More than one way to make a tail.

The vast majority of eukaryotic messenger mRNAs undergo 30 end cleavage and polyadenylation. The noncanonical poly(A) polymerase (PAP), STAR-PAP, adds a poly(A) tail to specific transcripts under oxidative stress conditions. These transcripts have the AAUAAA polyadenylation signal and use the same factors for cleavage, but are not polyadenylated by the canonical polyA polymerase. In this issue of The EMBO Journal, Leishram and Anderson ask what determinants are required for the modification of specific mRNAs by STAR-PAP. They report a surprising role for STAR-PAP in binding to the pre-mRNA and recruiting the processing factors required for cleavage, which suggests that there is still much to be discovered regarding the regulation of 30end processing. Eukaryotic messenger RNAs, with the exception of animal histone mRNAs, end in a poly(A) tail. The basic set of factors required for polyadenylation in mammalian cells was established by the work of the laboratories of Jim Manley and Walter Keller during the early 1990s (reviewed in Mandel et al, 2007). The polyadenylation signal AAUAAA located 50 of the site of cleavage is recognized by the cleavage polyadenylation specificity factor (CPSF) and the downstream sequence element is recognized by cleavage stimulatory factor (CstF), with cleavage occurring between these two elements catalyzed by CPSF73, a component of CPSF (Mandel et al, 2006). Coincident with cleavage, PAP adds the poly(A) tail to the 30 end. PAP is required in vitro for cleavage of many substrates, guaranteeing coupling of cleavage and polyadenylation. Two years ago, Richard Anderson and coworkers published the first example of a second PAP, termed STAR-PAP, that adds a polyA tail to the 30 end of a specific pre-mRNA, heme oxygenase (HO-1) mRNA (Mellman et al, 2008), when cells are subjected to oxidative stress. STAR-PAP is a non-canonical PAP (Speckle targeted PIPKIa regulated poly A polymerase) that requires PI4,5P2 for activity, linking phosphatidylinositol signalling pathways to RNA processing. Surprisingly, the canonical AAUAAA polyadenylation signal is present in HO-1 mRNA, and the same CPSF complex and other polyadenylation factors are involved in polyadenylation by STAR-PAP. The question then is what determines that an mRNA will be polyadenylated by STAR-PAP and what prevents polyadenylation by the canonical PAP. In this issue, Leishram and Anderson (2010) provide a partial answer to this question by showing that STAR-PAP is required not only for polyadenylation but also for cleavage of HO-1 mRNA. Using in vitro studies, they show that STAR-PAP specifically binds to the 30UTR of HO-1 mRNA 50 of the polyadenylation signal. STAR-PAP also binds to CPSF160 and helps recruit it stably to the HO-1 mRNA polyadenylation signal (Figure 1A). Using recombinant proteins, they then show that three polypeptides, STAR-PAP, CPSF-160 and CPSF-73, are sufficient to cleave HO-1 pre-mRNA in vitro, in a reaction that is dependent on the AAUAAA polyadenylation signal and CPSF-73, the endonuclease that cleaves pre-mRNAs (Mandel et al, 2006). Surprisingly, PI4,5P2 is not required for cleavage although it is required for polyadenylation.