Xiaojian Deng

The Histone Methyltransferase SDG724 Mediates H3K36me2/3 Deposition at MADS50 and RFT1 and Promotes Flowering in Rice

In rice, the two florigens Hd3a and RFT1 are separated by only 11.5 kb in the genome. This study reveals that rice SDG724 specifically affects the histone H3 lysine 36 me2/3 level of RFT1 but not its close paralog Hd3a. Therefore, RFT1 and Hd3a have functionally diverged to control flowering time under long-day and short-day conditions partly via a fine-tuned epigenetic mechanism. Chromatin modifications affect flowering time in the long-day plant Arabidopsis thaliana, but the role of histone methylation in flowering time regulation of rice (Oryza sativa), a short-day plant, remains to be elucidated. We identified a late-flowering long vegetative phase1 (lvp1) mutant in rice and used map-based cloning to reveal that lvp1 affects the SET domain group protein 724 (SDG724). SDG724 functions as a histone methyltransferase in vitro and contributes to a major fraction of global histone H3 lysine 36 (H3K36) methylation in vivo. Expression analyses of flowering time genes in wild-type and lvp1 mutants revealed that Early heading date1, but not Heading date1, are misregulated in lvp1 mutants. In addition, the double mutant of lvp1 with photoperiod sensitivity5 (se5) flowered later than the se5 single mutant, indicating that lvp1 delays flowering time irrespective of photoperiod. Chromatin immunoprecipitation assays showed that lvp1 had reduced levels of H3K36me2/3 at MADS50 and RFT1. This suggests that the divergent functions of paralogs RFT1 and Hd3a, and of MADS50 and MADS51, are in part due to differential H3K36me2/3 deposition, which also correlates with higher expression levels of MADS50 and RFT1 in flowering promotion in rice.

Understanding the genetic and epigenetic architecture in complex network of rice flowering pathways

ABSTRACTAlthough the molecular basis of flowering time control is well dissected in the long day (LD) plant Arabidopsis, it is still largely unknown in the short day (SD) plant rice. Rice flowering time (heading date) is an important agronomic trait for season adaption and grain yield, which is affected by both genetic and environmental factors. During the last decade, as the nature of florigen was identified, notable progress has been made on exploration how florigen gene expression is genetically controlled. In Arabidopsis expression of certain key flowering integrators such as FLOWERING LOCUS C (FLC) and FLOWERING LOCUS T (FT) are also epigenetically regulated by various chromatin modifications, however, very little is known in rice on this aspect until very recently. This review summarized the advances of both genetic networks and chromatin modifications in rice flowering time control, attempting to give a complete view of the genetic and epigenetic architecture in complex network of rice flowering pathways.

Identification of a Geranylgeranyl reductase gene for chlorophyll synthesis in rice

Geranylgeranyl reductase (CHL P) catalyzes the reduction of geranylgeranyl diphosphate to phytyl diphosphate, and provides phytol for both Chlorophyll (Chl) and tocopherol synthesis. In this study, we isolated a yellow-green leaf mutant, 502ys, in rice (Oryza sativa). The mutant exhibited reduced level of Chls, arrested development of chloroplasts, and retarded growth rate. The phenotype of the 502ys mutant was controlled by by a recessive mutation in a nuclear gene on the long arm of rice chromosome 2. Map-based cloning of the mutant resulted in the identification of an OsChl P gene (LOC_Os02g51080). In the 502ys mutant, a single base pair mutation was detected at residue 1279 in DNA sequence of the gene, resulting in an amino acid change (Gly-206 to Ser) in the encoded protein. HPLC analysis of Chls indicated that the majority of Chl molecules are conjugated with an unsaturated geranylgeraniol side chain, in addition to small amount of normal Chls in the mutant. Furthermore, the mutant phenotype was complemented by transformation with the wild-type gene. Therefore, this study has confirmed the 502ys mutant resulted from a single base pair mutation in OsChl P gene.

OsPOP5, a prolyl oligopeptidase family gene from rice confers abiotic stress tolerance in Escherichia coli.

The prolyl oligopeptidase family, which is a group of serine peptidases, can hydrolyze peptides smaller than 30 residues. The prolyl oligopeptidase family in plants includes four members, which are prolyl oligopeptidase (POP, EC3.4.21.26), dipeptidyl peptidase IV (DPPIV, EC3.4.14.5), oligopeptidase B (OPB, EC3.4.21.83), and acylaminoacyl peptidase (ACPH, EC3.4.19.1). POP is found in human and rat, and plays important roles in multiple biological processes, such as protein secretion, maturation and degradation of peptide hormones, and neuropathies, signal transduction and memory and learning. However, the function of POP is unclear in plants. In order to study POP function in plants, we cloned the cDNA of the OsPOP5 gene from rice by nested-PCR. Sequence analysis showed that the cDNA encodes a protein of 596 amino acid residues with Mw ≈ 67.29 kD. In order to analyze the protein function under different abiotic stresses, OsPOP5 was expressed in Escherichia coli. OsPOP5 protein enhanced the tolerance of E. coli to high salinity, high temperature and simulated drought. The results indicate that OsPOP5 is a stress-related gene in rice and it may play an important role in plant tolerance to abiotic stress.

