Nucleic Acids Research Advance Access originally published online on March 31, 2009
Nucleic Acids Research 2009 37(10):3418-3430; doi:10.1093/nar/gkp207
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Nucleic Acids Research, 2009, Vol. 37, No. 10 3418-3430
© 2009 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Molecular Biology |
Cotranscriptional recruitment of the nuclear poly(A)-binding protein Pab2 to nascent transcripts and association with translating mRNPs
RNA Group, Department of Biochemistry, Université de Sherbrooke, Québec, Canada
*To whom correspondence should be addressed. Tel: +1 819 820 6868 (12733); Fax: +1 819 564 5340; Email: f.bachand{at}usherbrooke.ca
Received December 31, 2008. Revised March 10, 2009. Accepted March 12, 2009.
Synthesis of the pre-mRNA poly(A) tail in the nucleus has important consequences on the translational activity of the mature mRNA in the cytoplasm. In most eukaryotes, nuclear polyadenylation of pre-mRNAs is thought to require the nuclear poly(A)-binding protein (PABP2/PABPN1) for poly(A) tail synthesis and ultimate length control. As yet, however, the extent of the association between PABP2 and the exported mRNA remains poorly understood. Here, we used chromatin immunoprecipitation (ChIP) assays to show that the fission yeast ortholog of mammalian PABP2 (Pab2) is cotranscriptionally recruited to active genes. Notably, the association of Pab2 to genes precedes that of a typical 3'-processing/polyadenylation factor, suggesting that Pab2 recruitment during the transcription cycle precedes polyadenylation. The inclusion of an RNase step in our ChIP and immunoprecipitation assays suggests that Pab2 is cotranscriptionally recruited via nascent mRNA ribonucleoprotein (mRNPs). Tandem affinity purification coupled with mass spectrometry also revealed that Pab2 associates with several ribosomal proteins as well as general translation factors. Importantly, whereas previous results suggest that the nuclear poly(A)-binding protein is not present on cytoplasmic mRNAs, we show that fission yeast Pab2 is associated with polysomes. Our findings suggest that Pab2 is recruited to nascent mRNPs during transcription and remains associated with translated mRNPs after nuclear export.