REDUCTION OF COPROPORPHYRINOGEN OXIDASE LEVEL BY ANTISENSE RNA-SYNTHESIS LEADS TO DEREGULATED GENE-EXPRESSION OF PLASTID PROTEINS AND AFFECTS THE OXIDATIVE DEFENSE SYSTEM

A full-length cDNA sequence encoding coproporphyrinogen oxidase was in serted in inverse orientation behind a CaMV promoter and transferred t o tobacco (Nicotiana tabacum) by standard transformation techniques. T ransformants showed reduced coproporphyrinogen oxidase activity and ac cumulation of photosensitive coproporphyrin(ogen), indicating antisens e RNA expression. An inverse correlation was observed between the leve l of coproporphyrinogen oxidase and transformant phenotype. The latter is characterized by a broad range of growth retardation and necrosis, indicating oxidative leaf damage. Coproporphyrinogen is an apparent c hromophore and its excitation finally leads to the production of react ive oxygen. Evidence is presented that indicates a direct correlation between the accumulation of nonmetabolized coproporphyrinogen and oxid ative damage to cellular structural components. Enzymatic and non-enzy matic antioxidants were investigated. Whereas superoxide dismutase act ivity increased in transgenic plants, catalase and ascorbate peroxidas e activity remained constant. Tocopherol, rather than carotene or zeax anthin, seemed to be involved in detoxification, indicating the putati ve localization and allocation of coproporphyrinogen. Expression of co proporphyrinogen oxidase antisense RNA did not significantly influence the level of other enzymes in the chlorophyll metabolic pathway, but deregulated gene expression of nuclear encoded plastid proteins. Accum ulation of coproporphyrinogen and/or the resulting effects, such as ox idative stress, impairs a plastid/nuclear signal which may adapt gene expression to the plastid state.