Three pathway-specific regulators control streptolydigin biosynthesis in Streptomyces lydicus

The streptolydigin biosynthetic gene cluster from Streptomyces lydicus NRRL 2433 contains three putative regulatory genes, slgR1, slgR2 and slgY, encoding proteins belonging to TetR and LuxR transcriptional regulator families and ATP/GTP-binding proteins of DNA and RNA helicase superfamily I, respectively. Inactivation of slgR1 or slgR2 resulted in the abolition of streptolydigin production, suggesting that these proteins are pathway-specific positive regulators. In the case of the slgR1 mutant, low amounts of streptolydigin C were produced instead of streptolydigin. RT-PCR transcription analysis of streptolydigin biosynthesis genes revealed a hierarchical regulation process. SlgY was found to control the expression of the regulator slgR2. SlgR2 regulates the expression of structural genes involved in the formation of the streptolydigin bicyclic ketal moiety, incorporation and processing of 3-methylaspartate, and the regulator slgR1. On the other hand, SlgR1 controls the expression of slgE1–E2, involved in the conversion of glutamate to 3-methylaspartate, and putative glycoside hydrolases slgC1 and slgC2. Ectopic expression of slgR1, slgR2 and slgY regulatory genes in S. lydicus led to considerable increases in streptolydigin yields, 18-, 11- and 8.5-fold, respectively. Ectopic expression of slgY in an slgR1 mutant led to a 14-fold increase of streptolydigin C yields, while no effect was observed to result from expression of slgR2.