The Sm-like protein Hfq regulates polyadenylation dependent mRNA decay in Escherichia coli.
In Escherichia coli, the post-transcriptional addition of poly(A) tails by poly(A) polymerase I (PAP I, pcnB) plays a significant role in cellular RNA metabolism. However, many important features of this system, including its regulation and the selection of polyadenylation sites, are still poorly understood. Here we show that the inactivation of Hfq (hfq), an abundant RNA-binding protein, leads to the reduction in the ability of PAP I to add poly(A) tails at the 3' termini of mRNAs containing Rho-independent transcription terminators even though PAP I protein levels remain unchanged. Those poly(A) tails that are synthesized in the absence of Hfq are shorter in length, even in the absence of polynucleotide phosphorylase (PNPase), RNase II and RNase E. In fact, the biosynthetic activity of PNPase in the hfq single mutant is enhanced and it becomes the primary polynucleotide polymerase, adding heteropolymeric tails almost exclusively to 3' truncated mRNAs. Surprisingly, both PNPase and Hfq co-purified with His-tagged PAP I under native conditions indicating a potential complex among these proteins. Immunoprecipitation experiments using PNPase- and Hfq-specific antibodies confirmed the protein-protein interactions among PAP I, PNPase and Hfq. Analysis of mRNA half-lives in hfq, deltapcnB and hfq deltapcnB mutants suggests that Hfq and PAP I function in the same mRNA decay pathway.
Mohanty
Bijoy K
BK
Department of Genetics, University of Georgia, Athens, GA 30602, USA.
Maples
Valerie F
VF
Kushner
Sidney R
SR
D001483:Base Sequence; D004722:Endoribonucleases / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D005095:Exoribonucleases / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D035001:Host Factor 1 Protein / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D009690:Nucleic Acid Conformation; D026723:Polyadenylation; D011062:Polynucleotide Adenylyltransferase / Q000235:genetics / Q000378:metabolism*; D011117:Polyribonucleotide Nucleotidyltransferase / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D020871:RNA Stability; D012333:RNA, Messenger / Q000378:metabolism*; D020741:rho GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism
D029968:Escherichia coli Proteins; C087373:Hfq protein, E coli; D035001:Host Factor 1 Protein; D012333:RNA, Messenger; D011062:Polynucleotide Adenylyltransferase; C063643:pcnB protein, E coli; D011117:Polyribonucleotide Nucleotidyltransferase; D004722:Endoribonucleases; D005095:Exoribonucleases; C022535:exoribonuclease II; C022537:ribonuclease E; D020741:rho GTP-Binding Proteins