Abstract
To assess the effect of the ectopic accumulation of 5-aminolevulinic acid (ALA), a gene encoding a Bradyrhizobium japonicum 5-aminolevulinic acid synthase (ALA-S) was introduced into the genome of rice under the control of the maize ubiquitin promoter. All transgenic rice lines were able to transcribe the B. japonicum ALA-S mRNA, with a higher level in transgenic lines P13 and P14, whereas no ALA-S mRNA was detected in wild-type. The B. japonicum ALA-S protein was expressed efficiently in transgenic plants and localized in the stroma. The detection of ALA-S activity in two transgenic lines demonstrates that the ALA-S gene produces an enzyme that is functionally active in the plants. The introduction of ALA-S genes into transgenic plants led to increases in ALA synthesis of 44% and 85% in transgenic lines P5 and P14, respectively, as compared to wild-type. Protoporphyrin IX (Proto IX) also greatly accumulated in transgenic plants, with up to a 2.3-fold increase in transgenic line P14 as compared to wild-type. The transgenic plants had 9–15% higher levels of chlorophylls (Chls), as compared to wild-type plants, indicating that part of the additional ALA from B. japonicum ALA-S was metabolized to Chls. When exposed to photosynthetic photon flux density (PPFD) of 350 μmol m −2 s −1, growth was retarded and pigment was bleached in all transgenic lines, especially in lines P13 and P14; wild-type plants had no noticeable differences in growth when compared to plants under the growth condition at PPFD of 150 μmol m −2 s −1. The expression of ALA-S in transgenic plants also caused a significant drop in the F v/ F m ratio and an increase in malonyldialdehyde (MDA) level, with a greater change occurring in line P14. The transgenic rice plants expressing B. japonicum ALA-S appeared to suffer from photodynamic damage, due to the accumulation of free Proto IX and Chls, caused by the elevated synthesis of ALA.
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