Os01g0757200

Gibberellin 2-oxidases (GA2oxs) regulate plant growth by inactivating endogenous bioactive gibberellins (GAs).

Annotated Information

Function

The members of the rice (Oryza sativa) GA2ox family are differentially regulated and act in concert or individually to control GA levels during flowering, tillering, and seed germination.
The three activation-tagged mutants, GA2ox5D335-341ACT,GA2ox6ACT, and GA2ox9ACT, were further characterized. Progenies displayed the same phenotypes as their parents, with GA2ox5D335-341ACT and GA2ox9ACT growing slightly shorter than the wild type, while GA2ox6ACT remained severely dwarfed throughout all growth stages (Figures 6A and 6B). GA2ox5D335-341ACT and GA2ox9ACT displayed a normal height but had longer roots and higher tiller numbers than the wild type (Table 1). Other traits significantly altered in the severe dwarf GA2ox6ACT mutant included shorter leaves, later heading date, reduced panicle length, higher tiller numbers, lower grain weight, and lower seed fertility compared with the wild type (Table 1). Germination of GA2ox6ACT seeds was also significantly delayed, as it took 20 d to reach 90% germination rate, while the wild-type and GA2ox9ACT mutant seeds took only 2 d to reach a germination rate of 97 and 98%, respectively (Figure 6C). Germination of GA2ox5D335-341ACT seeds was delayed for 4 d to reach a final 88% germination rate (Figure 6C).

To examine whether rice GA2oxs are functional in dicots,Ubi:GA2ox5 and Ubi:GA2ox6 constructs were used for tobacco transformation. Transgenic tobacco showed the same retardation of plant growth but to different extents. While Ubi:GA2ox5 reduced plant height to 32% and seed production to 62% and Ubi:GA2ox6 reduced plant height to 67% of the wild-type tobacco, Ubi:GA2ox6 had no effect on seed production (Figures 7C and 7D, Table 2). The flowering time was delayed ;2 to 4 weeks for all transgenic tobacco. Growth of hypocotyls and roots of 18-d-old T1 transgenic tobacco seedlings was slightly retarded by overexpression of GA2ox6 but significantly retarded by overexpression of GA2ox5, compared with the wild type(Figures 7E and 7F, Table 2). These studies demonstrated that the two rice GA2oxs have similar functions in monocots and dicots, with GA2ox5 being more potent in inactivation of GA than GA2ox6 in both transgenic rice and tobacco.

'Figure 6. Overexpression of GA2oxs Has Different Effects on Rice Seed Germination and Seedling Growth.(A) Morphology of T1 seedlings at 18 DAI.(B) Seedling heights of GA2ox5D335-341ACT and GA2ox9ACT mutants were slightly shorter, while seedlings of GA2ox6ACT were much shorter than the wild type. Heights of eight plants in each line were measured,and error bars indicate the SE of the mean at each time point.(C) Germination rate was normal for the GA2ox9ACT mutant, slightly delayed for the GA2ox5D335-341ACT mutant, and significantly delayed for the GA2ox6ACT mutant compared with the wild type. Numbers of seeds for determining germination rates were 154, 50, 156, and 49 for the wild type, GA2ox5D335-341ACT, GA2ox6ACT, and GA2ox9ACT, respectively'

'Figure 7. Overexpression of GA2ox5 in Transgenic Rice and Tobacco Causes More Severe Dwarfism Than Overexpression of GA2ox6.(A) and (B) Rice transformed with Ubi:GA2ox5 and Ubi:GA2ox6.(C)to (F) Tobacco transformed with Ubi:GA2ox5 and Ubi:GA2ox6.Transgenic plants showed different degrees of dwarfism compared with the control rice or tobacco transformed with vector only (MS).Photographs of transgenic tobacco were taken at the heading stage ([C]and [D]) and 18 d ([E] and [F]) after sowing of seeds.'

Expression

'Figure 3. Differential Expression of Two Groups of GA2oxs Regulates Flower and Tiller Development.(A) Developmental phases during the life cycle of rice. The timeline is measured in days after imbibition (DAI).(B) Temporal expression patterns of GA2oxs in rice. The last fully expanded leaves were collected from rice plants at different developmental stages. Total RNA was isolated and analyzed by RT-PCR using GA2ox and GA3ox2 gene-specific primers (see Supplemental Table 4 online). The 18S rRNA gene (rRNA) was used as a control.(C) Tiller development during the life cycle of rice. A total of eight plants were used for counting tiller number, and error bars indicate the SE of the mean at each time point.

Genes GA2ox1 to GA2ox9 were differentially expressed in leaves, and their expression was also temporally regulated (Figure 3B). However, mRNAs of GA2ox10 were not detected in any tissue at any growth stage, indicating that GA2ox10 could be a pseudogene or its mRNA level was too low to be detected Based on temporal mRNA accumulation patterns, GA2oxs could be classified into two groups. As can be seen in Figure 3B, for one group excluding GA2ox2 and GA2ox6, accumulation of their mRNAs in leaves was detected prior to the transition from vegetative to reproductive growth phases. By contrast, for another group including GA2ox2 and GA2ox6, their mRNAs accumulated in leaves after the phase transition from vegetative to reproductive growth. GA2ox6 mRNA could also be detected in leaves at early seedling stage and transiently at high level during the active tillering stage. Since expression of most GA2oxs terminated after the active tillering stage, the pattern of tiller growth throughout the rice life cycle was examined. Tiller number increased from 30 to 50 DAI (active tillering), remained constant until 75 DAI, and then increased again until 90 DAI (late tillering)when the experiment was terminated (Figure 3C). Expression of each group of GA2oxs paralleled the active and late tillering stages (cf. Figures 3B with 3C). Except for a slight reduction in the reproductive phase, the expression of GA3ox2, which encodes a GA3ox involved in GA biosynthesis, was not significantly altered in leaves throughout the rice life cycle.

Evolution

'Figure 2. Phylogenetic Tree Based on the Comparison of Plant GA2oxs.Amino acid sequences of 29 GA2oxs from nine plant species (see Supplemental Table 3 online). Plant species: At, Arabidopsis thaliana;Cm, Cucurbita maxima; Ls, Lactuca sativa; Nt, Nicotiana sylvestris; Pc,Phaseolus coccineus; PaPt, Populus alba 3 P. tremuloides; Ps, Pisum sativum; So, Spinacia oleracea. The scale value of 0.1 indicates 0.1 amino acid substitutions per site'

Labs working on this gene

• Institute of Molecular Biology, National Chung-Hsing University, Taichung 402, Taiwan, Republic of China
• Institute of Molecular Biology, Academia Sinica, Taipei 115, Taiwan, Republic of China
• Institute of Plant and Microbial Biology, Academia Sinica, Taipei 115, Taiwan, Republic of China
• Department of Energy Plant Research Laboratory and Department of Plant Biology, Michigan State University,East Lansing, Michigan 48824-1312

References

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