Bioinformatic analysis of the plasmid-linked gene cluster associated with biosynthesis of methylenomycin (Mm) suggested that part of the cluster directs synthesis of a gamma-butyrolactone-like autoregulator. Autoregulator activity could be extracted from culture fluids, but differed from gamma-butyrolactones in being alkali resistant. The activity has recently been shown to comprise a series of novel autoregulator molecules, the methylenomycin furans (termed MMF). MMF autoregulator activity is shown to account for the ability of certain Mm non-producing mutants to act as 'secretors' in cosynthesis with other 'convertor' mutants. Three genes implicated in MMF biosynthesis are flanked by two regulatory genes, which are related to genes for gamma-butyrolactone-binding proteins. Genetic evidence suggests that these two genes encode components of a hetero-oligomeric repressor of MMF and Mm biosynthesis. The Mm biosynthetic genes themselves depend on the activator gene mmyB, which appears to be repressed by the putative MmyR/MmfR complex until enough MMF accumulates to release repression. The presence of TTA codons in mmyB and the main MMF biosynthetic gene causes Mm production to be dependent on the pleiotropically acting bldA gene, which encodes the tRNA for the rarely used UUA codon.