Differential sensitivities of portions of the mRNA for ribosomal protein S20 to 3'-exonucleases dependent on oligoadenylation and RNA secondary structure.
The 3'-exonucleolytic decay of the mRNA for ribosomal protein S20 has been reconstituted in vitro using purified RNase II and crude extracts enriched for polynucleotide phosphorylase (PNPase) activity. We show that RNase II can catalyze the degradation of the 5' two-thirds of the S20 mRNA and that prior oligoadenylation of the 3' termini of truncated S20 mRNA substrates can significantly stimulate the initiation of degradation by RNase II. The intact S20 mRNA is, however, insensitive to attack by RNase II and polyadenylation of its 3'-end cannot overcome the natural resistance of the S20 mRNA to RNase II. Complete degradation of either the entire S20 mRNA without prior endonucleolytic cleavage or the 3'-terminal 147-residue fragment is dependent on both oligoadenylation and PNPase activity. Moreover, this process can take place in the absence of RNase E activity. Our data point to the importance of oligoadenylation in facilitating 3'-exonucleolytic activity and indicate that there are alternative degradative pathways. The implications for mRNA decay are discussed.
Coburn
G A
GA
Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, British Columbia, Canada V6T 1Z3.
Mackie
G A
GA
D000255:Adenosine Triphosphate / Q000378:metabolism; D001483:Base Sequence; D004722:Endoribonucleases / Q000378:metabolism; D004926:Escherichia coli; D005092:Exonucleases / Q000378:metabolism*; D005095:Exoribonucleases / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010710:Phosphates / Q000378:metabolism; D011061:Poly A / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D012269:Ribosomal Proteins / Q000235:genetics*
D010710:Phosphates; D012333:RNA, Messenger; D012269:Ribosomal Proteins; C029611:ribosomal protein S20; D011061:Poly A; D000255:Adenosine Triphosphate; D004722:Endoribonucleases; D005092:Exonucleases; D005095:Exoribonucleases; C022535:exoribonuclease II; C022537:ribonuclease E