In vivo targeting of the yeast Pop2 deadenylase subunit to reporter transcripts induces their rapid degradation and generates new decay intermediates.
Deadenylation is the rate-limiting step of mRNA decay, yet little is known about the mechanism regulating this process. In yeast, deadenylation is mainly mediated by the Pop2-Ccr4 complex. We tested whether the selective recruitment of this deadenylase to target mRNAs was sufficient to stimulate their decay in vivo. For this purpose, the Pop2 factor was fused to a U1A RNA binding domain while U1A binding sites were inserted in untranslated regions of a reporter transcript. Analysis of the reporter fate in strains expressing the Pop2-U1A-RBD fusion demonstrated a specific activation of target mRNA decay. Increased mRNA degradation involved accumulation of deadenylated mRNAs that was not detected when the control factors Dcp2 or Pub1 were tethered to the same transcript. The rapid target mRNA degradation was also accompanied by the appearance of new decay intermediates generated by the 3' -5' trimming of the corresponding 3' -untranslated region. Interestingly, this process was not mediated by the exosome but may result from the activity of the Pop2-Ccr4 deadenylase itself. These results indicate that selective recruitment of the Pop2-Ccr4 deadenylase is sufficient to activate mRNA decay, even though this process can also be stimulated by additional mechanisms. Furthermore, deadenylase recruitment affects the downstream path of mRNA decay.
Finoux
Anne-Laure
AL
Equipe Labelisée La Ligue, Centre de Génétique Moléculaire, CNRS Unite Propre de Recherche 2167 Associée à l'Université P. et M. Curie, Avenue de la Terrasse, 91198 Gif sur Yvette Cedex, France.
Séraphin
Bertrand
B
D000818:Animals; D001483:Base Sequence; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D017930:Genes, Reporter; D006801:Humans; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D012260:Ribonucleases / Q000235:genetics / Q000378:metabolism*; D017412:Ribonucleoprotein, U1 Small Nuclear / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
D005656:Fungal Proteins; D021122:Protein Subunits; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D017412:Ribonucleoprotein, U1 Small Nuclear; D029701:Saccharomyces cerevisiae Proteins; C075339:U1A protein; D012260:Ribonucleases; C079009:POP2 protein, S cerevisiae