our results provide new information regarding the gene network in mediating hypocotyl elongation.

Identification and characterization of the CONSTANS (CO)/CONSTANS-like (COL) genes related to photoperiodic signaling and flowering in tomato.

CO is an important regulator of photoperiodic response and flowering. However, the biological functions of CO and COL genes in tomato (Solanum lycopersicum) remain elusive. Here we identified 13 members in CO/COL family from the tomato genome. They were divided into three groups, and each group had specific characteristics in gene structures and protein domains. The SlCO/SlCOL genes showed different tissue-specific expression patterns and circadian rhythms, indicating their functional diversity in tomato. Moreover, among 13 members, the expression of SlCOL, SlCOL4a, and SlCOL4b was negatively correlated with flowering time variation in ten tomato lines. Through interaction network prediction, we found three FLOWERING LOCUS T (FT) orthologs, SINGLE FLOWER TRUSS (SFT), FT-like (FTL), and FT-like 1 (FTL1), which functioned as candidate interactors of SlCOL, SlCOL4a, and SlCOL4b. Further expression analyses suggested that SFT coincided with the three SlCOL genes in ten tomato lines with varied flowering time. These findings implied that SlCOL, SlCOL4a, and SlCOL4b are potential flowering inducers in tomato, and SFT may act as their downstream target. Thus, our study built a foundation for understanding the precise roles of SlCO/SlCOL family in plant growth and development of tomato, especially in flowering.

Identification and characterization of the CONSTANS-like gene family and its expression profiling under light treatment in Populus.

The CONSTANS-like (COL) genes play an important role in the photoperiodic flowering pathway. Poplar is a perennial woody plant with a long juvenile phase, but the molecular characterization of COL genes in Populus is limited. In this study, 14 COL genes were identified in the Populus genome. Phylogenetic analysis indicated the PtCOL proteins were divided into three subgroups, and the members of each subgroup had similar gene structure and motif composition. Chromosome distribution analysis showed that 14 PtCOL genes were distributed on 10 chromosomes. Multiple sequence alignment indicated that these proteins contained a highly conserved B-box1 and a conserved CCT domain, but the B-box2 structure was divided into three different types. Promoter analysis found that there were several light-responsive cis-elements in the PtCOL genes. Furthermore, tissue-specific expression showed that all nine PtCOL genes were widely expressed in various tissues and organs of Populus, and were preferentially expressed in the leaves. Additionally, the transcription level of PtCOL exhibited a diurnal oscillation pattern in different light conditions. This study not only provided comprehensive information for further analysis of the function of the PtCOL gene family, but also revealed the biological roles of PtCOL genes in the photoperiod-dependent flowering process of Populus.

The Arabidopsis thaliana CONSTANS-LIKE 4 (COL4) - A Modulator of Flowering Time.

Appropriate control of flowering time is crucial for crop yield and the reproductive success of plants. Flowering can be induced by a number of molecular pathways that respond to internal and external signals. In Arabidopsis, expression of the key florigenic signal FLOWERING LOCUS T (FT) is positively regulated by CONSTANS (CO) a BBX protein sharing high sequence similarity with 16 CO-like proteins. Within this study, we investigated the role of the Arabidopsis CONSTANS-LIKE 4 (COL4), whose role in flowering control was unknown. We demonstrate that, unlike CO, COL4 is a flowering repressor in long days (LD) and short days (SD) and acts on the expression of FT and FT-like genes as well as on SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1). Reduction of COL4 expression level leads to an increase of FT and APETALA 1 (AP1) expression and to accelerated flowering, while the increase of COL4 expression causes a flowering delay. Further, the observed co-localization of COL4 protein and CO in nuclear speckles supports the idea that the two act as an antagonistic pair of transcription factors. This interaction may serve the fine-tuning of flowering time control and other light dependent plant developmental processes.

Overexpression of CONSTANS-like 16 enhances chlorophyll accumulation in petunia corollas.

