PT  - JOURNAL ARTICLE
AU  - Katz, Noa
AU  - Cohen, Roni
AU  - Solomon, Oz
AU  - Kaufmann, Beate
AU  - Atar, Orna
AU  - Yakhini, Zohar
AU  - Goldberg, Sarah
AU  - Amit, Roee
TI  - RBP-RNA interactions in the 5’ UTR lead to structural changes that alter translation
AID  - 10.1101/174888
DP  - 2018 Jan 01
TA  - bioRxiv
PG  - 174888
4099  - http://biorxiv.org/content/early/2018/04/13/174888.short
4100  - http://biorxiv.org/content/early/2018/04/13/174888.full
AB  - We study translational regulation by a 5’ UTR sequence encoding the binding site of an RNA-binding protein (RBP) in bacteria, using a reporter assay and Selective 2’-hydroxyl acylation analysed by primer extension sequencing (SHAPE-Seq). We tested constructs containing a single hairpin, based on the binding sites of the coat RBPs of bacteriophages GA, MS2, PP7, and Qβ, positioned in the 5’ UTR of a reporter gene. With specifically-bound RBP present, either weak repression or up-regulation is observed, depending on the binding site and its flanking sequence. SHAPE-Seq data for a representative construct exhibiting up-regulation, indicates a partially-folded hairpin and non-reactive upstream and downstream flanking region, which we attribute to intermediate structures that apparently blocks translation. RBP binding stabilizes the fully-folded hairpin state and thus facilitates translation, suggesting that the up-regulating constructs are RBP-sensing riboswitches. This finding is further supported by lengthening the binding-site stem, which in turn destabilizes the translationally-inactive state, and abolishes the up-regulating behavior. Finally, we found that the combination of two binding sites, positioned in the 5’ UTR and gene-header of the same transcript, can yield a cooperative regulatory response. Together, we show that the interaction of an RBP with its RNA target facilitates structural changes in the RNA, which is reflected by a controllable range of binding affinities and dose response behaviors. Thus, demonstrating that RNA-RBP interactions can provide a platform for constructing gene regulatory networks that are based on translational, rather than transcriptional, regulation.