RPOS-DEPENDENT STATIONARY PHASE INDUCTION OF NITRATE REDUCTASE Z IN E. COLI

Abstract

During entry into stationary phase, Escherichia coli expresses many genes which impart cellular resistance to numerous environmental stresses such as oxidative or acid stress. Many ofthese genes are regulated by the alternative sigma factor, RpoS To identify additional genes regulated by RpoS, a phenotype independent genetic screen was previously employed (L. Wei Masters thesis). The identities ofthe ten most highly RpoSdependent fusions were determined by DNA sequencing and subsequent sequence analysis using the BLAST algorithm Three fusions map to genes previously known to be RpoSdependent while the remaining seven represent new members ofthe regulon The expression of many ofthe RpoS-dependent fusions remained growth phase dependent even in the rpoS background This suggests that other growth phase regulatory factors in addition to RpoS may coordinate stationary phase gene expression Upon sequencing the remaining rsd fusions, three mutants mapped to narY which is part ofthe narZYWV operon encoding the secondary nitrate reductase Z (NRZ) This operon was selected for further investigation since NRZ has been previously reported to be constitutively expressed Expression studies using promoter lacZ fusions and nitrate reductase assays reveal that NRZ is induced ten-fold at the onset ofstationary phase and twenty-fold in the presence of nitrate Like other rsd fusions, growth phase dependent expression was observed in an rpoS background indicating that other regulatory factors may be involvedn the regulation of NRZ Northern analyses using probes specific to NRZ confirmed that transcription of NRZ is indeed dependent on RpoS. These results suggest that RpoS mediated regulation of NRZ may be an important physiological adaptation to reduced oxygen levels during transition to stationary phase