We have previously found that a gene closely related to Arabidopsis CONSTANS-like 16 (COL16) was coordinately expressed with chlorophyll content in chrysanthemum petals and leaves. Here, to elucidate whether COL16 is involved in the regulation of chlorophyll biosynthesis and accumulation, we analyzed the function of COL16 in petunia (Petunia hybrida). We identified three petunia COL16 homologs: PhCOL16a, PhCOL16b, and PhCOL16c. Expression patterns of all three homologs were associated with chlorophyll content, with lower levels in white corollas than in pale green corollas, and relatively high levels in leaves. The result suggests that PhCOL16 homologs are involved in chlorophyll accumulation. We introduced a PhCOL16a overexpression construct into petunia. The transgenic plants had pale green corollas with a higher chlorophyll content than wild-type plants. Expression of genes encoding key enzymes of chlorophyll biosynthesis was significantly higher in the transgenic plants than in the wild-type plants. The results indicate that PhCOL16 positively regulates chlorophyll biosynthesis.

A member of the CONSTANS-Like protein family is a putative regulator of reactive oxygen species homeostasis and spaceflight physiological adaptation.

A feature of the physiological adaptation to spaceflight in Arabidopsis thaliana (Arabidopsis) is the induction of reactive oxygen species (ROS)-associated gene expression. The patterns of ROS-associated gene expression vary among Arabidopsis ecotypes, and the role of ROS signalling in spaceflight acclimation is unknown. What could differences in ROS gene regulation between ecotypes on orbit reveal about physiological adaptation to novel environments? Analyses of ecotype-dependent responses to spaceflight resulted in the elucidation of a previously uncharacterized gene (OMG1) as being ROS-associated. The OMG1 5' flanking region is an active promoter in cells where ROS activity is commonly observed, such as in pollen tubes, root hairs, and in other tissues upon wounding. qRT-PCR analyses revealed that upon wounding on Earth, OMG1 is an apparent transcriptional regulator of MYB77 and GRX480, which are associated with the ROS pathway. Fluorescence-based ROS assays show that OMG1 affects ROS production. Phylogenetic analysis of OMG1 and closely related homologs suggests that OMG1 is a distant, unrecognized member of the CONSTANS-Like protein family, a member that arose via gene duplication early in the angiosperm lineage and subsequently lost its first DNA-binding B-box1 domain. These data illustrate that members of the rapidly evolving COL protein family play a role in regulating ROS pathway functions, and their differential regulation on orbit suggests a role for ROS signalling in spaceflight physiological adaptation.

Phytochrome A signal transduction 1 and CONSTANS-LIKE 13 coordinately orchestrate shoot branching and flowering in leafy Brassica juncea.

Branching is a major determinant of crop yield, and enables vigorous shoot growth and the production of a dense canopy. Phytochrome A signal transduction 1 (PAT1) positively regulates phytochrome A signal transduction in response to light, but its effects on branching remain unknown. In this study, we mapped PAT1, and revealed a previously unknown role related to branching and flowering in leafy Brassica juncea. Earlier and increased branching was observed when PAT1 expression was down-regulated, implying that PAT1 negatively regulates shoot branching. Additionally, down-regulated PAT1 expression reversed the inhibited branching induced by far-red light, suggesting PAT1 is involved in the shade avoidance response. PAT1 negatively regulated branching only after bud initiation. The observed interaction between PAT1 and BRC1 implied that PAT1 influences bud outgrowth in a BRC1-dependent manner. Biochemical and genetic evidence indicate that PAT1 directly interacts with CONSTANS-LIKE 13 (COL13), which negatively regulates flowering, with the resulting PAT1-COL13 complex mediating shoot branching and flowering. Our findings reveal a new crosstalk modality between phytochrome signalling and flowering pathways during the regulation of shoot branching and flowering. The data presented herein may be useful for future studies involving the editing of the GRAS family transcription factor PAT1 gene to enhance crop productivity and enable earlier harvesting.