Identification of the Regulatory Region Responsible for Vascular Tissue-Specific Expression in the Rice Hd3a Promoter

Pasriga, Richa,Cho, Lae-Hyeon,Yoon, Jinmi,An, Gynheung Korean Society for Molecular and Cellular Biology 2018 Molecules and cells Vol.41 No.4

Flowering time is determined by florigens. These genes include, Heading date 3a (Hd3a) and Rice FT 1 (RFT1) in rice, which are specifically expressed in the vascular tissues of leaves at the floral transition stage. To study the cis-regulatory elements present in the promoter region of Hd3a, we generated transgenic plants carrying the 1.75-kb promoter fragment of Hd3a that was fused to the ${\beta}$-glucuronidase (GUS) reporter gene. Plants expressing this construct conferred a vascular cell-specific expression pattern for the reporter gene. However, GUS was expressed in leaves at all developmental stages, including the early seedling stage when Hd3a was not detected. Furthermore, the reporter was expressed in roots at all stages. This suggests that the 1.75-kb region lackings cis-elements that regulate leaf-specific expression at the appropriate developmental stages. Deletion analyses of the promoter region indicated that regulatory elements determining vascular cell-specific expression are present in the 200-bp region between -245 bp and -45 bp from the transcription initiation site. By transforming the Hd3a-GUS construct to rice cultivar 'Taichung 65' which is defective in Ehd1, we observed that Ehd1 is the major regulatory element that controls Hd3a promoter activity.

Overexpression of RICE FLOWERING LOCUS T 1 (RFT1) Induces Extremely Early Flowering in Rice

Pasriga, Richa,Yoon, Jinmi,Cho, Lae-Hyeon,An, Gynheung Korean Society for Molecular and Cellular Biology 2019 Molecules and cells Vol.42 No.5

RICE FLOWERING LOCUS T 1 (RFT1) is a major florigen that functions to induce reproductive development in the shoot apical meristem (SAM). To further our study of RFT1, we overexpressed the gene and examined the expression patterns of major regulatory genes during floral transition and inflorescence development. Overexpression induced extremely early flowering in the transgenics, and a majority of those calli directly formed spikelets with a few spikelets, thus bypassing normal vegetative development. FRUITFULL (FUL)-clade genes OsMADS14, OsMADS15, and OsMADS18 were highly induced in the RFT1-expressing meristems. OsMADS34 was also induced in the meristems. This indicated that RFT1 promotes the expression of major regulatory genes that are important for inflorescence development. RFT1 overexpression also induced SEPALLATA (SEP)-clade genes OsMADS1, OsMADS5, and OsMADS7 in the greening calli before floral transition occurred. This suggested their possible roles at the early reproductive stages. We found it interesting that expression of OsFD1 as well as OsFD2 and OsFD3 was strongly increased in the RFT1-expressing calli and spikelets. At a low frequency, those calli produced plants with a few leaves that generated a panicle with a small number of spikelets. In the transgenic leaves, the FUL-clade genes and OsMADS34 were induced, but SEP-clade gene expression was not increased. This indicated that OsMADS14, OsMADS15, OsMADS18, and OsMADS34 act immediately downstream of RFT1.

Overexpression of RICE FLOWERING LOCUS T 1 (RFT1) Induces Extremely Early Flowering in Rice