Map-based cloning and characterization of the novel yellow-green leaf gene ys83 in rice (Oryza sativa)

Leaf-color mutants have been extensively studied in rice, and many corresponding genes have been identified up to now. However, leaf-color mutation mechanisms are diverse and still need further research through identification of novel genes. In the present paper, we isolated a leaf-color mutant, ys83, in rice (Oryza sativa). The mutant displayed a yellow-green leaf phenotype at seedling stage, and then slowly turned into light-green leaf from late tillering stage. In its yellow leaves, photosynthetic pigment contents significantly decreased and the chloroplast development was retarded. The mutant phenotype was controlled by a recessive mutation in a nuclear gene on the short arm of rice chromosome 2. Map-based cloning and sequencing analysis suggested that the candidate gene was YS83 (LOC_Os02g05890) encoding a protein containing 165 amino acid residues. Gene YS83 was expressed in a wide range of tissues, and its encoded protein was targeted to the chloroplast. In the mutant, a T-to-A substitution occurred in coding sequence of gene YS83, which caused a premature translation of its encoded product. By introduction of the wild-type gene, the ys83 mutant recovered to normal green-leaf phenotype. Taken together, we successfully identified a novel yellow-green leaf gene YS83. In addition, number of productive panicles per plant and number of spikelets per panicle only reduced by 6.7% and 7.6%, respectively, meanwhile its seed setting rate and 1000-grain weight (seed size) were not significantly affected in the mutant, so leaf-color mutant gene ys83 could be used as a trait marker gene in commercial hybrid rice production.

Mutation of FdC2 gene encoding a ferredoxin-like protein with C-terminal extension causes yellow-green leaf phenotype in rice.

Ferredoxins (Fds) are small iron-sulfur proteins that mediate electron transfer in a wide range of metabolic reactions. Besides Fds, there is a type of Fd-like proteins designated as FdC, which have conserved elements of Fds, but contain a significant C-terminal extension. So far, only two FdC genes of Arabidopsis (Arabidopsis thaliana) have been identified in higher plants and thus the functions of FdC proteins remain largely unknown. In this study, we isolated a yellow-green leaf mutant, 501ys, in rice (Oryza sativa). The mutant exhibited yellow-green leaf phenotype and reduced chlorophyll level. The phenotype of 501ys was caused by mutation of a gene on rice chromosome 3. Map-based cloning of this mutant resulted in identification of OsFdC2 gene (LOC_Os03g48040) showing high identity with Arabidopsis FdC2 gene (AT1G32550). OsFdC2 was expressed most abundantly in leaves and its encoded protein was targeted to the chloroplast. In 501ys mutant, a missense mutation was detected in DNA sequence of the gene, resulting in an amino acid change in the encoded protein. The mutant phenotype was rescued by introduction of the wild-type gene. Therefore, we successfully identified FdC2 gene via map-based cloning approach, and demonstrated that mutation of this gene caused yellow-green leaf phenotype in rice.

GRY79 encoding a putative metallo-β-lactamase-trihelix chimera is involved in chloroplast development at early seedling stage of rice.

Key messageThegreen-revertible yellow79mutant resulting from a single-base mutation suggested that theGRY79gene encoding a putative metallo-β-lactamase-trihelix chimera is involved in chloroplast development at early seedling stage of rice.AbstractFunctional studies of metallo-β-lactamases and trihelix transcription factors in higher plants remain very sparse. In this study, we isolated the green-revertible yellow79 (gry79) mutant in rice. The mutant developed yellow-green leaves before the three-leaf stage but recovered to normal green at the sixth-leaf stage. Meanwhile, the mutant exhibited reduced level of chlorophylls and arrested development of chloroplasts in the yellow leaves. Genetic analysis suggested that the mutant phenotype was controlled by a single recessive nuclear gene on rice chromosome 2. Map-based cloning revealed that the candidate gene was Os02g33610 encoding a putative metallo-β-lactamase-trihelix chimera. In the gry79 mutant, a single-base mutation occurred in coding region of the gene, resulting in an amino acid change in the encoded protein. Furthermore, the mutant phenotype was rescued by transformation with the wild-type gene. Therefore, we have confirmed that the gry79 mutant phenotype resulted from a single-base mutation in GRY79 (Os02g33610) gene, suggesting that the gene encoding a putative metallo-β-lactamase-trihelix chimera is involved in chloroplast development at early seedling stage of rice. In addition, we considered that the gry79 mutant gene could be applicable as a leaf-color marker gene for efficient identification and elimination of false hybrids in commercial hybrid rice production.

Fine mapping and candidate gene analysis of the dwarf gene d162(t) in rice (Oryza sativa L.)

