Early Recruitment of AU-Rich Element-Containing mRNAs Determines Their Cytosolic Fate during Iron Deficiency

Abstract

The yeast Cth2 protein is a CX₈CX₅CX₃H tandem zinc finger protein that binds AU-rich element (ARE)-containing transcripts to enhance their decay in response to iron (Fe) deficiency. Mammalian members of this family of proteins are known to undergo nucleocytoplasmic shuttling, but little is known about the role of shuttling in the mechanism of ARE-dependent mRNA decay. Here we demonstrate that, like its mammalian homologues, Cth2 is a nucleocytoplasmic shuttling protein whose nuclear export depends on mRNA transport to the cytosol. The nuclear import information of Cth2 is contained within its tandem zinc finger domain, but it is independent of mRNA-binding function. Moreover, we also demonstrate that nucleocytoplasmic shuttling of Cth2 requires active transcription and that disruption of shuttling leads to defects in Cth2 function in mRNA decay under Fe deficiency. Taken together, our data suggest that under conditions of Fe deficiency Cth2 travels into the nucleus to recruit target mRNAs, perhaps cotranscriptionally, that are destined for cytosolic degradation as part of the mechanism of adaptation to growth under Fe limitation. These data also suggest an important role for nucleocytoplasmic shuttling in this conserved family of proteins in the mechanism of ARE-mediated mRNA decay.

ACKNOWLEDGMENTS

We are grateful to Jack Keene and Roy Parker for helpful discussions and comments on the manuscript and to Sam Johnson and Yasheng Gao at the Duke Light Microscopy Facility for advice. We thank Roy Parker for providing the DCP2-RFP plasmid; Karsten Weis for providing the GFP-NES/NLS plasmid and the mex67-5^ts, xpo1-1^ts, mex67-5/xpo1-1, and rbp1-1^ts mutant strains; Anita Corbett for the YAP1-GFP plasmid and the crm1-2 and crm1-3 mutant strains; and Maurice Swanson for pPAB1-GFP.

S.V.V. is a trainee of the Duke University Program in Genetics and Genomics. This work was supported by NIH grant GM41840 to D.J.T., NIH predoctoral fellowship FDK081304A to S.V.V., and BIO2008-02835 to S.P.