Two-component signal transduction system (TCS) is a common mechanism of signal transduction and exists in almost all bacterial species. The putative two-component genes SCO6421 (ecrE(1)) and SCO6422 (ecrE(2)) in Streptomyces coelicolor, which located in the vicinity of the red locus, could modulate the antibiotic undecylprodigiosin (Red) biosynthesis. In this study, the mutants of M145E1 (ecrE(1) knock-out) and M145E2 (ecrE(2) knock-out) were constructed using homologous recombination by double crossover. Red production of two mutants decreased dramatically compared with that of the wild-type strain. However, the production of actinorhodin (Act) of two mutants had not changed significantly. The results suggested that ecrE(1)/ecrE(2) participated in regulating the antibiotic Red synthesis. Northern blotting analysis indicated that ecrE(1)/ecrE(2) may regulate the biosynthesis of Red through influencing the transcription of redD and redZ during cell growth. Therefore, ecrE(1)/ecrE(2), as a specific pair of TCS, most likely regulates the red gene cluster positively.