IDENTIFICATION AND ANALYSIS OF AARP, A TRANSCRIPTIONAL ACTIVATOR OF THE 2'-N-ACETYLTRANSFERASE IN PROVIDENCIA-STUARTII

The aarP gene has been identified in a search for activators of the 2- N-acetyltransferase [encoded by aac(2')-Ia] in Providencia stuartii, I ntroduction of aarP into P, stuartii on a multicopy plasmid resulted i n a 9.9-fold increase in the accumulation of beta-galactosidase from a n aac(2')-lacZ fusion, Northern (RNA) blot analysis demonstrated that this increased aac(2')-Ia expression occurred at the level of mRNA acc umulation, The deduced AarP protein was 15,898 Da in size and exhibite d significant homology to a number of transcriptional activators in th e AraC/XyIS family, including TetD, Rob, MarA, and SoxS, The similarit y of AarP to the MarA and SoxS proteins prompted an investigation to d etermine whether AarP is involved in activation of genes in either the multiple antibiotic resistance (Mar) phenotype or redox stress (SoxRS ) system, Introduction of aarP on a multicopy plasmid into either P. s tuartii or Escherichia coli conferred a Mar phenotype with higher leve ls of resistance to tetracycline, chloramphenicol, and ciprofloxacin, Multiple copies of aarP in E, coli also resulted in activation of the endonuclease IV gene (nfo), a gene in the SoxRS regulon of E. coli, Th e function of aarP in its single-copy state was addressed by using all elic replacement to construct an aarP::Cm disruption, which resulted i n a fivefold reduction in the accumulation of aac(2')-Ia mRNA, Analysi s of aarP regulation showed that aarP mRNA accumulation,vas slightly i ncreased by exposure to tetracycline and dramatically increased in cel ls containing the aarB3 (aar3) mutation, which was previously shown to increase transcription of the aac(2')-Ia gene