CstF-64 and 3′-UTR cis-element determine Star-PAP specificity for target mRNA selection by excluding PAPα

Almost all eukaryotic mRNAs have a poly (A) tail at the 3'-end. Canonical PAPs (PAP alpha/gamma) polyadenylate nuclear pre-mRNAs. The recent identification of the non-canonical Star-PAP revealed specificity of nuclear PAPs for pre-mRNAs, yet the mechanism how Star-PAP selects mRNA targets is still elusive. Moreover, how Star-PAP target mRNAs having canonical AAUAAA signal are not regulated by PAP alpha is unclear. We investigate specificity mechanisms of Star-PAP that selects pre-mRNA targets for polyadenylation. Star-PAP assembles distinct 3'-end processing complex and controls pre-mRNAs independent of PAP alpha. We identified a Star-PAP recognition nucleotide motif and showed that suboptimal DSE on Star-PAP target pre-mRNA 3'-UTRs inhibit CstF-64 binding, thus preventing PAP alpha recruitment onto it. Altering 3'-UTR cis-elements on a Star-PAP target pre-mRNA can switch the regulatory PAP from Star-PAP to PAP alpha. Our results suggest a mechanism of poly (A) site selection that has potential implication on the regulation of alternative polyadenylation.