FIGURE 6.

Toxicity of N,N-dimethyldoxorubicin. (A) G001, MAG301, MAG302, MAG303 and MAG304 were streaked on SFM agar plates (supplemented with 20 μg mL⁻¹ thiostrepton for strains harbouring pRDS). The development is blocked by the production of anthracyclines. MAG304 exhibited the most pronounced inhibition of development. In contrast, deletion of the glycosyltransferase gene dnrS in MAG302 stimulated development. (B) MAG305 pWHM3-oriT and MAG309 (drrAB) were spotted on SFM agar plates supplemented with increasing concentrations of doxorubicin (6) or N,N-dimethyldoxorubicin (12). For each strain, 5 µL of spore or mycelium stock was spotted at a concentration of 1.0.10⁴ CFU per spot and the plates were incubated at 30 °C for 4 days. Dashed circles indicate the highest concentration that does not inhibit growth. For MAG305, the inhibitory concentration of N,N-dimethyldoxorubicin (12) is 16-fold lower than that of doxorubicin (6). Overexpression of drrAB increased resistance to both compounds eight-fold. (C) LC-MS analysis of crude extracts of MAG305 pWHM3-oriT and MAG309 (drrAB) cultivated in E1 medium. Extracted ion chromatograms showing the mass peaks [M + H]⁺ of compounds 7–12. Overexpression of drrAB resulted in a 3.7-fold increased production of N,N-dimethyldaunorubicin (11).