Ccr4 and Pop2 control poly(A) tail length in Saccharomyces cerevisiae

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Abstract

ABSTRACT Messenger RNA degradation is an important aspect of post-transcriptional gene regulation and shortening the poly(A) tail is suggested to be the rate-limiting step in mRNA degradation. In Saccharomyces cerevisiae , the Ccr4-Not complex is the major deadenylase and contains two subunits with exoribonuclease domains, Ccr4 and Pop2. Although the role of Ccr4 and Pop2 in deadenylation has previously been studied using individual reporter mRNAs, their activity has not been studied transcriptome-wide. Here, we describe END-seq, a method to accurately measure poly(A) tail lengths of individual mRNAs transcriptome-wide, and have used this assay to examine the impact of deleting or mutating CCR4 and POP2 on steady state poly(A) tail length. We found that Ccr4 and Pop2 have differential effects on the poly(A) tail lengths of individual mRNAs. Additionally, though Pop2 has previously been reported to have exonuclease activity, mutations that render it catalytically inactive have no effect on steady-state poly(A) tail lengths. Furthermore, mutations that disrupt the interaction between Ccr4 and Pop2 result in longer poly(A) tails. We also observe an inverse correlation between codon optimality and poly(A) tail length – transcripts containing predominantly optimal codons display fewer changes in poly(A) tail length upon deletion of Ccr4 or Pop2 than those containing less optimal codons. Together, these results indicate that Pop2 modulates poly(A) tail length, at least partially, via its association with Ccr4 and that Pop2 improves the function of Ccr4 in regulating poly(A) tail length. These data provide important insights into poly(A) tail length dynamics in yeast and demonstrate that END-seq is an efficient and accurate method to study poly(A) tail length.

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