Post-transcriptional processes play a crucial role in controlling gene expression in all organisms. Our research is aimed at elucidating the mechanisms by which such control is imposed. To identify and characterize the proteins, RNA elements, and molecular mechanisms that govern these key regulatory processes, we employ a comprehensive approach combining biochemical, molecular biological, and genetic methods. We are particularly interested in understanding how gene expression in bacterial and mammalian cells is regulated by mRNA degradation.
5′-end-dependent mRNA degradation
In bacteria, the lifetimes of individual mRNAs can differ by more than an order of magnitude, with profound consequences for gene expression. For many years it had been assumed that bacterial mRNA degradation always begins with endonucleolytic cleavage at internal sites. However, our findings have overturned that view by showing that mRNA decay is often triggered by a prior non-nucleolytic event that marks transcripts for rapid turnover: the conversion of the 5′ terminus from a triphosphate to a monophosphate. In Escherichia coli and related organisms, this modification creates better substrates for the endonuclease RNase E, whose cleavage activity is greatly enhanced when the RNA 5′ end is monophosphorylated, whereas in Bacillus subtilis and other bacterial species that lack RNase E, it triggers 5′-exonucleolytic degradation by RNase J. We have identified a pyrophosphate-removing hydrolase, RppH, that is critical for this initial 5′-terminal event, the first such enzyme ever discovered. We have also characterized the substrate specificity and phylogeny of E. coli and B. subtilis RppH, which differ significantly. The inability of RppH to bind 5′ ends that are structurally sequestered by a stem-loop helps to explain the stabilizing influence of 5′-terminal base pairing on mRNA lifetimes in vivo. Interestingly, this master regulator of 5′-end-dependent mRNA degradation in bacteria not only catalyzes a process functionally reminiscent of eukaryotic mRNA decapping but also bears an evolutionary relationship to the eukaryotic decapping enzyme Dcp2.
RNase E autoregulation
RNase E is an essential regulatory enzyme whose overproduction or underproduction can impair cell growth. To ensure a steady supply of this protein, E. coli has evolved a homeostatic mechanism for tightly regulating its synthesis by modulating the decay rate of rne (RNase E) mRNA in response to changes in cellular RNase E activity. Our studies have shown that feedback regulation by RNase E is mediated in cis by the 361-nucleotide rne 5′ untranslated region (UTR), which can confer this property onto heterologous messages to which it is fused. The rne 5′ UTR is composed of six structural domains, two of which play crucial roles in feedback regulation. In vitro studies with purified components indicate that these 5′ UTR elements, like a 5′ monophosphate, expedite RNA degradation by binding to RNase E and guiding it to nearby cleavage sites.
Professor, Department of Microbiology
PhD from Harvard University
Fellowship, Stanford University, Genetics
Fellowship, Harvard University, Chemistry
Molecular cell. 2017 Jun 29; 67(1):44-54.e6
Journal of biological chemistry. 2016 Dec 14; 292(5):1934-1950
Journal of biological chemistry. 2016 Mar 04; 291(10):5038-5048
Journal of biological chemistry. 2015 Apr 10; 290(15):9478-9486
Nucleic acids research. 2015 Jan; 43(1):309-323
Proceedings of the National Academy of Sciences of the United States of America (PNAS). 2013 May 28; 110(22):8864-8869
Molecular cell. 2011 Sep 16; 43(6):940-949
Nature. 2008 Jan 17; 451(7176):355-358