RIP-Seq Suggests Translational Regulation by L7Ae in Archaea

L7Ae is a universal archaeal protein that recognizes and stabilizes kink-turn (k-turn) motifs in RNA substrates. These structural motifs are widespread in nature and are found in many functional RNA species, including ribosomal RNAs. Synthetic biology approaches utilize L7Ae/k-turn interactions to control gene expression in eukaryotes. Here, we present results of comprehensive RNA immunoprecipitation sequencing (RIP-Seq) analysis of genomically tagged L7Ae from the hyperthermophilic archaeon Sulfolobus acidocaldarius. A large set of interacting noncoding RNAs was identified. In addition, several mRNAs, including the l7ae transcript, were found to contain k-turn motifs that facilitate L7Ae binding. In vivo studies showed that L7Ae autoregulates the translation of its mRNA by binding to a k-turn motif present in the 5′ untranslated region (UTR). A green fluorescent protein (GFP) reporter system was established in Escherichia coli and verified conservation of L7Ae-mediated feedback regulation in Archaea. Mobility shift assays confirmed binding to a k-turn in the transcript of nop5-fibrillarin, suggesting that the expression of all C/D box sRNP core proteins is regulated by L7Ae. These studies revealed that L7Ae-mediated gene regulation evolved in archaeal organisms, generating new tools for the modulation of synthetic gene circuits in bacteria.