Os06g0651100

The gene YK1(HC-toxin reductase like rice gene)is associated with metabolic alteration of rice.

Annotated Information

Function

• Alteration of metabolites as well as proteins may contribute to multiple stress tolerances in transgenic YK1 rice. Ectopic over-expression of a single gene (YK1) in rice cells might affect the expression of unrelated proteins and metabolites.

• The overexpression of YK1 in calli enhanced synthesis and the reduction state of GSH concomitantly with NAD(P)(H) production. Since the ratio of GSH to GSSG increased, YK1 calli might enhance tolerance to oxidative stresses. On the other hand, the amounts of GSH in YK1 plants were only marginally increased and the activities of GPX and GR were not altered.

• The YK1-overexpressing rice plants showed enhanced resistance to rice blast disease and to abiotic stresses like UV radiation, increased salinity, and submergence. YK1 possesses the catalytic activity to convert taxifolin to leucocyanidin.

• The overexpression of YK1 (DFR) was associated with slight changes in the amounts of several metabolites analysed in whole plants, whilst glutathione derivatives were substantially increased in suspensioncultured cells. Transgenic rice plants harboring the YK1 gene showed high resistance against blast infection. The combination of dihydroflavonol-4-reductase activity and the elevated level of NAD(P)H pool may confer the prevention of induced cell death in planta.

GO assignment(s): GO:0003824, GO:0009225, GO:0051287

Mutation

YK1-transformed plants:

L-1

L-2

• The amount of GSH was increased in YK1 lines by 291% for L-1 and 204% for L-2 over the control. Likewise, the amount of GSSG was increased by 131% (L-1) and 134% (L-2), respectively.

• The total amounts of NAD(H) and NADP(H) in 5-day-old YK1 calli (L-1 and L-2) were higher than the control. In the case of NAD synthetase and NAD kinase, both L-1 and L-2 callus lines showed higher activities than the control.

• ADH, GAPDH, and ME activity were enhanced in YK1-overexpressing cultured cells (L-1). L-1 and L-2 transgenic cells showed apparent H₂O₂ stress tolerance (Fig. 4A). Moreover, an ion leakage assay using excised leaves also demonstrated that both L-1 and L-2 lines leaked fewer ions than the control.

• Both L-1 and L-2 lines exhibited relatively reduced disease severity compared with the control plants where actively spreading lesions were developed on leaves of the control plants, whereas unexpanded lesions were formed on the YK1 lines.

• Although transgenic cells over-expressing YK1 (L-1 and L-2) showed similar production of H₂O₂, the level was significantly lower than the control cells after equilibration for at least 3h.

Expression

• Ectopic overexpression of YK1 protein can be used for the molecular breeding of biotic- and abiotic-resistant rice breeding. YK1 is not involved in the anti-oxidant activities, but may inhibit some ROS generating enzymes.

• The overexpression of YK1 in both suspension-cultured cells and rice plants increased NAD(H) and NADP(H) levels by causing an increase in NAD synthetase and NAD kinase activities.

• The overexpression of YK1 in rice altered the activities of enzymes engaged in NAD biosynthesis, and of enzymes requiring NAD(P) as coenzymes. Moreover, overexpression of YK1 in rice conferred the prevention of cell death caused by the hydrogen peroxide as well as the bacterial disease.

• In YK1 (DFR)-overexpressing plants, the concentrations of cis-aconitate, isocitrate and 2-oxoglutarate were higher in leaves, whereas those of fructose-1,6-bisphosphate and glyceraldehyde-3-phosphate were lower in roots. In seeds, the amounts of free amino acids and metals were altered, whereas sugars in seeds were kept constant. In YK1 calli, an approx. 3-fold increase in glutathione was observed, whereas the activities of glutathione peroxidase and glutathione reductase were concomitantly increased.

• The YK1 gene expression has less or no effects for tolerance to infection with the bacterial blight. Overexpression of YK1 conferred resistance phenotype to the bacterial brown stripe.

Figure 1. Diagram illustrating the biochemical functions of YK1 protein in relation to flavonoid and NAD homeostasis in the metabolic pathways.(from reference ).

• The overexpression of a single gene can cause metabolic alterations by changing the activities of other enzymes related to NAD homeostasis. It may be presumed that overexpressed YK1 protein may use more NAD(P)H than normal level, which in turn pumps up nicotinamide coenzymes by activating responsible enzymes. Such concomitant relation leading to cell-death tolerance is presented in Fig. 1. In addition, high-throughput analysis of metabolites by Fourier transform ion cyclotron resonance MS in transgenic YK1 plants suggested alteration in other metabolite levels (27).

Evolution

The deduced 363 amino acid sequences shared 78 percent homology with maize HC-toxin reductase (Figure 1). The α β α-dinucleotide binding consensus sequence in YK1 gene is also identical at the N-terminal side to its homologues. Such homology can be seen with dehydroflavonol-4-reductase (DHFR), and NADPH dependent carbonyl reductase.

Knowledge Extension

Metabolome analysis in plants has been performed using different types of mass spectrometers. Non-target metabolome analysis of strawberry using a Fourier transform ion cyclotron mass spectrometer (FT-MS) allowed the detection of thousands of metabolites. Mungur et al. showed alterations of metabolites such as amino acids, sugars, lipids, nucleic acids and other organic acids in transgenic tobacco plants exhibiting higher glutamate dehydrogenase activity. Furthermore, fingerprints obtained from FT-MS analysis provided a tremendous amount of information by matching gene expression profiles.

Additionally, CE/MS revealed that the amount of metabolites engaged in the tricarboxylic acid (TCA) cycle was altered in transgenic rice expressing the antisense methylmalonate-semialdehyde dehydrogenase gene.

Labs working on this gene

• Institute of Molecular and Cellular Biosciences,The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan
• Department of Molecular Genetics, National Institute of Agrobiological Resources, 2-1-9 Kannondai, Tsukuba, Ibaraki 305-8601,Japan
• Plant Molecular Biology Group, Laboratory of Environmental Science, Central Research Institute Electric Power Industry, 1646 Abiko, Chiba 270-1194, Japan
• Institute for Advanced Biosciences, University of Keio, Tsuruoka, Yamagata 997-0017, Japan
• Iwate Biotechnology Research Center, Kitakami, Iwate 024-0003, Japan
• Shiraoka Research Station of Biological Science, Nissan Chemical Industries, LTD., 1470, Shiraoka, Minamisaitama, Saitama, 349-0294, Japan
• Department of Applied Biological Science, Science University of Tokyo, Noda, Chiba, 278-8510, Japan;
• Biological Science, Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8601, Japan