To investigate the role of cytosine methylation in regulation of polyketide compounds biosynthesis in larvae of Strongylocentrotus intermedius. Treatment of S. intermedius larvae with 100 and 200 A mu M 5-azacytidine (5A) as a DNA demethylating agent significantly increased the amounts of spinochrome D and spinochrome E, as the number of pigmented cells per studied larva, in a dose-depended manner. The data on SiPks gene expression showed enhancement in 16- and 67-fold in S. intermedius larvae treated with 100 and 200 A mu M 5A, respectively, relative to untreated ones. Moreover, the activation of transcription factors SiGcm, SiGatae and SiKrl gene expression involved in regulation of SiPks was observed in S. intermedius larvae upon treatment with 5A, suggesting DNA methylation being powerful regulator of polyketide compounds biosynthesis. This is the first study to describe the role of cytosine DNA methylation in the regulation of polyketide compounds biosynthesis in sea urchins. Current study implies a negative control provided by cytosine DNA methylation machinery as a key regulator of polyketide compound biosynthesis.