Role of terminator hairpin in biogenesis of functional Hfq-binding sRNAs

Abstract

Many Rho-independent transcription terminators of the genes encoding bacterial Hfq-binding sRNAs possess a set of 7 or more T residues at the 3′ end (Otaka et al. 2011; Ishikawa et al. 2012). Here, we have studied the role of terminator hairpin in biogenesis of sRNAs focusing on SgrS and RyhB in Esherichia coli. We constructed variant sRNA genes in which the GC-rich inverted repeat sequences are extended to stabilize the terminator hairpins. We demonstrate that the extension of the hairpin stem leads to generation of heterogeneous transcripts in which the polyU tail is shortened. The transcripts with shortened polyU tails no longer bind to Hfq and lose the ability to repress the target mRNAs. The shortened transcripts are generated in an in vitro transcription system with purified RNA polymerase, indicating that the generation of shortened transcripts is caused by premature transcription termination. We conclude that the terminator structure of sRNA genes is optimized to generate functional sRNAs. Thus, the Rho-independent terminators of sRNA genes possess two common features: a long T residue stretch that is prerequisite for generation of functional sRNAs and a moderate strength of hairpin structure that ensures the termination at the seventh or longer position within the consecutive T stretch. The modulation of termination position at the Rho-independent terminators is critical for biosynthesis of functional sRNAs.