The transcriptional regulator of landomycin A biosynthesis encoded by lanI gene has been inactivated within the chromosome of Streptomyces cyanogenus S136. The obtained mutant strain did not produce landomycin A and its known intermediates. Loss of landomycin A production caused significant changes in morphology of the lanI deficient strain. RT-PCR analysis confirmed complete cessation of transcription of certain lan genes, including lanJ (encoding putative proton dependent transporter) and lanK (presumably involved in lanJ expression regulation). Introduction of either lanI or lndI [lanI homologue controlling landomycin E biosynthesis in Streptomyces globisporus 1912, both encoding Streptomyces antibiotic regulatory proteins (SARPs)] restored landomycin A production in the mutant strain. Chimeric constructs ladI and ladR were generated by exchanging the DNA sequences corresponding to N- and C-terminal parts of LndI and LanI. None of these genes were able to activate the production of landomycins in regulatory mutants of S. cyanogenus and S. globisporus. Nevertheless, the production of novel unidentified compound was observed in the case of S. cyanogenus harboring ladI gene. Various genes encoding SARPs have been expressed in S. globisporus and S. cyanogenus regulatory mutants and the results of these complementation experiments are discussed.