Richa Pasriga,윤진미,조래현,안균흥 한국분자세포생물학회 2019 Molecules and cells Vol.42 No.5

RICE FLOWERING LOCUS T 1 (RFT1) is a major florigen that functions to induce reproductive development in the shoot apical meristem (SAM). To further our study of RFT1, we overexpressed the gene and examined the expression patterns of major regulatory genes during floral transition and inflorescence development. Overexpression induced extremely early flowering in the transgenics, and a majority of those calli directly formed spikelets with a few spikelets, thus bypassing normal vegetative development. FRUITFULL (FUL)-clade genes OsMADS14, OsMADS15, and OsMADS18 were highly induced in the RFT1-expressing meristems. Os-MADS34 was also induced in the meristems. This indicated that RFT1 promotes the expression of major regulatory genes that are important for inflorescence development. RFT1 overexpression also induced SEPALLATA (SEP)-clade genes OsMADS1, OsMADS5, and OsMADS7 in the greening calli before floral transition occurred. This suggested their possible roles at the early reproductive stages. We found it interesting that expression of OsFD1 as well as OsFD2 and OsFD3 was strongly increased in the RFT1-expressing calli and spikelets. At a low frequency, those calli produced plants with a few leaves that generated a panicle with a small number of spikelets. In the transgenic leaves, the FULclade genes and OsMADS34 were induced, but SEP-clade gene expression was not increased. This indicated that Os-MADS14, OsMADS15, OsMADS18, and OsMADS34 act immediately downstream of RFT1.

Identification of the Regulatory Region Responsible for Vascular Tissue–Specific Expression in the Rice Hd3a Promoter

Richa Pasriga,조래현,윤진미,안진흥 한국분자세포생물학회 2018 Molecules and cells Vol.41 No.4

Flowering time is determined by florigens. These genes include, Heading date 3a (Hd3a) and Rice FT 1 (RFT1) in rice, which are specifically expressed in the vascular tissues of leaves at the floral transition stage. To study the cis-regulatory elements present in the promoter region of Hd3a, we generated transgenic plants carrying the 1.75-kb promoter fragment of Hd3a that was fused to the β-glucuronidase (GUS) reporter gene. Plants expressing this construct conferred a vascular cell-specific expression pat-tern for the reporter gene. However, GUS was expressed in leaves at all developmental stages, including the early seedling stage when Hd3a was not detected. Furthermore, the reporter was expressed in roots at all stages. This suggests that the 1.75-kb region lackings cis-elements that regulate leaf-specific expression at the appropriate developmental stages. Deletion analyses of the promoter region indicated that regulatory elements determining vascular cell-specific expression are present in the 200-bp region between -245 bp and -45 bp from the transcription initiation site. By transforming the Hd3a-GUS construct to rice cultivar ‘Taichung 65’ which is defective in Ehd1, we observed that Ehd1 is the major regulatory element that controls Hd3a promoter activity.

Roles of Sugars in Controlling Flowering Time

Gynheung An,전종성,조래현,Richa Pasriga,Jinmi Yoon 한국식물학회 2018 Journal of Plant Biology Vol.61 No.3

Flowering time is influenced by environmentalfactors such as photosynthesis, temperature, nutrition, andwater. The main products of photosynthesis are sugars thatare mobilized to sink tissues to support plant growth anddifferentiation. They also function as signals to controlvarious types of metabolism and developmental processes. One of the most important transitions in the plant life cycleis from the vegetative to reproductive phase. During thattransition, sucrose levels rise rapidly but transiently in thephloem and shoot apexes. For several species, the addition ofexogenous sucrose promotes flowering, possibly by acting asa main signal. Although other sugars, including glucose, alsoappear to be involved in this transition, evidence for theirroles in flowering is limited. In Arabidopsis thaliana,trehalose-6-phosphate serves as a signal to induce flowering. However, its roles in other plants have not been reported. Sucrose seems to function primarily in the leaf phloem toenhance the generation of florigens such as Flowering LocusT (FT) while trehalose-6-phosphate functions in the shootapical meristem to promote the flowering signal pathwaydownstream of those florigens.

Homodimerization of Ehd1 Is Required to Induce Flowering in Rice

Cho, Lae-Hyeon,Yoon, Jinmi,Pasriga, Richa,An, Gynheung American Society of Plant Biologists 2016 Plant Physiology Vol.170 No.4

