The phytochrome-deficient aurea mutant of tomato has been widely used for the study of both phytochrome function and the role of other photoreceptors in the control of development in higher plants. To date the exact nature of the aurea mutation has remained unknown, though this information is clearly important for the interpretation of these studies. It has been proposed that aurea and yellow-green-2, another mutant of tomato that has a similar phenotype to aurea, could be deficient in phytochrome chromophore synthesis. We have examined this hypothesis by measuring the activity of the enzymes committed to phytochrome chromophore synthesis in these mutants. The approach takes advantage of a recently developed high pressure liquid chromatography-based assay for the synthesis of the free phytochrome chromophore, phytochromobilin from its immediate precursors biliverdin IXalpha and heme. Isolated etioplasts from aurea and yellow-green-2 seedlings were specifically unable to convert biliverdin IXalpha to 3Z-phytochromobilin and heme to biliverdin IXalpha, respectively. In addition, the level of total noncovalently bound heme in the mutants was the same as in wild type seedlings. Together, these results identify both aurea and yellow-green-2 as mutants that are deficient in phytochrome chromophore synthesis.

The aurea (au) and yellow-green-2 (yg-2) mutants of tomato (Solanum lycopersicum L.) are unable to synthesize the linear tetrapyrrole chromophore of phytochrome, resulting in plants with a yellow-green phenotype. To understand the basis of this phenotype, we investigated the consequences of the au and yg-2 mutations on tetrapyrrole metabolism. Dark-grown seedlings of both mutants have reduced levels of protochlorophyllide (Pchlide) due to an inhibition of Pchlide synthesis. Feeding experiments with the tetrapyrrole precursor 5-aminolevulinic acid (ALA) demonstrate that the pathway between ALA and Pchlide is intact in au and yg-2 and suggest that the reduction in Pchlide is a result of the inhibition of ALA synthesis. This inhibition was independent of any deficiency in seed phytochrome, and experiments using an iron chelator to block heme synthesis demonstrated that both mutations inhibited the degradation of the physiologically active heme pool, suggesting that the reduction in Pchlide synthesis is a consequence of feedback inhibition by heme. We discuss the significance of these results in understanding the chlorophyll-deficient phenotype of the au and yg-2 mutants.

Inhibition of chromophore synthesis in the phytochrome-deficient aurea (au) and yellow-green-2 (yg-2) mutants of tomato (Solanum lycopersicum L.) results in a severe reduction of protochlorophyllide (Pchlide) accumulation in dark-grown hypocotyls. Experiments with apophytochrome-deficient mutants indicate that the inhibition of Pchlide accumulation results from two separate effects: one dependent on the activity of phytochromes A and B1 and one phytochrome-independent effect that is attributed to a feedback inhibition of the tetrapyrrole biosynthesis pathway. Cotyledons only show phytochrome-independent inhibition of Pchlide synthesis. Analysis of NADPH:protochlorophyllide oxidoreductase levels by western blotting showed that the reduction in Pchlide in au and yg-2 is accompanied by a correlative, but less substantial, decrease in NADPH:protochlorophyllide oxidoreductase. Consistent with this result, in vivo fluorescence spectra demonstrate that both mutants are primarily deficient in non-phototransformable Pchlide. Analysis of etioplast structure indicates that plastid development in au and yg-2 is retarded in hypocotyls and partially impaired in cotyledons, again correlating with the reduction in Pchlide. Since Pchlide synthesis is also reduced in chromophore-deficient mutants of pea (Pisum sativum L.) and Arabidopsis thaliana (L.) Heynh. (Landsberg erecta) these results may be significant for explaining aspects of the phenotype of this mutant class that are independent of the loss of phytochrome.

The aurea mutants of tomato have been widely used as phytochrome-deficient mutants for photomorphogenetic and photobiological studies. By expressed sequence tag (EST)-based screening of sequence databases, we found a tomato gene that encodes a protein homologous to Arabidopsis HY2 for phytochromobilin synthase catalyzing the last step of phytochrome chromophore biosynthesis. The tomato protein expressed in Escherichia coli showed phytochromobilin synthase activity. The corresponding loci in all aurea mutants tested have nucleotide substitutions, deletions or DNA rearrangements. These results indicate that aurea is a mutant of phytochromobilin synthase in tomato. We also discuss a phylogenetic analysis of phytochromobilin synthases in the bilin reductase family.