Characterization of conserved sequence elements in eukaryotic RNase P RNA reveals roles in holoenzyme assembly and tRNA processing.
RNase P is a ubiquitous endoribonuclease responsible for cleavage of the 5' leader of precursor tRNAs (pre-tRNAs). Although the protein composition of RNase P holoenzymes varies significantly among Bacteria, Archaea, and Eukarya, the holoenzymes have essential RNA subunits with several sequences and structural features that are common to all three kingdoms of life. Additional structural elements of the RNA subunits have been found that are conserved in eukaryotes, but not in bacteria, and might have functions specifically required by the more complex eukaryotic holoenzymes. In this study, we have mutated four eukaryotic-specific conserved regions in Saccharomyces cerevisiae nuclear RNase P RNA and characterized the effects of the mutations on cell growth, enzyme function, and biogenesis of RNase P. RNase P with mutations in each of the four regions tested is sufficiently functional to support life although growth of the resulting yeast strains was compromised to varying extents. Further analysis revealed that mutations in three different regions cause differential defects in holoenzyme assembly, localization, and pre-tRNA processing in vivo and in vitro. These data suggest that most, but not all, eukaryotic-specific conserved regions of RNase P RNA are important for the maturation and function of the holoenzyme.
Xiao
Shaohua
S
Department of Biological Chemistry, University of Michigan, Ann Arbor, 48109-0606, USA.
Day-Storms
Jeremy J
JJ
Srisawat
Chatchawan
C
Fierke
Carol A
CA
Engelke
David R
DR
D001483:Base Sequence; D017124:Conserved Sequence; D020035:Holoenzymes / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D012322:RNA Precursors / Q000378:metabolism*; D012323:RNA Processing, Post-Transcriptional; D012331:RNA, Fungal / Q000737:chemistry*; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D012343:RNA, Transfer / Q000378:metabolism*; D043262:Ribonuclease P / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism / Q000502:physiology
D020035:Holoenzymes; D012322:RNA Precursors; D012331:RNA, Fungal; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; D012343:RNA, Transfer; D043262:Ribonuclease P