Three Chymotrypsin Genes Are Members of the AdpA Regulon in the A-Factor Regulatory Cascade in Streptomyces griseus

Abstract

ABSTRACT AdpA is a key transcriptional activator in the A-factor regulatory cascade in Streptomyces griseus , activating a number of genes required for secondary metabolism and morphological differentiation. Of the five chymotrypsin-type serine protease genes, sprA , sprB , and sprD were transcribed in response to AdpA, showing that these protease genes are members of the AdpA regulon. These proteases were predicted to play the same physiological role, since these protease genes were transcribed in a similar time course during growth and the matured enzymes showed high end-to-end similarity to one another. AdpA bound two sites upstream of the sprA promoter approximately at positions −375 and −50 with respect to the transcriptional start point of sprA . Mutational analysis of the AdpA-binding sites showed that both AdpA-binding sites were essential for transcriptional activation. AdpA bound a single site at position −50 in front of the sprB promoter and greatly enhanced the transcription of sprB . The AdpA-binding site at position −40 was essential for transcription of sprD , although there was an additional AdpA-binding site at position −180. Most chymotrypsin activity excreted by S. griseus was attributed to SprA and SprB, because mutant Δ sprAB , having a deletion in both sprA and sprB , lost almost all chymotrypsin activity, as did mutant Δ adpA . Even the double mutant Δ sprAB and triple mutant Δ sprABD grew normally and developed aerial hyphae and spores over the same time course as the wild-type strain.