The RNA-binding protein HF-I, known as a host factor for phage Qβ RNA replication, is essential for rpoS translation in Escherichia coli

Abstract

The rpoS-encoded σ(s) subunit of RNA polymerase in Escherichia coli is a global regulatory factor involved in several stress responses. Mainly because of increased rpoS translation and stabilization of σ(s), which in nonstressed cells is a highly unstable protein, the cellular σ(s) content increases during entry into stationary phase and in response to hyperosmolarity. Here, we identify the hfq-encoded RNA-binding protein HF-I, which has been known previously only as a host factor for the replication of phage Qβ RNA, as an essential factor for rpoS translation. An hfq null mutant exhibits strongly reduced σ(s) levels under all conditions tested and is deficient lot growth phase-related and osmotic induction of σ(s). Using a combination of gene fusion analysis and pulse-chase experiments, we demonstrate that the hfq mutant is specifically impaired in rpoS translation. We also present evidence that the H-NS protein, which has been shown to affect rpoS translation, acts in the same regulatory pathway as HF-I at a position upstream of HF-I or in conjunction with HF-I. In addition, we show that expression and heat induction of the heat shock σ factor σ32 (encoded by rpoH) is not dependent on HF-I, although rpoH and rpoS are both subject to translational regulation probably mediated by changes in mRNA secondary structure. HF-I is the first factor known to be specifically involved in rpoS translation, and this role is the first cellular function to be identified for this abundant ribosome-associated RNA-binding protein in E. coli.