A transcriptional regulator of a pristinamycin resistance gene in Streptomyces coelicolor

Pip is a pristinamycin-induced transcriptional regulator protein detected i n many Streptomyces species by its ability to specifically bind sequence mo tifs within the promoter of a Streptomyces pristinaespiralis multidrug resi stance gene (ptr). To investigate the possible role of Pip in regulating mu ltidrug resistance, it was purified from a genetically characterized specie s, Streptomyces coelicolor, utilizing an affinity matrix of the ptr promote r conjugated to magnetic beads. Reverse genetics identified the correspondi ng locus and confirmed that it encoded Pip, a protein belonging to the TetR family of procaryotic transcriptional repressors. Pip binding motifs were located upstream of the adjacent gene pep, encoding a major facilitator ant iporter homologous to ptr. bz vivo analysis of antibiotic susceptibility pr ofiles demonstrated that pep conferred elevated levels of resistance only t o pristinamycin I (PI), a streptogramin B antibiotic having clinical import ance. Purified recombinant Pip was a dimer tin the presence or absence of P I) and displayed a high affinity for its palindromic binding motifs within the ptr promoter and the upstream region of pep, The Pip/ptr promoter compl ex was dissociated by PI but not by any of the other nonstreptogramin antib iotics that were described previously as transcriptional inducers. These pr ocaryotic regulatory elements served as the basis for the development of sy stems allowing repression or induction of cloned genes in mammalian and pla nt cells in response to streptogramin antibiotics (including pristinamycin, virginiamycin, and Synercid(R).