Molecular mechanisms underlying the regulation of gene expression by non-coding RNAs in archaea

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This project investigates the role of small non-coding RNAs (sRNAs) in the response of microorganisms to environmental stress. Hundreds of sRNAs have been discovered in Archaea, a group of microorganisms with a close evolutionary relationship with Eukaryotes, but little is known about their mechanisms of action and how they modulate the functions of living organisms through evolution. The research will elucidate the regulatory landscape of an sRNA in a model archaeon in order to address this gap in knowledge. A deeper understanding of archaeal sRNA functions and regulation will enable manipulation of sRNA molecules as synthetic biology tools and rational design of biological systems for biotechnology applications. The broader impact activities will engage students from diverse backgrounds early in their STEM careers with laboratory research, especially first-generation college students, women, and underrepresented minorities. Students will be recruited through FiGURE (First Generation Undergrads Research Experience), an innovative research experience and cohort building program at Johns Hopkins University. These activities will also provide mentorship training for graduate students.This research aims to gain a mechanistic understanding of sRNA regulation in a model archaeon, Haloferax volcanii (Hv) in response to oxidative stress. The specific goals are to (1) characterize post-transcriptional mechanisms of the SHOxi sRNA-mediated oxidative stress response and determine how SHOxi regulates putative targets by modulating mRNA stability or interfering with translation; (2) identify chaperone proteins and other factors involved in the interactions of oxidative stress-responsive sRNAs with their targets; and (3) elucidate how the stress-responsive sRNA SHOxi is regulated under oxidative stress (particularly testing the hypothesis that canonical archaeal transcription factors regulate SHOxi). The research will employ a combination of genetic and biochemical approaches, site-directed mutagenesis combined with sRNA-mRNA interaction modeling, and high-throughput phenotyping. The outcomes are expected to reveal mechanisms by which an archaeal sRNA binds and regulates its targets, identify novel RNA binding proteins, and discover how sRNA-mediated regulatory networks are integrated with those of transcription regulators in archaea. The resulting knowledge will be of interest to the archaeal community and the non-coding RNA and RNA communities.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.