<P>In plants, flowering time is elaborately controlled by various environment factors. Ultimately, florigens such as FLOWERING LOCUS T (FT) or FT-like molecules induce flowering. In rice (Oryza sativa), Early heading date 1 (Ehd1) is a major inducer of florigen gene expression. Although Ehd1 is highly homologous to the type-B response regulator (RR) family in the cytokinin signaling pathway, its precise molecular mechanism is not well understood. In this study, we showed that the C-terminal portion of the protein containing the GARP DNA-binding (G) domain can promote flowering when overexpressed. We also observed that the N-terminal portion of Ehd1, carrying the receiver (R) domain, delays flowering by inhibiting endogenous Ehd1 activity. Ehd1 protein forms a homomer via a 16-amino acid region in the inter domain between R and G. From the site-directed mutagenesis analyses, we demonstrated that phosphorylation of the Asp-63 residue within the R domain induces the homomerization of Ehd1, which is crucial for Ehd1 activity. A type-A RR, OsRR1, physically interacts with Ehd1 to form a heterodimer. In addition, OsRR1-overexpressing plants show a late-flowering phenotype. Based on these observations, we conclude that OsRR1 inhibits Ehd1 activity by binding to form an inactive complex.</P>

Chromatin Interacting Factor OsVIL2 Is Required for Outgrowth of Axillary Buds in Rice

Yoon, Jinmi,Cho, Lae-Hyeon,Lee, Sichul,Pasriga, Richa,Tun, Win,Yang, Jungil,Yoon, Hyeryung,Jeong, Hee Joong,Jeon, Jong-Seong,An, Gynheung Korean Society for Molecular and Cellular Biology 2019 Molecules and cells Vol.42 No.12

Shoot branching is an essential agronomic trait that impacts on plant architecture and yield. Shoot branching is determined by two independent steps: axillary meristem formation and axillary bud outgrowth. Although several genes and regulatory mechanism have been studied with respect to shoot branching, the roles of chromatin-remodeling factors in the developmental process have not been reported in rice. We previously identified a chromatin-remodeling factor OsVIL2 that controls the trimethylation of histone H3 lysine 27 (H3K27me3) at target genes. In this study, we report that loss-of-function mutants in OsVIL2 showed a phenotype of reduced tiller number in rice. The reduction was due to a defect in axillary bud (tiller) outgrowth rather than axillary meristem initiation. Analysis of the expression patterns of the tiller-related genes revealed that expression of OsTB1, which is a negative regulator of bud outgrowth, was increased in osvil2 mutants. Chromatin immunoprecipitation assays showed that OsVIL2 binds to the promoter region of OsTB1 chromatin in wild-type rice, but the binding was not observed in osvil2 mutants. Tiller number of double mutant osvil2 ostb1 was similar to that of ostb1, suggesting that osvil2 is epistatic to ostb1. These observations indicate that OsVIL2 suppresses OsTB1 expression by chromatin modification, thereby inducing bud outgrowth.

Chromatin Interacting Factor OsVIL2 Is Required for Outgrowth of Axillary Buds in Rice

Gynheung An,Jinmi Yoon,조래현,이시철,Richa Pasriga,윈툰,Jungil Yang,Hyeryung Yoon,Hee Joong Jeong,전종성 한국분자세포생물학회 2019 Molecules and cells Vol.42 No.12

Shoot branching is an essential agronomic trait that impacts on plant architecture and yield. Shoot branching is determined by two independent steps: axillary meristem formation and axillary bud outgrowth. Although several genes and regulatory mechanism have been studied with respect to shoot branching, the roles of chromatin-remodeling factors in the developmental process have not been reported in rice. We previously identified a chromatin-remodeling factor OsVIL2 that controls the trimethylation of histone H3 lysine 27 (H3K27me3) at target genes. In this study, we report that loss-of-function mutants in OsVIL2 showed a phenotype of reduced tiller number in rice. The reduction was due to a defect in axillary bud (tiller) outgrowth rather than axillary meristem initiation. Analysis of the expression patterns of the tiller-related genes revealed that expression of OsTB1, which is a negative regulator of bud outgrowth, was increased in osvil2 mutants. Chromatin immunoprecipitation assays showed that OsVIL2 binds to the promoter region of OsTB1 chromatin in wild-type rice, but the binding was not observed in osvil2 mutants. Tiller number of double mutant osvil2 ostb1 was similar to that of ostb1, suggesting that osvil2 is epistatic to ostb1. These observations indicate that OsVIL2 suppresses OsTB1 expression by chromatin modification, thereby inducing bud outgrowth.