Identification of TmcN as a pathway-specific positive regulator of tautomycetin biosynthesis in Streptomyces sp. CK4412.

Tautomycetin (TMC) is a novel activated T-cell-specific immunosuppressive compound with a unique structure, containing an ester bond linkage between a terminal cyclic anhydride moiety and a linear polyketide chain bearing an unusual terminal alkene. A 3 kb gene, tmcN, with a deduced product of 1029 amino acid residues, located on the 3'-terminus of an approximately 70 kb contiguous TMC biosynthetic gene cluster, was found to have amino acid sequence homology with bacterial regulatory proteins. In silico database comparisons revealed that TmcN belongs to the large ATP-binding regulators of the LuxR protein family. Gene disruption of tmcN from the Streptomyces sp. CK4412 chromosome resulted in significantly reduced antifungal activity against Aspergillus niger, as well as the absence of TMC. In addition, complementation by an integrative plasmid carrying tmcN restored TMC biosynthesis, strongly suggesting that TmcN is a positive regulator of TMC biosynthesis. Gene expression analysis by RT-PCR of the TMC biosynthetic genes revealed that a TmcN mutant strain exhibited reduced expression levels for most of the biosynthetic genes except for its own tmcN. It is thus suggested that TmcN is a pathway-specific positive regulator that activates transcription of the TMC biosynthetic pathway genes in Streptomyces sp. CK4412.