Characterization of the alnumycin gene cluster reveals unusual gene products for pyran ring formation and dioxan biosynthesis.

Alnumycin is closely related to the benzoisochromanequinone (BIQ) polyketides such as actinorhodin. Exceptional structural features include differences in aglycone tailoring that result in the unique alnumycin chromophore and the existence of an unusual 4-hydroxymethyl-5-hydroxy-1,3-dioxan moiety. Cloning and sequencing of the alnumycin gene cluster from Streptomyces sp. CM020 revealed expected biosynthesis genes for polyketide assembly, but several genes encoding subsequent tailoring enzymes were highly atypical. Heterologous expression studies confirmed that all of the genes required for alnumycin biosynthesis resided within the sequenced clone. Inactivation of genes aln4 and aln5 showed that the mechanism of pyran ring formation differs from actinorhodin and granaticin pathways. Further inactivation studies identified two genes, alnA and alnB, involved in the synthesis and attachment of the dioxan moiety, and resulted in the production of the polyketide prealnumycin.

Diverse roles for chloroplast stromal and thylakoid-bound ascorbate peroxidases in plant stress responses.

Photosynthetic light reactions comprise a significant source of hydrogen peroxide (H(2)O(2)) in illuminated leaves. APXs (ascorbate peroxidases) reduce H(2)O(2) to water and play an important role in the antioxidant system of plants. In the present study we addressed the significance of chloroplast APXs in stress tolerance and signalling in Arabidopsis thaliana. To this end, T-DNA (transfer DNA) insertion mutants tapx, sapx and tapx sapx, lacking the tAPX (thylakoid-bound APX), sAPX (stromal APX) or both respectively, were characterized. Photo-oxidative stress during germination led to bleaching of chloroplasts in sapx single-mutant and particularly in the tapx sapx double-mutant plants, whereas the greening process of wild-type and tapx plants was only partially impaired. Mature leaves of tapx sapx double mutants were also susceptible to short-term photo-oxidative stress induced by high light or methyl viologen treatments. After a 2-week acclimation period under high light or under low temperature, none of the mutants exhibited enhanced stress symptoms. Immunoblot analysis revealed that high-light-stress-acclimated tapx sapx double mutants compensated for the absence of tAPX and sAPX by increasing the level of 2-cysteine peroxiredoxin. Furthermore, the absence of tAPX and sAPX induced alterations in the transcriptomic profile of tapx sapx double-mutant plants already under quite optimal growth conditions. We conclude that sAPX is particularly important for photoprotection during the early greening process. In mature leaves, tAPX and sAPX are functionally redundant, and crucial upon sudden onset of oxidative stress. Moreover, chloroplast APXs contribute to chloroplast retrograde signalling pathways upon slight fluctuations in the accumulation of H(2)O(2) in chloroplasts.