Poster: Oxidative Stress

Abs # 135: Post-Transcriptional Regulation of Ascorbate Peroxidase during Light Adaptation of Euglena gracilis Z

Presenter:	Madhusudhan, Rapolu , rapolumadhu@hotmail.com
Authors	Madhusudhan, Rapolu  (A) (B)  Sawa, Yoshihiro  (A)   Shibata, Hitoshi  (A)   Shigeoka, Shigeru  (B)   Ishikawa, Takahiro  (A)
Affiliations:	(A): Department of Life Science and Biotechnology, Shimane University (B): Department of Food and Nutrition, Kinki University

When Euglena cells are grown heterotrophically in the dark, chloroplast development is stopped at the proplastid stage. Upon exposure to light, the cells rapidly develop chloroplasts and the photosynthesis resumes. We have investigated the regulation of cytosolic ascorbate peroxidase (APX) in E. gracilis during light adaptation of dark-grown cells. Both the activity and protein levels of APX increased by nearly 4 fold in about 24 hours of illumination. Cycloheximide almost completely inhibited the APX induction whereas transcription inhibitors did not have a significant effect, suggesting that the light induction of APX is post-transcriptionally regulated. We have isolated a partial cDNA of Euglena APX, which contained an open reading frame encoding for 296 amino acids. Interestingly, the partial cDNA sequence contained the code for almost the whole of the catalytic region. When expressed in E. coli, the cDNA encoded a catalytically active protein. Northern hybridization with the partial cDNA as the probe revealed a constant level of APX transcripts during the light adaptation. The APX induction was abolished by norflurazon, which inhibits the carotenoid synthesis. However, treatment of the dark-grown cells with H₂O₂, or methyl viologen did not induce APX. Our results suggest that the light induction of APX is dependent on the development of chloroplast.