In our previous study, d162(t), a single recessive gene, which caused rice dwarf mutant, had been mapped on the short arm of chromosome 3. In this study, the d162(t) gene was fine mapped to a confined region about 0.82 cM by RM14641 and RM3134, and co-segregated with InDel361-2, InDel361-3, InDel361-5, RM14645, RM1022 and RM14643, where no known gene involved in plant height has been identified. Based on the annotation results of TIGR, dozens of open reading frames (ORFs) were predicted in this region, among them, five ORFs were the most possible genes related to the phenotype. In these ORFs, Os03g13010, related to U-box domain containing protein, had a 62bp segment deletion in the coding region in 162d (mutant type, MT). The results of RT-PCR showed that the transcriptional level of Os03g13010 was significantly different between Shuhui162 (wild type, WT) and 162d (MT). Therefore, the gene (Os03g13010) encoding a U-box domain containing protein was considered as the candidate gene of d162(t).

Genetic Analysis and Fine Mapping of Gene ygl98 for Yellow-Green Leaf of Rice

Abstract A yellow-green leaf mutant ygl98 was isolated by ethyl methane sulfonate (EMS) mutagenesis. The whole plant exhibited yellow-green character throughout the growing period. The contents of chlorophyll and carotenoid decreased by 45.3% and 45.6%, respectively, compared with its wild-type parent 10079. At maturity stage, the number of productive panicles per plant, seed-setting rate, and plant height of the mutant were reduced by 14.4%, 10.7%, and 7.4%, respectively. Under electron microscope, the chloroplasts in the ygl98 mutant were out-of-shape. A lot of cystic structures and poor thylakoids were observed in the chloroplasts of the ygl98 mutant, and grana stacks appeared to be less dense compared to those of the wild type. Genetic analysis showed that the yellow-green leaf character of the ygl98 mutant was controlled by one pair of recessive nuclear genes. Genetic mapping of the mutant gene was conducted using 771 yellow-green leaf individuals from the F 2 mapping population of ygl98 /Zhefu 802. Finally, the mutant gene was mapped between insert/deletion (InDel) markers I3 and I4 on the long arm of chromosome 3 with the genetic distances of 0.07 cM and 0.19 cM, respectively. The physical distance between the 2 markers was 44.2 kb harboring 8 predicted genes annotated. Sequencing analysis of these candidate genes between the mutant and its wild type revealed that the single base change (G1 522A) of the gene for magnesium-chelatase ChlD subunit resulted in a missense mutation (A508T) in the encoded product. The same gene mutation caused by OsChlD ( Chlorina-1 ) was documented previously. The Chlorina-1 mutant displays a severe yellowish-green leaf phenotype only at seedling stage, and the abnormal leaf color is first observed on the leaves of 2- to 3-week-old seedlings, while the ygl98 mutant exhibits yellow-green character throughout the growing period. The different phenotypes of the 2 mutants may be caused by the different mutational sites of OsChlD genomic sequence.

A single nucleotide mutation of IspF gene involved in the MEP pathway for isoprenoid biosynthesis causes yellow-green leaf phenotype in rice

Key messageWe identified IspF gene through yellow-green leaf mutant 505ys in rice. OsIspF was expressed in all tissues detected, and its encoded protein was targeted to the chloroplast. On expression levels of genes in this mutant, OsIspF itself and the genes encoding other enzymes of the MEP pathway and chlorophyll synthase were all up-regulated, however, among eight genes associated with photosynthesis, only psaA, psaN and psbA genes for three reaction center subunits of photosystem obviously changed.AbstractIsoprenoids are the most abundant natural compounds in all organisms, which originate from the basic five-carbon units isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP). In plants, IPP and DMAPP are synthesized through two independent pathways, the mevalonic acid pathway in cytoplasm and the 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway in plastids. The MEP pathway comprises seven enzymatic steps, in which IspF is the fifth enzyme. So far, no IspF gene has been identified in monocotyledonous plants. In this study, we isolated a leaf-color mutant, 505ys, in rice (Oryza sativa). The mutant displayed yellow-green leaf phenotype, reduced level of photosynthetic pigments, and arrested development of chloroplasts. By map-based cloning of this mutant, we identified OsIspF gene (LOC_Os02g45660) showing significant similarity to IspF gene of Arabidopsis, in which a missense mutation occurred in the mutant, resulting in an amino acid change in the encoded protein. OsIspF gene was expressed in all tissues detected, and its encoded protein was targeted to the chloroplast. Further, the mutant phenotype of 505ys was complemented by transformation with the wild-type OsIspF gene. Therefore, we successfully identified an IspF gene in monocotyledonous plants. In addition, real-time quantitative RT-PCR implied that a positive regulation could exist between the OsIspF gene and the genes encoding other enzymes of the MEP pathway and chlorophyll synthase. At the same time, it also implied that the individual genes involved in the MEP pathway might differentially regulated expression levels of the genes associated with photosynthesis.