Star-PAP is a nuclear non-canonical poly(A) polymerase (PAP) that shows specificity toward mRNA targets. Star-PAP activity is stimulated by lipid messenger phosphatidyl inositol 4,5 bisphoshate (PI4,5P2) and is regulated by the associated Type I phosphatidylinositol-4-phosphate 5-kinase that synthesizes PI4,5P2 as well as protein kinases. These associated kinases act as coactivators of Star-PAP that regulates its activity and specificity toward mRNAs, yet the mechanism of control of these interactions are not defined. We identified a phosphorylated residue (serine 6, S6) on Star-PAP in the zinc finger region, the domain required for PIPKIα interaction. We show that S6 is phosphorylated by CKIα within the nucleus which is required for Star-PAP nuclear retention and interaction with PIPKIα. Unlike the CKIα mediated phosphorylation at the catalytic domain, Star-PAP S6 phosphorylation is insensitive to oxidative stress suggesting a signal mediated regulation of CKIα activity. S6 phosphorylation together with coactivator PIPKIα controlled select subset of Star-PAP target messages by regulating Star-PAP-mRNA association. Our results establish a novel role for phosphorylation in determining Star-PAP target mRNA specificity and regulation of 3′-end processing.

Highlights d RBM10 interacts with Star-PAP and directs Star-PAP activity toward polyadenylation d RBM10 is enriched in the heart and controls expression of anti-hypertrophy regulators d RBM10 depletion or compromised Star-PAP interaction results in myocyte hypertrophy d RBM10 expression is lost in the in vivo model of hypertrophic heart in Wistar rats

Alternative polyadenylation (APA)-mediated 3′-untranslated region (UTR) shortening is known to increase protein expression due to the loss of miRNA regulatory sites. Yet, mRNAs with longer 3′-UTR also show enhanced protein expression. Here, we identify a mechanism by which longer transcripts generated by the distal-most APA site leads to increased protein expression compared to the shorter transcripts and the longer transcripts are positioned to regulate heart failure (HF). A Star-PAP target gene, NQO1 has three poly(A) sites (PA-sites) at the terminal exon on the pre-mRNA. Star-PAP selects the distal-most site that results in the expression of the longest isoform. We show that the NQO1 distal-specific mRNA isoform accounts for the majority of cellular NQO1 protein. Star-PAP control of the distal-specific isoform is stimulated by oxidative stress and the toxin dioxin. The longest NQO1 transcript has increased poly(A) tail (PA-tail) length that accounts for the difference in translation potentials of the three NQO1 isoforms. This mechanism is involved in the regulation of cardiac hypertrophy (CH), an antecedent condition to HF where NQO1 downregulation stems from the loss of the distal-specific transcript. The loss of NQO1 during hypertrophy was rescued by ectopic expression of the distal- but not the proximal- or middle-specific NQO1 mRNA isoforms in the presence of Star-PAP expression, and reverses molecular events of hypertrophy in cardiomyocytes.