title: Determinants in the rpsT mRNAs recognized by the 5'-sensor domain of RNase E.
abstract: RNase E plays a central role in processing virtually all classes of cellular RNA in many bacterial species. A characteristic feature of RNase E and its paralogue RNase G, as well as several other unrelated ribonucleases, is their preference for 5'-monophosphorylated substrates. The basis for this property has been explored in vitro. At limiting substrate, cleavage of the rpsT mRNA by RNase E (residues 1-529) is inefficient, requiring excess enzyme. The rpsT mRNA is cleaved sequentially in a 5' to 3' direction, with the initial cleavage(s) at positions 116/117 or 190/191 being largely driven by direct entry, independent of the 5'-terminus or the 5'-sensor domain of RNase E. Generation of the 147 nt 3'-limit product requires sequential cleavages that generate 5'-monophosphorylated termini on intermediates, and the 5'-sensor domain of RNase E. These requirements can be bypassed with limiting enzyme by deleting a stem-loop structure adjacent to the site of the major, most distal cleavage. Alternatively, this specific cleavage can be activated substantially by a 5'-phosphorylated oligonucleotide annealed 5' to the cleavage site. This finding suggests that monophosphorylated small RNAs may destabilize their mRNA targets by recruiting the 5-sensor domain of RNase E 'in trans'.
authors.affiliation: Department of Biochemistry & Molecular Biology, Life Sciences Centre, The University of British Columbia, Vancouver, BC, Canada. george.mackie@ubc.ca
authors.forename: George A
authors.initials: GA
authors.lastname: Mackie
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D020134:Catalytic Domain; D004722:Endoribonucleases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D009841:Oligonucleotides / Q000235:genetics; D012329:RNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012269:Ribosomal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D009841:Oligonucleotides; D012329:RNA, Bacterial; D012333:RNA, Messenger; D012269:Ribosomal Proteins; C029611:ribosomal protein S20; D004722:Endoribonucleases; C022537:ribonuclease E