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Nucleic Acids Research Advance Access published online on November 25, 2008
Nucleic Acids Research, doi:10.1093/nar/gkn929
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© 2008 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.

RNA

Identification of a signature motif in target mRNAs of RNA-binding protein AUF1

Krystyna Mazan-Mamczarz1,2, Yuki Kuwano1, Ming Zhan3, Elizabeth J. White1, Jennifer L. Martindale1, Ashish Lal1,4 and Myriam Gorospe1,*

1Laboratory of Cellular and Molecular Biology, National Institute on Aging-Intramural Research Program, National Institutes of Health, Baltimore, MD 21224, 2University of Maryland, Marlene and Stewart Greenebaum Cancer Center, Baltimore, MD 21201, 3Research Resources Branch, National Institute on Aging-Intramural Research Program, National Institutes of Health, Baltimore, MD 21224 and 4Immune Disease Institute and Department of Pediatrics, Harvard Medical School, Boston, MA 02115, USA

*To whom correspondence should be addressed. Tel: +1 410 558 8443; Fax: +1 410 558 8386; Email: myriam-gorospe{at}nih.gov

Received September 4, 2008. Revised October 15, 2008. Accepted November 4, 2008.

The ubiquitous RNA-binding protein AUF1 promotes the degradation of some target mRNAs, but increases the stability and translation of other targets. Here, we isolated AUF1-associated mRNAs by immunoprecipitation of (AUF1–RNA) ribonucleoprotein (RNP) complexes from HeLa cells, identified them using microarrays, and used them to elucidate a signature motif shared among AUF1 target transcripts. The predicted AUF1 motif (29–39 nucleotides) contained 79% As and Us, consistent with the AU-rich sequences of reported AUF1 targets. Importantly, 10 out of 15 previously reported AUF1 target mRNAs contained the AUF1 motif. The predicted interactions between AUF1 and target mRNAs were recapitulated in vitro using biotinylated RNAs. Interestingly, further validation of predicted AUF1 target transcripts revealed that AUF1 associates with both the pre-mRNA and the mature mRNA forms. The consequences of AUF1 binding to 10 predicted target mRNAs were tested by silencing AUF1, which elevated the steady-state levels of only four mRNAs, and by overexpressing AUF1, which also lowered the levels of only four mRNAs. In total, we have identified a signature motif in AUF1 target mRNAs, have found that AUF1 also associates with the corresponding pre-mRNAs, and have discovered that altering AUF1 levels alone only modifies the levels of subsets of target mRNAs.


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