Porphyrin Accumulation and Export by Isolated Barley (Hordeum vulgare) Plastids (Effect of Diphenyl Ether Herbicides)

Abstract

We have investigated the formation of porphyrin intermediates by isolated barley (Hordeum vulgare) plastids incubated for 40 min with the porphyrin precursor 5-aminolevulinate and in the presente and absence of a diphenylether herbicide that blocks protoporphyrinogen oxidase, the enzyme in chlorophyll and heme synthesis that oxidizes protoporphyrinogen I X to protoporphyrin IX.In the absence of herbicide, about 50% of the protoporphyrin I X formed was found in the extraplastidic medium, which was separated from intact plastids by centrifugation at the end of the incubation period. In contrast, uroporphyrinogen, an earlier intermediate, and magnesium protoporphyrin IX, a later intermediate, were located mainly within the plastid. When the incubation was carried out in the-presence of a herbicide that inhibits protoporphyrinogen oxidase, protoporphyrin IX formation by the plastids was completely abolished, but large amounts of protoporphyrinogen accumulated in the extraplastidic medium. To detect extraplastidic protoporphyrinogen, it was necessary to first oxidize it to protoporphyrin IX with the use of a herbicide-resistant protoporphyrinogen oxidase enzyme present in Escbericbia coli membranes.Protoporphyrinogen is not detected by some commonly used methods for porphyrin analysis unless it is first oxidized to protoporphyrin IX. Protoporphyrin I X and protoporphyrinogen found outside the plastid did not arise from plastid lysis, because the percentage of plastid lysis, measured with a stromal marker enzyme, was far less than the percentage of these porphyrins in the extraplastidic fraction. These findings suggest that of the tetrapyrrolic intermediates synthesized by the plastids, protoporphyrinogen and protoporphyrin IX, are the most likely to be exported from the plastid to the cytoplasm. These results help explain the extraplastidic accumulation of protoporphyrin I X in plants treated with photobleaching herbicides. In addition, these findings suggest that plastids may export protoporphyrinogen or protoporphyrin IX for mitochondrial heme synthesis.There has been increased interest in the mechanism of plant Chl and heme synthesis due to recent studies showing that the light-dependent action of photobleaching diphenylether herbicides involves the accumulation of protoporphyrin IX, an intermediate in the heme and Chl synthetic pathways. This photoactive porphyrin can generate membrane-damag-

Mechanism and regulation of Mg-chelatase

“…There is a good physiological argument that this may be the case : mitochondria cannot synthesize haem, although they do contain ferrochelatase, and therefore chloroplasts must supply either haem or haem precursors to mitochondria and the rest of the cell. The efflux of haem and other porphyrins from isolated chloroplasts has been observed [61][62][63] . Possibly there are two haem-synthesis pathways in the chloroplast : one for photosynthetic uses and the other for non-photosynthetic and\or extrachloroplastidic haem.…”

Level of protoporphyrinogen oxidase activity tightly correlates with photodynamic and defense responses in oxyfluorfen-treated transgenic rice

“…3) Peroxidizing herbicides, including oxyfluorfen and acifluorfen, are photodynamically active and competitively block the substrate-binding region of Protox. [4][5][6][7] The acute toxicity of oxyfluorfen is a result of the accumulation of Protogen IX, which is oxidized in the cytoplasm to Proto IX by a nonspecific plasma membrane-bound peroxidase. 8,9) Proto IX in the cytoplasm is not metabolized by the porphyrin biosynthetic pathway because Mg-chelatase and Fe-chelatase, which use Proto IX as a substrate, are located in chloroplasts and mitochondria.…”

Cellular Localization of Protoporphyrinogen-Oxidizing Activities of Etiolated Barley (Hordeum vulgare L.) Leaves (Relationship to Mechanism of Action of Protoporphyrinogen Oxidase-Inhibiting Herbicides)

“…These data supported the view that, when plastid Protox is inhibited, Protogen IX migrates from the plastid to be oxidized to Proto IX by a PMbound Protox-like activity. Jacobs and Jacobs (1993) recently demonstrated that Protogen IX is exported at high levels from intact etioplasts treated with a Protox-inhibiting diphenyl ether herbicide. The accumulated extraplastidic Proto IX apparently does not readily reenter the porphyrin pathway; Plant Physiol.…”